Stimulus novelty, task relevance and the visual evoked potential in man. Visual evoked potentials (VEPs) were recorded from normal adult subjects performing in a visual discrimination task. Subjects counted the number of presentations of the numeral 4 which was interposed rarely and randomly within a sequence of tachistoscopically flashed background stimuli (numeral 2s). Intrusive, task-irrelevant (not counted) stimuli were also interspersed rarely and randomly in the sequence of 2s; these stimuli were of two types: simples, which were easily recognizable (e.g., geometric figures), and novels, which were completely unrecognizable (i.e., complex, colorful patterns). It was found that the simples and the counted 4s evoked posteriorly distributed P3 waves (latency 380-430 msec) while the irrelevant novels evoked large, frontally distributed P3 waves (latency 360-380 msec). These large, frontal P3 waves to novels were also found to be preceded by large N2 waves (latency 278 msec). These findings indicate that "the P3" wave is not a unitary phenomenon but should be considered in terms of a family of waves, differing in their brain generators and in their psychological correlates. These late positive components are discussed in terms of task-relevance, recognition and Pavlov's "what is it" response.


Sleep studies on a 90-minute day.

Sleep studies on a 90-minute day. After 2 adaptation and 2 baseline all-night sleep recordings, 5 normal young adult subjects (3 males) were placed on a schedule alternating 60 min of wakefulness and 30 min of sleep for 5 1/3 24-h periods. A 2-day recovery period followed. One male subject (MA15) was later placed on the identical protocol with the exception that he was allotted periods of 75 min of wakefulness and 15 min of sleep during the experimental period. One male narcolepsy-cataplexy patient was placed on the 60-30 schedule for 48 h. All subjects showed REM sleep during the schedule manipulation. REMM sleep occurred within 10 min of sleep onset (SOREMP) on 79 of 110 REM sleep occasions in the normals, on all 29 REM episodes in MA15, and on 16 of 17 REM periods in the narcoleptic. In the normals, REM sleep showed a tendency to recur on alternate 90-min cycles, while in the narcoleptic REM recurred on consecutive periods. Compared to baseline, REM sleep 24 h was decreased in the normals and increased in the narcoleptic. Time spent in slow wave sleep and stage 2 was also reduced in the normal subjects on the 90-min schedule, and stage 1 sleep time was increased. Peak sleep times for the 5 normals occurred between 09.00 and 12.30 and lowest sleep times from 21.00 to 02.00. During the first recovery night, sleep times ranged from 11.5 to 18.5 h, including significant increases of slow wave sleep and REM sleep. Except for SOREMPs, no signs of the narcolepsy-cataplexy syndrome were seen in any of the normal subjects.


Electroencephalographic signs of ethanol tolerance and physical dependence in the cat. Spontaneous electrical activity was recorded from three cortical and five subcortical sites, via permanently implanted electrodes, in five conscious, freely-moving adult cats. Initial observations were made during and after an intravenous infusion of ethanol, 1 g/kg. The animals then received ethanol by gastric intubation, in doses of 1.5 g/kg every 8, 12 or 24 h, for a period of 5 weeks. Electrical and behavioral observations were repeated on the day following the last gavage, before and during another intravenous infusion of ethanol 1 g/kg. All animals showed EEG changes which are interpreted as signs of tolerance to and dependence on alcohol. EEG changes appeared up to a day earlier than gross behavioral signs of alcohol withdrawal.


Differential effect of thalamic and subthalamic lesions on early and late components of the somatic evoked potentials in man. The effect of thalamic (Vo), subthalamic (Raprl) and combined (Vo-Raprl) unilateral lesions on early and late components of the somatic evoked potentials (SEPs) was investigated in a group of 16 cases operated on for the treatment of contralateral dyskinetic movements. In these cases, SEPs were independently produced by stimulation of the left and right median nerves and recorded at the corresponding somatosensory scalp regions. In addition, EEG frequencies and reaction time (RT) were independently and bilaterally determined. These tests were performed before and after operation and changes in SEP, EEG and RT were quantitatively evaluated in relation to both ipsilateral preoperative and contralateral postoperative controls. 1. All cases with either thalamic or combined lesions involving Vo nucleus showed an ipsilateral reduction in amplitude of late SEP components and EEG frequency and a contralateral increase RT. A peculiar form of "inattention" to the contralateral hand was also found. Quantitative evaluation of the total group showed significant amplitude reduction in late SEP components in relation to both ipsilateral preoperative and contralateral postoperative controls. Changes in EEG and RT were only significant in relation to their preoperative ipsilateral controls. 2. Two cases with subthalamic lesions and quick postoperative recovery showed no apparent change in SEP, EEG and RT. Three cases with similar lesions and slow postoperative recovery showed bilateral decrease in amplitude of late SEP components, EEG frequency and increase in RT. Clinical inattention to contralateral hand was also found in all these cases. Quantitative evaluation of the total group showed no significant changes in these parameters in relation to preoperative controls. 3. None of these lesions produced changes in early SEP components or somatosensory deficits at the contralateral hand.


EEG changes after withdrawal of medication in epileptic patients. Fifty-five patients with intractable partial seizures whose on-medication EEGs demonstrated either predominantly focal epileptiform lesions or absence of paroxysmal activity, were studied, and the effect of withdrawing all anticonvulsive drugs on their EEGs was observed. Four types of response were encountered: (1) no effect (20%); (2) specific (focal) activation (25%); (3) complex activation (29%) with widespread of the initial on-medication focus or appearance of the additional independent epileptogenic foci; and (4) "non-specific" activation (63%), consisting of bursts of either bilaterally synchronous and frontally dominant spike and waves, triphasic waves, or sharp slow complexes, or smaller amplitude rapid and diffuse spike-and-wave complexes. This latter effect is thought to be secondary to metabolic derangements resulting from the withdrawal of neurotropic agents and not directly related to the specific epileptogenic process. No association was found between type of effect and any of the following parameters: topography of on-medication focus, duration of therapy, type of anticonvulsant used, suspected underlying etiopathology, or median age when medication was withdrawnn. Furthermore no evidence could be found that the development of a "complex" or "non-specific" EEG effect carried with it a bad prognosis for surgical cure following focal cortical excision. Performing off-medication tracings seems to be of greatest value in patients with partial seizures and EEGs revealing either a relative paucity of definite absence of epileptiform discharges. The occurrence of a "non-specific" response in a questionable epileptic during the off-medication period, on the other hand, should be interpreted with caution.


[Rhythmic slow waves and multi-unit activity during sleep-waking cycle in the rat ventral tegmentum]. 1. Multi-unit activity (MUA) was recorded together with gross activity in different structures during various states of sleep in 43 rats. The records were made by means of telemetry from lateral hypothalamus, nucleus ruber, formatio reticularis, dorsal hippocampus and the ventral mesencephalic tegmentum (VMT) surrounding the nucleus interpeduncularis. 2. The records during paradoxical sleep showed slow rhythmic theta waves located in the VMT and the dorsal hippocampus. In these two structures the MUA was mainly organized in periodic patterns of discharges. In the other structures a continuous increment of the basic level of discharge was superimposed on the periodic patterns. 3. The similarity of the hippocampal and VMT records strengthens the hypothesis that the VMT region has anatomical and functional connections with the limbic system.


A device for the maintenance of a constant body temperature, with some special features. A device for the maintenance of constant body temperature of laboratory animals during experiments is presented. Some special features make it especially suitable for experiments in which small electrical events are to be recorded, because no 50 or 60 c/sec interference or spurious signals will occur from it, nor is the animal earthed through the apparatus. In 2 years of daily use it has proved to be a reliable and accurate working device.


Distortion of averaged post-auricular muscle responses due to system bandwidth limits. Substantially different waveform details have been reported in various studies of the post-auricular muscle (PAM) response. By examining the high frequency limits of the pass-bands used to record the response it would appear that the system bandwidth limits have affected the waveform. Recordings were made using a very wide bandwidth, and were used to obtain power spectra of these PAM responses. These determined the minimum bandwidth requirements to avoid distortion of the waveform. It seems clear that a substantial proportion of the differences reported have been due to the use of a recording bandwidth which has distorted the response.


Chloralose induced alteration of visually evoked response from specific and non-specific regions of cat neocortex. Visually evoked response (VER) and EEG from the motor cortex (precruciate gyrus) and the visual cortex (marginal gyrus) of cats were recorded from 4 to 7 h after the injection of anesthetic doses of alpha-chloralose. During the recording period the VER from the precruciat gyrus showed a 200-300% increase in amplitude while the VER from the marginal gyrus rarely varied more than 50% in amplitude, and did so independent of the changes in the VER from the precruciate gyrus. The number of large amplitude spikes in the EEG from the precruciate gyrus also increased dramatically during the recording period, but no definite correlation between changes in VER amplitude and in the number of spikes in the EEG could be demonstrated. These observations suggest a functional separation between specific and nonspecific sensory pathways, with the latter showing a considerably greater sensitivity to level of anesthesia.


REM sleep and contingent negative variation development. Three subjects demonstrated the ability to respond in a contingent negative variation (CNV) paradigm during rapid eye movement (REM) sleep. However, the CNV did not appear as it does in the waking state. This failure of the CNV to develop during REM sleep might be attributed to electrophysiological changes accompanying REM sleep.


Analysis of motor conduction velocity in the human median nerve by computer simulation of compound muscle action potentials. A digital computer was used to reconstruct compound muscle action potentials recorded from the human thenar eminence after stimulation of the median nerve. The programme allowed the following parameters to be varied: (1) the dimensions of a representative single motor unit potential; (2) the number of motor units in the muscle and the range and distribution of conduction velocities in their nerve fibres; and (3) the distance along the nerve from the point of stimulation to the muscle. The reconstructed compound muscle action potentials were similar to real compound potentials recorded from normal subjects. The number of single motor units and the range of conduction velocities required for the reconstruction correlated with quantitative histologic studies of the recurrent branch of the median nerve to the thenar muscles. By altering the distribution of conduction velocities it was possible to study the effect of abnormal patterns of nerve conduction on the configuration of the simulated compound muscle action potentials. It was found that abnormally slow conduction caused an increased discrepancy between the main parameters of compound potentials corresponding to stimulation of the nerve at proximal and distal sites. These observations suggest that a careful analysis of the differences between pairs of compound muscle action potentials may provide a method for more detailed assessment of conduction velocity in clinical studies of peripheral nerve disorders.


Effects of time and tasks upon auditory and somatosensory evoked potentials in man. Auditory evoked potentials (AEPs) recorded from the vertex and somatosensory evoked potentials (SEPs) recorded from the somatosensory and motor areas and vertex were examined during visual and auditory discrimination tasks, with and without motor responses, and during motor tasks alone in normal human subjects. These procedures allowed the separation of sensory discrimination from motor activity effects on the evoked potentials. It was observed that EPs were modified systematically by task and by temporal variables, even though vigilance, as evaluated through EEG recording and performance level, was stable throughout the experimental recording session. 1. AEPs were minimally influenced by time, but very sensitive to task. Inversely, the SEP amplitude decreased considerably with time and less with task. There was evidence of time/task interaction. 2. The magnitude of SEP attenuation in time was reduced by intervening rest periods. 3. The sensory modality in which the discrimination task was performed did not influence the effect on EPs. A discrimination task involving a motor response reduced EPs more than a pure discrimination or a pure motor task. The task effect seems to involve a general, mechanism (load imposed upon the subject) not dependent on the particular sensory channel used to deliver task-relevant information.


Effect of voluntary self-paced movements upon auditory and somatosensory evoked potentials in man. The effect of voluntary self-paced movements upon auditory (AEPs) and somatosensory (SEPs) evoked potentials has been investigated according to the temporal relationship between movement and delivery of test stimuli. EPs were recorded in 7 subjects and averaged in 10 successive epochs extending from 880 msec before to 2500 msec after movement. AEPs were attenuated in all epochs. The decrease was greatest in the 220 msec epoch just following movement and involved components N85 and P170. SEPs were attenuated similarly to AEPs when movements were performed by the hand contralateral to somatosensory stimulation. Of the 5 SEP components, only P40 failed to reflect the attenuation, while P95 showed the greatest amplitude decrease. When stimulation was ipsilateral, SEP amplitude was attenuated only when close to the movement. N65 and P95 decreased while N130 increased. In all subjects the results were consistent for treatments of AEP and SEP (with contralateral movements), whereas large inter-individual differences were observed for the SEP with ipsilateral movements.


Correlation analysis of EEG changes during recognition of images of objects. This work reports the results of both psychological and electrophysiological experiments. It was carried out on 15 normal subjects by the method of cross-correlation analysis of EEGs recorded during recognition of tachistoscopically presented images of objects. Changes in the random component Cr of the cross-correlation were larger during recognition than during simple mobilization of the subject's attention.


Post-activation potentiation and the kindling phenomenon. Potentials evoked in the hippocampus and preoptic region of rats by single biphasic pulses applied to the amygdala were compared during recruiting, after post-tetanic potentiation (PTP) and after amygdaloid kindling. The same components were enhanced temporarily by recruiting and PTP as were enhanced permanently by kindling. Trains of tetanic stimulation with parameters which partially mimicked the cellular discharge parameters during an amygdaloid afterdischarge (AD) were applied to the amygdala at a frequency of 1 per 5 sec continuously for 2.5 h or for 15 min a day for 10 days to produce a total of 1800 trains. Amygdaloid kindling rates were then measured and compared with control groups. Animals pretreated with tetanic stimulation required significantly fewer ADs to develop maximal seizures. Further experiments showed that tetanic stimulation, but not recruiting stimulation (10 c/sec), low frequency stimulation (1 c/sec), or handling, would produce a permanent change in potentials evoked in secondary sites by single pulses applied to the amygdala. This change in evoked potential amplitude was significant but smaller than that produced by kindling. Also tetanic stimulation, but not recruiting or single pulses, facilitated subsequent kindling.


The number and duration of the episodes of the various EEG stages of sleep in young and older people. Episodes of any one electrophysiological sleep stage and of intervening wakefulness were examined as to their numbers and durations in sleep of ten young normal subjects (mean age 22) and in significantly more disturbed sleep of fourteen late middle age normal people (mean age 55). No significant difference between the two age groups was found for any of the stages in the mean number of episodes taken from 4 nights. For any of the stages, the individual numbers of episodes accumulated over the 4 nights varied in a wide range and showed significant consistency with similar data obtained in another set of 4 nights. There were significant differences between the two age groups in episode duration. The sleep of the older group contained significantly higher proportions of longer episodes of wakefulness and drowsiness and significantly lower proportions of longer episodes of slow wave sleep and REM sleep. Thus, the initiation of episodes of EEG stages was on average similar in the better sleep of young and impaired sleep of older subjects, but the maintaining or termination of the episodes were different.


On hemispheric differences in evoked potentials to speech stimuli. Eight subjects listened to lists of speech sounds (pa or ba) or pure tones (250 or 600 c/sec). Within each list one of the sounds (the "frequent") occurred more often than the other (the "target") in a ratio of approximately 4:1. Subjects were required to count the targets in each list; concurrently, evoked responses to both targets and frequents were being separately averaged from electrodes at vertex at symmetrical left and right parietal locations. The evoked responses show the expected sequence of deflections at all three electrode sites, including large P3 waves (about 350 msec latency) to the target stimuli. However, the left and right hemispheric responses to speech or tones, either frequent or target, were strikingly similar, both to the eye and by statistical tests intended to reveal differences between them.


Two-dimensional eye movement monitor for clinical and laboratory recordings. A photo-electric device designed to monitor simultaneously vertical and horizontal eye movements within a 20 degrees range is presented with illustrating experimental data. Four small infrared detecting cells are mounted on a light spectacle-like frame together with a miniature true infrared (9000 A) emitting diode. This original design which eliminates separate source illumination artifacts is extremely light, preserves maximum vision field size, and has particularly straightforward operation. The instrument resolution is less than 1 minute of arc with a 1000 c/sec bandwidth and a 5% linearity over the maximum operating range.


Modification of the direct cortical response by taurine. The effects of taurine on the direct cortical response have been studied in immobilized cats (with local analgesia). The primary negative component of the response was inverted in polarity by the topical application of 25 mM taurine while the later slow negative component was considerably augmented. These effects are identical to those observed with 25 mM GABA. Pentobarbital anesthesia produced little qualitative change in the effects of taurine. With stronger stimuli in the presence of taurine, rhythmic waves followed each stimulus both at the cortical surface and at a depth of 1000 mum. An increase in the EEG amplitude following topical taurine was generally localized to the cortical surface but in certain experiments, could be recorded at cortical depths up to 1000 mum.


The feedback effects of sex steroid hormones on pituitary gonadotropin release in Turner's syndrome and Klinefelter's syndrome. The present study was designed to elucidate the feedback relationship between the release of pituitary gonadotropins and sex steroid hormones in Turner's syndrome and Klinefelter's syndrome. LH-RH stimulation test was employed to evaluate the effects of sex steroids on the release of gonadotropins. The release of gonadotropins in response to LH-RH as well as in baseline level was suppressed after the treatment with estrogen (mestranol 0.08 mg/day) for 10 days, followed by the treatment of the same period with estrogen (mestranol 0.08 mg/day) and progesterone (chlormadinone acetate 2.0 mg/day) in combination in both syndromes. The inhibitory effect of the combined treatment was greater than that of the treatment with estrogen alone. Administration of testosterone propionate (25 mg/day) for 3 days resulted in suppression of the release of both gonadotropins in baseline level and in response to LH-RH in both syndromes, but the suppressive effect appeared to be less complete as compared with that of estrogen or estrogen-progesterone. It was thus verified that the feedback interaction between the pituitary gonadotropin release and sex steroids such as estrogen, estrogen-progesterone or testosterone was operative in the same fashion in the patients with Turner's syndrome and Klinefelter's syndrome.


[Cytological and immunocytochemical studies of the anterior pituitary gland of the beagle bitch during various reproduction phases]. The changes in anterior pituitary (AP) of pregnant and lactating dogs as compared with pituitary of animals in metestrus-anestrus phase are described with special reference to the relative proportion, topography and morphology of prolactin cells, somatotrophs, corticotrophs, thyrotrophs and gonadotrophs. For demonstration of these cells, suitable histological, histochemical and immunoenzyme cytochemical methods are used. The prolactin cells show progressive hyperplasia during pregnancy, so that at the end of this phase and during lactation, they comprise the most predominant glandular cells in AP. At the same time, they reveal massive hypertrophy with marked morphological features of high secretory activity, After transient or continous interruption of the suckling stimulus they show signs of functional inhibition on involution. The corticotrophs appear at 20. and 30. days of pregnancy to be relatively increased in number. While in the last third of pregnancy and during lactation, they only seem to be more active than those in pituitary of metestrus-anestrus dogs. The somatotrophs appear to be progressively reduced in relative number during pregnancy and lactation. However, they show some morphological signs of active secretion. The thyrotrophs did not show any morphological alterations during the different reproductive phases. The gonadotrophs reveal during pregnancy, especially at 30. day morphological signs of stimulation. On the contrary they appear atrophied during lactation. This may be a result of suckling stimulus and morphological expression of the inverse relationship in the secretion pattern of gonadotrophins and prolactin in dogs during suckling. The estrogen and progesterone levels in plasma as well as the changes in their relative concentrations may largely account for the structural changes on AP of pregnant dogs. However, neuroendocrine reflexes (e.g. suckling stimulus) seem to be of a great importance for the maintainance of stimulation of prolactin cells during lactation.


[Enzyme histochemical studies on the rat adrenal cortex, ovary, uterus and vagina following chlormadinone acetate administration, especially cholinesterase activity in myometrium and endometrium]. 1. Female albino rats were treated with a total of 28 mg of chlormadinone acetate (CMA) for 28 days. In the adrenal cortex, the ovary, the vagina, and the uterus the activities of 3-beta-ol-steroiddehydrogenase, of dl-beta-OH-butric acid dehydrogenase, of alcaline and acid phosphatases, of DPN-diaphorase, of ATP-ase, and of non-specific esterases do not differ from untreated controls. 2. In the external muscle layer of the myometrium strong cholinesterase (ChE) activity was induced by C.M.A. A corresponding high ChE activity is normally found in immature rats or in estrus. 3. Furthermore, by treatment with CMA, ChE activity was induced in the tubular glands of the endometrium. This activity is found in the small parts of glomerate glandular terminals only but not in the rest of the glandular epithelium, nor in the epithelium of the cavum. It could be demonstrated that a corresponding ChE activity normally appears in the second third of pregnancy. The ChE activity induced by CMA was considerably higher and more widespread than during normal pregnancy. 4. It is concluded that in the endometrial glands a development similar to pregnancy is initiated by CMA. But development stops at the stage of ChE activity, thus leading to accumulation of ChE active cells.


Interactions of antiepileptic drugs.

Interactions of antiepileptic drugs. Several interactions involving antiepileptic drugs are based on changes in the rate of their metabolism and elimination, with concomitant rise or fall of plasma levels. Thus, phenobarbital generally induces the production of the DPH metabolizing enzyme, but its presence inhibits the action of that enzyme. The net result depends upon the balance between these factors in individual patients. Either a decline, a rise, or no change of the DPH plasma level may occur after the onset of administration of phenobarbital. Drugs that may cause elevation of the DPH plasma level include disulfiram, sulthiame, bishydroxycoumarin, chloramphenicol, phenyramidol, benzodiazepines, sulfamethizole, and isoniazid. Isoniazid has been shown experimentally to be a strong inhibitor of DPH metabolism. The extent of DPH plasma level elevation by INH is related to the genetic make-up of individual patients. The highest and frequently toxic DPH plasma levels were seen in very slow INH inactivators. The incidence of clinically significant interactions is not high with most drug combinations; marked changes of antiepileptic drug levels occur only in apparently susceptible individuals. The effects of interactions are not necessarily detrimental; elevation of a low ineffective level may improve seizure control. A rise to a toxic level range requires reduction of the dose of primary drug or elimination of interfering drugs. Monitoring the blood levels of anti-epileptic drugs provides the best means to anticipate interactions and to regulate the doses when multiple medications have to be used.


Antigenic determinants in the disulfide regions of bovine fibrinogen. Rabbit antibodies to bovine fibrinogen were used to study the antigenic activity of four cyanogen bromide peptides containing the disulfide regions of this molecule. In precipitation tests the highest activity was associated with the peptide F-CB3 which is exclusively derived from the alpha-chain. Reduction of the single disulfide bridge in peptide F-CB3 did not influence its serologic activity. Weaker reactions were observed with the N-terminal multichain peptide F-CB1. The antigenicity of peptide F-CB1 was not affected by removal of fibrinopeptides A and B but it was lost after reduction. The immunological activity of the multichain peptide F-CB2 was even less than that of peptide F-CB1 and the antigenic determinants were destroyed by reduction. A large fragment essentially composed of peptides F-CB1 and F-CB2 could be obtained by limited cyanogen bromide cleavage and showed considerably better immunological activity than peptides F-CB1 and F-CB2 together. Apparently no activity was associated with a mixture of small hydrophobic, disulfide-loop peptides tentatively called peptide F-CB4. The large loss of antigenic activity in cyanogen bromide digests of fibrinogen suggests that the disulfide-stabilized regions do not have an important role in maintaining conformational antigenic determinants of fibroinogen. Changes in noncovalently stabilized conformation requiring uncleaved chains is considered as a possible reason for the findings observed.


Barrier impairment and immune reaction in the cerebrospinal fluid. Investigations on pathophysiological processes in the CSF space due to impairment of the blood-CSF barrier and due to an immune reaction are reported. Barrier disturbances and immune reaction are demonstrated by means of absolute values of CSF protein electrophoresis and by measurement of immunoglobulins. With inflammatory diseases of the CNS, an altered blood-CSF barrier will often be seen at the moment of the first lumbar puncture. Multiple sclerosis, subacute leucoencephalitis and neurosyphilis show signs of a distinct immune reaction together with an increase of the gamma-globulins as well as the immunoglobulin G in the absence or slight presence of a barrier impairment. Non-inflammatory diseases of the CNS have variable barrier disturbances.


Alpha subunit of glycoprotein hormone: presence in peripheral serum after LHRH. A method is described for the selective measurement of human luteinizing hormone (hLH) and alpha subunit. The assays employ highly purified tracers of hLH and of subunit of human chorionic gonadotropin (hCGalpha) and a "mixed population" of antibody: Population I (directed against determinants on beta subunit of hCG) and Population II (directed against determinants on alpha subunit of hCG). The former is present in greater concentration than the latter. When the mixed antibody is used at higher dilution (1:1.2x10(6)), Population II is effectively diluted out, and using 125I-labelled hLH as tracer, the assay recognize hLH, hCG and their beta subunits 20-50 times more sensitively than hCGalpha. When the mixed antibody is used at fivefold higher concentration, Population I is present in relative excess and acts as a "sink" for hCG, hLH and their beta subunits. Under these conditions, using 125I-hCGalpha as tracer, the assay recognize the alpha subunit 20 to 50-fold more sensitively than hLH and hCG. These assays have been employed in the study of sera which have been filtered over Sephadex G-100. Alpha subunit was detected in serum within minutes after intravenous injection of luteinizing hormone releasing hormone (LHRH) in four subjects tested.


Granulocyte collagenase, elastase and plasma protease inhibitors in purulent sputum. Sputum was collected from patients with purulent chronic bronchitis. Immuno-chemical techniques using rabbit antiserum against human granulocyte collagenase and elastase showed the presence of both enzymes. Also the serum protease ingibitors alpha1-antitrypsin and alpha2-macroglobulin were demonstrated. Their protease inhibiting capacity was saturated. Granulocyte elastase and collagenase occurred not only in complexes with the inhibitors, but also as free enzymes. All sputa showed free proteolytic, elastolytic and collagenolytic activity. The concentration of collagenase was equal in the sol phase and in the gel phase of the sputa, but most of the elastase was bound to the gel phase.


A comparison of subjective measurements of human sperm motility and viability with two live-dead staining techniques. Two commonly used live-dead stains (eosin-nigrosin (EN) and eosin-opal blue (EOB)) were compared with the estimated active spermatozoa in semen samples from patients attending an infertility service. Twenty-eight semen samples were analyzed throughout the day of their collection by estimating the number of active spermatozoa and by staining a portion of the incubated sample (37 degrees C) with each stain. The samples were analyzed 30, 60, 120, 240, and 360 minutes after the initial collection. At 30 minutes there were no significant differences between the estimated values and those of either stain. The slope of the EN stain closely paralleled that of the estimated measurements throughout the remainder of the time periods, while the EOB slope was somewhat steeper. Repeatability of values for semen samples obtained on different days was generally good for each stain. A comparison of semen samples from 85 patients, 30 minutes after collection, showed no significant differences between the numbers of estimated active sperm and the percentage live using the EN stain. Studies of 132 semen samples using the EN stain showed a positive correlation with the over-all quality of sperm motility. The results indicate that there is a role for live-dead staining in assessing semen quality.


Bleomycin therapy in mycosis fungoides.

Bleomycin therapy in mycosis fungoides. Nine patients with mycosis fungoides in different stages were treated with Bleomycin. Much better results were obtained with this new drug in the six patients with the infiltrative or beginning tumour stage than in the patients in the advanced tumour stage. Complete remission was not seen. In one case the results were objectivized by DNA cytophotometry. The role of Bleomycin in the treatment of mycosis fungoides is discussed. It is concluded that Bleomycin is not the medicament of choice for the treatment of mycosis fungoides.


Heterogeneity of allergenic extracts and of the antibody response in allergic patients. An attempt to provide an overview of intrinsic problems encountered in the definition of the immunochemical parameters involved in the allergic response in terms of the appropriate in vivo and in vitro manifestations. The primary difficulty stems from the complexity of the chemical composition of most extracts of allergenic materials and from the lack of simple laboratory methods for establishing the allergenic profile of each patient as a function of the different components of a given extract. The principles of inhibition of the Prausnitz-Küstner passive transfer test and of the radioallergosorbent technique will be described; from the results one may deduce the minimum number of distinct allergenic components and of the corresponding reaginic antibodies of IgE type. The possible use of monovalent, hapten-like, low molecular weight constituents of allergens for blocking the triggering of the release of vasoactive amines from mast cells and basophils of allergic patients will be discussed. In addition of IgE antibodies, the spectrum of antibodies produced by the allergic patient, both before and after hyposensitization, involves immunoglobulins of the IgG, IgM and IgA classes, which can be measured by radioimmunoelectrophoretic and -immunodiffusion techniques; IgM antibodies can be readily differentiated from IgG antibodies by the hemagglutination technique, or by adaptation of reverse immunosorbent procedures. Similarly, the heterogeneity in relative affinities of the different antibodies for a given allergenic fraction may be established by the use of immunosorbents. The effectiveness of hyposensitization therapy could be thus correlated with objectively measurable immunological parameters. Possible mechanisms responsible for the effectiveness of this treatment will be discussed in the light of recent experiments in animal systems, suggesting the mandatory cooperation between T and B cells for the induction of the reaginic response.


Problems raised by potency evaluation of allergen preparations. Wide variations are observed in the potency of allergen preparations available for use on humans. There are numerous methods of in vivo or in vitro testing: skin testing, passive anaphylaxis testing on animals, study of precipitins, histamine release tests on human leukocytes, radio-immunological assay test (RAST), blasticlymphocyte transformation test (TTB). The first investigations seemed to show that the allergenic potency of a large number of inducing substances was due to proteic activity; this gave rise to the theory that potency could be evaluated by protein nitrogen units (PNU). However, the protein content of an allergenic extract is not always active. In addition, natural specific proteins are altered during the extraction process or subsequently during preservation. Where the active antigen is known, comparison of different testing with various preparations may be made. The results show excellent correlation between the two biological results (cutaneous reaction and histamine release) and the content of E antigen. No relation with PNU has been observed. Certain authors propose the following: skin testing, PNU titration, dosage of certain elements (polysaccharides, lipids, phosphates, etc.), RAST. The great differences observed between the value of skin testing and the RAST may be due to the large number of active antigens in different 'dusts' and to the fact that the RAST method measures only IgE whereas allergic activity of these allergens is no doubt associated with other types of antibodies. The complexity of the problem shows that a great many studies are still necessary to provide a satisfactory solution.


Preliminary characterization of a major allergen of timothy grass pollen. Previous studies with timothy pollen extracts demonstrated that a low molecular weight dialyzable fraction, antigen D, possessed the allergenic determinant of the major allergen of timothy pollen (allergen B). Bio-gel P-2 gel-filtration of the dialysate fraction resulted in the isolation of a fragment, antigen D3 with a molecular weight near 1000. The relationship of antigen D3 to the antigenic and allergenic determinants of allergen B was evaluated by passive cutaneous anaphylaxis (PCA) in guinea pigs sensitized with rabbit antitimothy sera and by allergen-induced histamine release from monkey lung tissue passively sensitized with the serum of timothy-sensitive patients. These studies demonstrated that 10 units of antigen D3 gave 1) a 70% inhibition of PCA reactions in guinea pigs challenged with allergen B, and 2) a 73% inhibition of allergen-induced histamine release from monkey lung tissue sensitized with timothy reagin and challenged with the crude pollen extract (WST). Although chemical characterization studies of the antigen D3 fragment are still being carried out it appears that the major components fo this fragment are 1) a flavonoid pigment - quercitin, 2) a disaccharide moiety - cellobiose, and 3) the amino acid threonine linked together in an )O-glycosidic type linkage.


Standardization of allergens by an improved skin-testing technique. From the seemingly most appropriate methods for standardization procedures of allergen extracts, the most serious candidates are in sequence of age: the old atopic skin test, the histamine-release test, using human basophil cells and the radio-allergosorbent test. At the moment it is not yet certain which method gives the most exact results, although, especially in the last ones - as part of the whole test - highly sophisticated methods are used. The skin test is not dependent on modern laboratory methods, is used on a world-wide scale, but as method studied very poorly. This is a matter of regret, because at any rate the skin test gives the clinician a lot of valuable information. The major drawbacks of the skin test are that the inclination of the log-dose response line is rather small and that the results of tests done with the same dose may vary sometimes not inconsiderably. Furthermore, the results are dependent on the sites of the skin where the tests are done and on the amount of fluid injected intracutaneously. There is a diurnal variability, a certain influence of the menstrual cycle and of course influence of medicaments (histamines). Nevertheless, taking all these factors into consideration and using a highly perfected method of skin testing, it appeared possible in 16 experiments, with not more than six volunteers per experiment, to reduce the variability of the bioassay between the limits of +50 and minus 50 per cent.


Chemistry of a low molecular weight ragweed pollen allergen RA5, and association of human responsiveness to RA5 with HL-A7 cross-reacting group. A protein allergen, Ra5, was isolated by column chromatographic procedures from the aqueous extract of short ragweed pollen by Lapkoff and Goodfriend. The allergen was obtained in a highly homogenous form, was devoid of carbohydrate and had the low molecular weight of 5100 daltons. A report will be given of studies by Marsh, Bias, Hsu and Goodfriend on the cutaneous activity of Ra5 in 105 antigen E-sensitive patients along with HL-A typing of these patients. The results have demonstrated a highly significant association between sensitivity to Ra5 and possession of a histocompatibility antigen of the HL-A7 cross-reacting group (creg). By analogy with studies in inbred strains of mice, this finding suggests control of human sensitivity to allergens by immune response genes. A brief account will also be given of studies on the primary structure of allergen Ra5 by Mole, Goodfriend, Lapkoff, Kehoe and Capra which have to date yielded the complete amino acid sequence of this protein.


Optimum fractionation regimen for bleomycin treatment. A theoretical analysis was made on the optimum Bleomycin treatment regimen on the basis of the "binding-saturation model" which was proposed for the Bleomycin dose-cell survival relation. The surviving fraction of tumor cells decreased as a function of the number of fractionated treatments up to the optimum fractionation number if the tumor was treated with the same total dose. The effect of cellular sensitivity to the antibiotic, tumor doubling time, treatment interval, and total doses on the optimum regimen was analyzed. The importance of treatment interval and tumor doubling time was emphasized and the short treatment interval was recommended for the clinical use of this antibiotic. The optimum number of fractions increased linearly with the increase of the total dose while the optimum single dose was independent of the total dose. A concept of the tumor control probability of tumors treated with the optimum fractionation regimen was introduced and implications of these analyses in the clinical cancer chemotherapy were discussed.


Effect of various factors on induction of liver tumors in animals by the alpha-isomer of benzene hexachloride. The tumorigenic effect of a diet containing the alpha-isomer of benzene hexachloride (alpha-BHC) on the liver of various animals was examined. It was found that alpha-BHC induced liver tumors in male and female mice but not in rats or hamsters in the present observations. Histological changes in the liver of mice induced by alpha-BHC were also much greater than those induced in rats or hamsters. Male animals were more susceptible to the tumorigenic action of alpha-BHC than females. Among different strains of mice, the DDY showed greatest susceptibility and the C57BL/6 showed the least. Induction oflpha-BHC was not inhibited by concomitant feeding of 1-naphthyl isothiocyanate or p- hydroxypropiophenone. However, 3-methylcholanthrene slightly inhibited their induction by alpha-BHC.


alpha-Fetoprotein detected in rat transplantable hepatomas by radioimmunoassay. Production of rat alpha-fetoprotein by transplantable ascites hepatoma was studied by radioimmunoassay method. alpha-Fetoprotein was detected in sera from rats bearing 22 out of 50 ascites hepatoma cell lines that were previously negative for the presence of alpha-fetoprotein by the Ouchterlony test. alpha-Fetoprotein was also detected with high prevalency in Morris hepatomas and sublines of Yoshida sarcoma by this assay system.


Intramural injection of ethanol under direct vision for the treatment of protuberant lesions of the stomach. The experimental production of gastric ulcers in dogs by subnucosal injection of ulcerogeic agents is described. The most satisfactory agent tested was 95% ethanol, because it produced immediate changes in the mucosa which delineated the size of the ulcer. The depth of ulceration could be controlled and the ulcer healed readily. Using this technique of ulcer formation, treatment of protuberant gastric lesions was attempted. Eradication of the pedunculated and narrow-based polyps in stomach was almost totally successful by injection into the base. Wide-based lesions, including atypical hyperplasia and the elevated type of early gastric cancer, were also completely eradicated in 96% of the treated cases. There have been no complications. An initial assessment of the practicality of this method in poor risk patients is presented.


Clinical observations during a relatively early stage of hepatocellular carcinoma, with special reference to serum alpha-fetoprotein levels. Five cases of hepatocellular carcinoma in whom diagnosis was made when the tumor was relatively small, are described. In 2 cases, serum alpha-fetoprotein (AFP) strted to rise sharply, which enabled early detection and surgical removal of the tumor. Serum AFP was below 100 ng per ml, but above the upper normal limit by radioimmunoassay, and was unfluctuating for a considerable period of time before it began to rise in 2 cases. It was negative throughout in 1 case, who lived more than 4 years after the tumor had reached a detectable size. In 4 of 5 cases, the tumor seemed to have evolved during a stage of chronic hepatitis or its transition to cirrhosis. In 1 case with chronic schistosomiasis and advanced mixed macro- and micronodular cirrhosis, a 1.5-cm tumor was detected by celiac angiography. These observations on time relationship of oncogenesis may be generalized to modify the cirrhotic liver. Necessity is emphasized for the early detection of this type of carcinoma to monitor serum AFP in chronic hepatitis patients, particularly in those with unfluctuating, mildly abnormal levels of AFP.


Formal genetics of the HL-A region.

Formal genetics of the HL-A region. The extreme polymorphism of the HL-A system is due to the presence of two (SD1, SD2) and perhaps three linked polyallelic genes. The distinction of "bridging antibodies" (reacting with several HL-A specificities recognizing separate sites on the HL-A molecule) from the main HL-A determinant as it is demonstrated by absorption/inhibition experiments increases this complexity. The HL-A linkage group is composed of other systems: LD1, LD2, PGM3, ADA (?), P, ME1, IPO-B and possibly a "hay fever gene". No gametic or zygotic selection was found in spite of the presence of HL-A antigens on spermatozoa. Mixed lymphocyte reaction (MLR) is principally governed by LD genes. The main (LD2) gene is probably situated outside the interval SD1, SD2, near SD2. Other LD genes (LD1 inside the interval SD1-SD2 and LD3) are suspected. The presence of an immune response gene (Ir) has not yet been demonstrated although several diseases associated with specific SD2 antigens are known. These different genes (SD1, SD2, LD1, LD2, LD3 and Ir) probably form a functional unit in the allo-immunozation.


Antibodies to double-stranded (native) DNA in active chronic hepatitis. Sera from 36 patients with active chronic hepatitis were studied for the presence of antibodies to double-stranded (native) DNA. These antibodies are a specific antinuclear antibody previously shown to have a high degree of specificity for systemic lupus erythematosus. Fifteen of the 36 patients (42%) were found to have levels of antibody usually only reported in systemic lupus erythematosus and higher than those seen in a control population. Anti-DNA antibodies were not found in a group of 22 patients with other forms of liver disease.


Tumour immunology and the gut.

Tumour immunology and the gut. This review set out to answer several questions related to tumour immunology and the gut. It is evident that in patients with gastrointestinal cancer there is a general depression of the immune response and this seems to be correlated with the stage of the disease. Paradoxically a specific immune response against definable tumour antigens can be demonstrated, both cellular and humoral mechanisms being involved although the complexities of this paradox require further analysis. Immunotherapy has been employed in gastrointestinal tumours in a sporadic way. The results suggest that gastrointestinal neoplasms may respond at least as well as other tumours. A firm conclusion awaits the results of controlled trials in which the bulk of the tumour has been effectively dealt with by other means or where combined immunochemotherapy is being used.


New aspects of radioimmunochemical measurement of human parathyroid hormone using the labelled antibody technique. Two forms of heterogeneity of parathyroid hormone (PTH) have given rise to conflicting results: one due to the heterogeneity of the secreted species from the gland and their peripheral metabolism and the other representing the immunochemical heterogeneity of the available antibodies. We have developed sequence specific assays using the technique of labelled antibodies. Therefore, results of assays measuring the C-terminal part and the (1-34)-N-terminal part of the molecule could be compared to those of an assay for hormone bearing both N- and C-terminal antigenic determinants. This assay is supposed to detect predominantly (1-84)-intact hormone. The immunoradiometric assay of (1-34)-PTH has a sensitivity of 0.04 ng/ml. This technique avoids the critical iodination of the hormone fragment containing no tyrosine. There is the expected overlap between normal subjects and patients with primary and secondary hyperparathyroidism. The most important finding are results from patients undergoing neck catheterization. We demonstrated nonuniform secretion of different species of PTH by parathyroid adenomata and normal glands. This supports the hypothesis of cleavage of the (1-84)-molecule in the gland.


[On the role of pentachlorocyclohexene in the metabolism and action of hexachlorocyclohexane. I. Synthesis of beta-pentachlorocyclohexene and its identification as the monodehydrochlorination product of alpha-hexachlorocyclohexane (author's transl)]. It is the aim of a series of investigations to test whether or not beta-pentachloro-1-cyclohexene is an intermediate in the biodegradation of alpha-hexachlorocyclohexane. This paper describes attempts to synthesize this intermediate by chemical methods. 1) Pentachlorocyclohexene was synthesized by partial additive chlorination of chlorobenzene. Combined gas chromatography-mass spectrometry revealed that at least five different isomers of pentachlorocyclohexene had been formed. 2) Treatment of alpha-hexachlorocyclohexane with alkaline buffer (pH 8) produced trichlorobenzenes and, in small yield (4%), a pentachlorocyclohexene. This was isolated and identified as the beta-isomer by melting point (71.8 - 72.6 degrees C, uncorr.), IR- and mass spectrum. Dehydrochlorination of beta-pentachlorocyclohexene produced the trichlorobenzene isomers in a pattern which is characteristic of alpha-hexachlorocyclohexane. The position of the chlorine substituents in the beta-pentachlorocyclohexene molecule as judged from NMR studies is e-aeee. This confirms that it is the monodehydrochlorination product of alpha-hexachlorocyclohexane. The configurations of gamma- and delta-pentachlorocyclohexene, determined for comparison, are e-eeaa and e-eeee, respectively. The kinetics of dehydrochlorination of both alpha-hexachlorocyclohexane and beta-pentachlorocyclohexene in alkaline acetone/water (3 + 2) was studied by means of conductometry. Both reactions are of second order:  kappa alpha-HCH 0.0495 [1 times mol- minus 1 times s- minus 1[; kappa beta-PCH 0.905 [1 times mol- minus 1 times s- minus 1] (3.6 degrees C). 3) Dehydrochlorination of alpha-hexachlorocyclohexane in pyridine/xylene (3 + 4) was also studied. An earlier report claiming that gamma-pentachlorocyclohexene (and not the beta isomer) is produced in this medium was confirmed, if the reaction was performed at high temperature (120 - 140 degrees C). Moreover, the ratio of trichlorobenzene isomers formed from alpha-hexachlorocyclohexane shifted to a pattern characteristic of the gamma (or gamma) isomer. However, at temperatures of 90 degrees C or less, beta-pentachlorocyclohexene was the main product. The results strongly suggest that in pyridine/xylene, the same isomer is primarily produced from alpha-hexachlorocyclohexane and is isomerized to the gamma, delta and at least two other isomers of pentachlorocyclohexene before further dehydrochlorination ensues. A simple method for the synthesis of beta-pentachlorocyclohexene is presented.


Symposium on pathology of the gastrointestinal tract-Part II. Small intestinal biopsy. A practical approach to the interpretation of peroral small intestinal biopsy specimens is presented. Biopsy technique and tissure handling are described. Interpretation of normal and abnormal biopsy specimens is discussed. A practical classification of abnormal small intestinal biopsies is presented and illustrated.


Whipple's disease. An example of the value of the electron microscope in diagnosis, follow-up, and correlation of a pathologic process. Two cases of Whipple's disease were followed by electron microscopic study of periodic peroral suction biopsy specimens of small intestine to show the presence and disappearance of the interstitial lamina proprial organisms, the sequential changes of the macrophages, and the return to normal leukocytic population of the lamina propria following prolonged treatment with tetracycline. The value of electron microscopy in the detection of small numbers of micro-organisms is demonstrated. Ultrastructural study is the most efficient method of demonstrating the presence of diagnostic micro-organisms, measuring the adequacy of treatment, and identifying possible early reactivation of infection.


The clinician's view of diagnostic electron microscopy. From the clinician's standpoint, it has become increasingly evident that use of the electron microscope should no longer be confined to research institutes, but should be applied as an adjunct to clinical diagnosis, a guide to therapy, and a means of elucidating pathogenetic mechanisms. The usefulness of electron microscopy is stressed in relation to hematology, nephrology, virology, gastroenterology, and the study of miscellaneous metabolic conditions such as the storage diseases.


[Late replication and X-autosome traslocation a case with banding patterns autoradiographic and B.U.D.R. studies (author's transl)]. A patient with ovarian failure was found to have a segment of the long arm of an X chromosome translocated to the distal end of the short arm of a 2nd chromosome: 46,Xt(X;2) (q21;p25). The Barr bodies were of normal size. Autoradiographic and B.U.D.R. studies showed that the normal X was the late-replicating X. The translocated segment was late replicating in 5% without spreading effect. It is suggested that preferential inactivation of the normal X chromosome is observed when a segment of X chromosome is translocated onto an autosome. When a segment of autosome is translocated onto an X chromosome the abnormal X chromosome is consistently late-replicating. These findings may be available for other mammalian species in the balanced translocations.


[Four new cases of X-autosome translocation in man (author's transl)]. Four new cases of translocations involving the X chromosome are reported. The first is a balanced t(1;X) observed in an abnormal girl. In most of the cells, the normal X is inactivated. The second case is a woman with oligomenorrhea, who has had two spontaneous abortions. She carries an umbalanced t(X-autosome). In all cells, the abnormal X, including the autosomal segment, is inactivated. The third and fourth cases are a mother, and her son. The mother has oligomenorrhea, carries a t(X,X), and has a karyotype of 46,X,+der t(X;X) and that of her son is 47,XY,+der t(X;X) and shows behavioral abnormalities. The abnormal X chromosome is inactivated in all the cells analyzed.


Biochemical and cytogenetic studies on the nucleolus organizing regions (NOR) of man. II. A family with the 15/21 translocation. The amount of ribosomal DNA (rDNA) was determined quantitatively by RNA-DNA hybridization in the genomes of a mother and her daughter, both with the karyotype 45,XX,t(15q21q). The saturation values found were 0.030% (mother), 0.023% (daughter), and 0.022% for the husband and father of the daughter. A detailed cytogenetic analysis of the short arms of the acrocentric chromosomes of these probands allowed the biochemical results to be interpreted in terms of the size of the individual set of nucleolus organizing regions (NORs) present in each proband. The correlation existing between the biochemical and the cytogenetic findings shows that the amount of rDNA in the human genome is not primarily a function of the number of acrocentric chromosomes, but depends on the individual combination of variant NORs occurring in the human genome.


Variants of normal human alpha2-macroglobulin. Immunoelectrophoresis and enzyme-binding effect. Three phenotypical variants of normal human serum alpha2-macroglobulin revealed by immunoelectrophoresis and specific antibodies obtained in rabbits are presented. The variants are characterized by differences in electrophoretic mobility: one being fast, one slow, and one of an intermediate rate. To find out possible differences with respect to the effect of the trypsin and chymotrypsin on the three variants, they were treated with the enzymes before immunoelectrophoresis. Migration was accelerated in all cases, after complexing with the enzymes, but the differences in the relative positions of the variants were maintained. The trypsin- and chymotrypsin-binding capacities of these three forms seem to differ, as suggested by the results presented in this report.


[Cat-eye syndrome. Clinical and cytogenetical differentialdiagnosis (author's transl)]. We report a 5 1/2-year-old girl whose clinical symptoms are consistent with diagnosis of the cat-eye syndrome. The prominent symptoms are: anal stenosis, preauricular tags and pits, coloboma of the iris, doubling of the pelvis and ureter on both sides, vesicourethral reflux on the right side and normal mental development. Leucocyte alkaline phosphatase is normal. Chromosomal analysis shows a supernumerary submetacentric chromosome. This extra chromosome is smaller than the G-group chromosomes and has satellites on the short and long arms. Autoradiography after 3H-thymidine incorporation shows a late-labeling marker chromosome. After using the Giemsa-banding technique, the chromatides demonstrate dark bandings with only soft, unstained satellites. With the fluorescence method, one can see spotlike fluorescence of the satellites on both arms and diffuse fluorescence of the hetero-chromatic segments. In addition, the C-bandings demonstrate a homogeneous dark staining of the chromatids, but we did not find stained satellites. Using the Giemsa-11 technique one can see the 47th chromosome with predominantly heterochromatic parts, but small euchromatic segments are visible between them. Satellites are unstained. Using currently accepted cytogenetical methods, it is not possible to identify the origin of this supernumerary marker chromosome.


Satellite DNA III and alkaline Geimsa staining. Satellite DNA III visualized by staining chromosomes with Giemsa at pH 10-12. Evidence is presented that besides the secondary constriction of chromosome 9, satellite III contained in considerable amount in the long arms of chromosome 20, giving rise to a clearly visible secondary constriction just below the centromere. The latter finding confirms that reported by Bobrow et al. (1972). The long arms of the Y chromosome also show strong staining with alkaline Giemsa, the region of staining corresponding exactly with the intensely flourescing area. This is interpreted as possible evidence for the presence of satellite DNA III in the distal long arms of the human Y chromosome.


High amniotic fluid alpha-1-fetoprotein in a case of fetal sacrococcygeal teratoma. Amniocentesis was performed in a pregnancy of 26 weeks because of hydramnios. The amniotic fluid was examined for fetal karyotype and AFP content. The latter was elevated to 300 mug/ml. Fetal death occurred shortly after amniocentesis and a malformed male fetus with a large sacrococcygeal teratoma was stillborn 3 days later.


[Non-random position of homologous chromosomes (no. 9 and YY) in interphase nuclei of human fibroblasts (author's transl)]. The position of chromosomes No. 9 and of the Y chromosomes in interphase nuclei was observed by means of the Giemsa-11 staining and the quinacrine mustard fluorescence staining respectively. 3 fibroblast cultures from normal female persons and 1 culture from a person with the karyotype 47/XYY were used. The distance between the two homologous chromosomes was compared with the theoretical expected distance between two points which are randomly positioned in a circular area. The distance between the chromosomes No. 9 as well as between the two Y chromosomes is significantly smaller than expected with a random position. This tendency for a somatic association is stronger in the case of the two Y chromosomes than in the two chromosomes No. 9.


Giemsa banding patterns in chronic lymphocytic leukaemia. The banding patterns of chromosomes from 20 patients with chronic lymphocytic leukaemia (C.L.L.) have been analyzed. 97 of 100 metaphases examined had a normal banding pattern. The 3 remaining metaphases, all from one patient had bands similar to those seen after aging. It is concluded that the chromosomes in C.L.L. have normal banding patterns. The majority of cytogenetic studies in chronic lymphocytic leukaemia have reported normal chromosomes (Fitzgerald and Adams 1965; Oppenheim et al., 1965; Lawler et al., 1968). An inherited abnormality of G group chromosome (No. 22) has been reported in a family, three members of whom developed C.L.L. (Fitzgerald and Hamer, 1969), but further investigations of cases of familial leukaemia failed to reveal a similar abnormality (Fitzgerald et. al., 1966). The development of new techniques which allow the positive identification of individual chromosomes (Caspersson et al., 1969; Dutrillaux and Lejeune, 1971; Sumner et al., 1971; Seabright, 1971), has revolutionised human cutogenetics and revealed additional information regarding chromosome abnormalities and leukaemia (Rowley, 1973; Lobb et al., 1972; Milligan and Garson, 1974). The purpose of this investigation was to determine whether  the chromosomes in C.L.L. have normal banding patterns.


Two cases of C-group balanced translocations. The present study describes 2 cases of a balanced reciprocal translocation in the C-group of autosomes. Familial translocation 46,XX,t(6;7) was found both in a woman cytogenetically examined because of a developmental anomaly of the internal genitals (uterus bicornis subseptus) and in her healthy mother. Chromosomal complement 46,XYt(8;10) was proved in the healthy father of a child showing clinical features of Down's syndrome with a karyotype 47,XY,21+. Q- and G-banding techniques were used for precise identification of the chromosomes involved in translocations.


The G-banded prophase chromosomes of man. Using a simple G-banding technique developed in our laboratory, analysis of late prophases enables the visualization of approximately 1000 bands in the haploid set of human chromosomes.  These bands have been classified according to the recommendations of the Paris Conference.  The increased resolution offered by this technique is likely to be useful in the study of the structure and molecular organization of chromosomes and in identifying minute chromosome defects in birth defects and neoplasia.


Frequency and distribution of sister-chromatid exchanges in a case of Fanconi's anemia. In lymphocytes of a 7-year-old boy with Fanconi's anemia the frequencies and sites of sister-chromatid exchanges (SCE) were studied with the BrdU-Giemsa method.  The average frequency of SCE (8.8 per metaphase) and the inter- and intrachromosomal distribution of SCE was not significantly different from the controls.


Ring chromosome 15:r(15).  Identification by R banding. Upon investigating the mental retardment of a small girl who was not dysmorphic, a ring chromosome D was discovered.  Thermic moderate denaturation enabled us to confirm it as being an r(15).  The r(15) syndrome scarcely affects the phenotype.  On the other hand, height is consistently reduced.


[Analysis of the karyotype of Pan paniscus. Comparison with other Pongidae and man (author's transl)]. The analysis of the karyotype of pygmee Chimpanzee (Pan paniscus), and its comparison with the one of Pan troglodytes shows some differences on chromosomes 2q, 7, 13, and 22. The study of the chromosomal rearrangements differentiating the Chimpanzees and the others Anthropoids and Man allows us to propose a filiation of ancestral species.


The differential distribution of X-ray induced chromosome lesions in trypsin-banded preparations from human subjects. The distribution of X-ray induced aberrations between trypsin-banded regions within human chromosomes has been studied in over a thousand cells from normal and abnormal individuals. An analysis of this data has been performed for each type of aberration separately, taking into account the relative amount of material at risk in reach region. A number of significant deviations from expectation were detected but many of these could be explained in terms of the failure of detection of some aberrations.


Dynamic aspects of trypsin-giemsa banding. The trypsin-Giemsa banding procedure was adapted so that chromosomes could be observed through the microscope during treatment and staining. Trypsin treatment resulted only in a swelling of the chromatids. Chromosome bands which appear as raised structures with interference contrast optics emerged only after staining with Giemsa. These structures remain after Giemsa destaining, suggesting that an irreversable change in chromosome structure is induced by Giemsa. Observations of the stain flow indicate that the positioning of the chromosomes has an effect on the quality of band production. These studies also revealed that bands appear in a reproducible sequence on individual chromosomes, which suggests that alterations take place at different rates along the length of the chromosomes.


A study of idiotypic suppression in adult rabbits immunized with Salmonella abortus-equi. It is possible to induce idiotypic suppression in adult rabbits immunized with Salmonella abortus-equi (S.a.e.).  Ten months after priming we injected the rabbit with anti-idiotypic serum prepared against its own antibodies to S.a.e. and, 3 weeks later, gave it a booster injection of bacteria.  A new anti-idiotypic serum was preparedwith the serum to S.a.e. collected after this boost and was used for the following isiotypic suppression attempt made 10 months after the first one.  Using this procedurewe succeeded in two successive idiotypic suppression attempts in the same rabbit.  Inthe three attempts we carried out, idiotypic suppression was totally effective, i.e. idiotypes detected by the serum used for the suppression totally disappeared after thesuppression, and the suppression lasted during the life of the rabbits (maximum 10 months).  This observation is consistent with a suppression resulti-g from an interaction of anti-idiotypic antibodies with the complementary receptors at the surface of memory cells.  This suppression was without effect on antibody to S.a.e. titre and on IgG concentration.  Idiotypes detected by the anti-idiotypic serum prepared withthe serum to S.a.e. collected after the suppression.  These idiotypes were different from those detected by the anti-idiotypic serum used for the suppression.  This observation confirms that idiotypic recognition is confined to a limited number of clonal products, despite the fact that a very heterogeneous antibody population was used forthe anti-idiotypic immunization.  Thus we did not observe the appearance of new idiotypes produced previously silent cell clones.  All the different idiotypes we detected during the successive idiotypic suppression attempts carried determinants shich remained peculiar to each individual rabbit.


The antigenic interrelations of some mammalian IgG subclasses detected with cross-reacting fowl antisera to human and mouse IgG-Fc. Two fowl antisera to plyclonal human IgG and one to a mouse IgG1 myeloma protein, after absorption either with homologous F(ab)2 alone, or also with myeloma proteins of known subclasses, cross-reacted in immunoelectrophoresis or immunodiffision withthe IgG of many mammalian species.  Often more than one IgG precipitation line was formed in the corss-reacting systems.  By testing isolated IgG1 and IgG2 fractions from mouse, rat, guinea-pig, bovine and dog sera, it was shown that the distinct arcs seen with whole serum corresponded to the known IgG subclasses of these species.  With rabbit serum as antigen, the two anti-human-Fc sera diffusing from neighboring wells formed a 'spur', showing that they were reacting with determinants on different molecules, and that therefore rabbit IgG contains two antigenically distinct Fcpopulations.  With the possible exception of canine IgG1 and human IgG4, no antigenic homologies were found between the subclasses of different species, thus supportingthe view obtained from sequence data, namely that IgG subclasses within species havearisen independently.


I. In vitrol transformation of rat embryo cells: correlations with the known tumorigenic activities of chemicals in rodents. Susceptibility to chemically induced transformation changed as a rat embryo cell culture was passaged. For the first 35 to 60 passages, the cultures were diploid and resistant to transformation by chemical carcinogens. However, cultures infected with a murine leukemia virus were transformed by chemicals. For the next 60 passages, the cultures were heteroploid, but retained contact inhibition and were not tumorigenic. Even without addition of heterotypic viruses, these heteroploid cultures could be transformed by chemicals, but the endogenous rat C-type virus could be demonstrated in the transformed cultures. At higher passages, the rates of spontaneous transformation gradually increased so that the cultures could not be used for transformation studies. Chemically induced transformation of the stable heteroploid cell line (F1706) was manifested by an easy to read focal alteration. Initial observations based on these foci were confirmed by inoculating the morphologically altered cells into isogeneic newborn rats. A number of carcinogenic and noncarcinogenic chemical analogues were tested for their ability to transform F1706 cultures. The compounds tested included 4 azo dyes, 12 polycyclic hydrocarbons, 12 aromatic amines, and 7 miscellaneous compounds. Based on the known activities of the same chemicals in rodents, certain active compounds failed to induce transformation in any test, and others induced transformation in only some tests, but these in vitro tests, if used as a screening assay, would have been correct in 82% of all individual tests, and over-all, would have correctly predicated the carcinogenic activity of 33 of the 35 agents tested.


Immunological properties of procollagens obtained from the culture medium of dermatosparactic cells. Rabbit antisera were produced against purified calf dermatosparatic procollagen and against the purified procollagens obtained from the culture medium of calf dermatosparatic cells.  These antisera and their derived gamma-globulins were characterized by immunoprecipitation, double immunodiffusion and immunoelectrophoresis.  Antiserum directed against dermatosparatic procollagen cross-reacted with the two different forms of procollagen obtained from the culture medium of dermatosparatic calf cells.  Antiserum directed against onw of these procollagens, namely (pro alpha1)2 pro alpha2, cross reacted with dermatosparatic procollagen and also cross-reacted with the other procollagen,(pro alpha1)3.  Antiserum directed against procollagen (pro alpha1)3 cross-reacted with dermatosparatic procollagen and with the procollagen (pro alpha1)2 pro alpha2.  None of the antisera reacted with authentic calf skin collagen, or with the collagen extracted from the cell layer of the dermatosparatic calf cells in culture.  Reduction andalkylation of the procollagens abolished the antigen-antibody reactions, while prior digestion of the antigens with bacterial collagenase did not eliminate the immunological reaction.  Antigenic determinants in the cell culture procollagens were found at the COOH-terminal non-collagen peptide as well as at the NH2-terminal non-collagen peptide.


Antibody response in the parotid fluid and serum of Irus monkeys (Macaca fascicularis) after local immunization with Streptococcus mutans. The antibody response of Macaca fascicularis in parotid saliva and serum to local immunization by two routes with Streptococcus mutans was studied and compared over 1 year. Antibodies were titrated and classified by indirect immunofluorescent staining using specific antiglobulin conjugates. Antiglucosyltransferase activity was assayed by an enzyme inhibition test. Animals were immunized first by injecting formalin-killed bacterial cells and cell products subcutaneously into the vicinity of the four major salivary glands. The monkeys were next immunized by retrograde instillation of antigen into the parotid duct. Extensive subcutaneous local immunization gave a serum response only. After parotid duct immunization, high titers of immunoglobulin A (IgA) antibody, along with traces of immunoglobulin G (IgG) immunoglobulin M (IgM) antibody, appeared in the parotid saliva, and in the serum high titers of IgG antibody were present along with lower titers of IgA and IgM. IgA antibodies in parotid fluid were shown by double immunofluorescent staining to be associated with antigenic determinants which cross-reacted with an antiserum directed to human secretory component. Titers in parotid fluids and sera fell sharply when immunization was stopped. This response pattern was reproducible. High concentrations of antibody capable of inhibiting glucosyltransferase prepared from S. mutans were found in the sera, but relatively little was detected in the parotid fluids. Extensive immunization via the parotid duct resulted in transient functional impairment of the gland, as evidenced by diminished salivary flow rates. We conclude that parotid ductal immunization can be an effective method for stimulating a salivary secretory IgA antibacterial antibody response.


Inhibition by nicotinamide of an homologous PCA reaction and antigen-induced histamine release from rat peritoneal mast cells. The antigen-induced release of histamine both in vitro from rat peritoneal cell suspensions containing mast cells actively sensitized with IgE antibody and in vivo from passively sensitized skin can be inhibited by relatively high doses of nicotinamide. The action of nicotinamide on the cellular mechanism involved in the regulation of antigen-induced histamine release is discussed.


Comparison of lymphocyte populations bearing surface immunoglobulins in avian bone marrow, bursa, spleen and thymus. Rabbit anti-chicken gamma-globulin was labeled with 125I and then incubated with cells from the bursa, thymus, spleen, and bone marrow of 4- and 8-week old birds. The same procedure was carried out on 11-week-old agammaglobulinemic chickens. Autoradiography revealed that the majority of large, medium, and small bursal lymphocytes bind the antibodies while labeled lymphocytes of each type in the spleen and thymus never exceeded 11 or 4 percent, respectively. Labeled medium and small lymphocytes in the bone marrow increased from 4.2 and 1.7%, respectively, at 4 weeks of age, to 9.5 and 8.8%, respectively, at 8 weeks of age. Labeled lymphocytes of all sizes were completely absent in all tissues of agammaglobulinemic chicks, including the marrow. Therefore, the increase in frequency of labeled lymphocytes in the bone marrow with age may be a result of recruitment of cells from the bursa of Fabricius. The majority of lymphocytes in the bone marrow do not label. Therefore, lymphocytes from the bone marrow may be T cells, subsets of B cells, or neither T or B cells.


Drug inhibition of anaphylactic histamine release from peritoneal cells of rats infected with Toxocara canis. An experimental model for studying anaphylactic histamine release from rat peritoneal cells in vitro is described. Rats were sensitized by feeding the infective eggs of ttoxocara canis, the antigen being prepared from the infective eggs. The characteristics of histamine release are comparable with those found for other anaphylactic systems. Addition of 0.2% inactivated normal rat serum to the incubation media reduced the spontaneous histamine release from 11.5 +/- 1.9% to 3.1 +/- 1.6% (mean +/- SD) of total histamine, whereas the corresponding figure for anaphylactic histamine release was 41.3 +/- 19.7%. Both disodium cromoglycate (DSCG) and levamisole produced a dose-related inhibition of histamine release, with DSCG being the more potent drug.


Soluble tumour-specific antigen and its relationship to tumour growth. Hooded rats bearing a syngeneic methylcholanthrene-induced tumour were evaluated for extent of in vivo host immunity and this was correlated by in vitro techniques with the levels of circulating tumour antigen and specific antibody. Early tumour growth was associated with detectable immunity, as measured by the capacity of the animal to reject a second direct challenge of the same tumour at a remote site. Radioimmunoassay for circulating tumour antigen and indirect membrane immunofluorescence for antitumour antibody did not detect either component at this stage. Animals with advanced tumours lost immunity as detected by direct tumour challenge, and this closely coincided with the appearance of rising levels of circulating soluble tumour antigen. Although the host possessed the immunologic ability to react against its own neoplasm, this ability was insufficient to produce tumour rejection. Active immunotherapy initiated at the time of, or up to 10 days after, intramuscular challenge with tumour, increased tumour immunity sufficiently for tumour growth to be prevented. Successful immunization was associated with the early appearance (16 days) of measurable levels of antitumour antibody and absence of circulating antigen. It is concluded that soluble tumour antigen present in the local microenvironment of the tumour in the early stages of tumour growth interferes with the ability of immune cells to cause tumour rejection. As the tumour progressively grows, sufficient soluble antigen is produced and released systemically to suppress the effector arm of the host's tumour immune response at distant sites. The levels of circulating soluble tumour antigen attained may be of critical importance in the suppression of rejection responses that prevent metastasis.


Are methylcholanthrene-induced sarcoma-associated, rejection-inducing (TSTA) antigens, modified forms of H-2 or linked determinants? TA3Ha/MSWBS hybrid cells have been derived from the fusion of the TA3Ha ascites carcinoma (H-2a) and the methylcholanthrene-induced MSWBS ascites sarcoma (H-2s). MSWBS expresses a strong tumor-specific transplantation antigen (TSTA), capable of inducing a rejection reaction in the syngeneic A.SW host. The genetic determinants of the H-2 complex are known to be localized on chromosome No. 17. TA3Ha contributes two normal, telocentric chromosomes No. 17 to the hybrid. In contrast, both chromosomes No. 17 of MSWBS are localized on readily identifiable translocations (17/1 and 17/M1 ; see Wiener et al., 1974). We have previously shown that the chromosomes No. 17 of one parental strain, or the other (but not both) can be removed from the hybrid by selective passage in the opposite parental strain. The present paper examined the possibility, often suggested in the literature, that the MC-sarcoma-associated TSTA could be a modified form of H-2. MSWBS, unselected TA3Ha/MSWBS and YACIR/MSWBS hybrids were compared with TA3Ha/MSWBS-derived isoantigen loss variants, with regard to their immunogenicity in the TSTA test, i.e. their ability to induce rejection of MSWBS target cells in ASW mice. Whereas the unselected hybrids were as immunogenic as the parental MSWBS line itself, two strain A compatible and two strain A.SW compatible variants which had lost chromosome No. 17 of the opposite strain showed a residual, but clearly weakened immunogenicity. Since there was no systematic difference between the reciprocal types, it is concluded that the genetic determinant of TSTA is not localized on the chromosome No. 17 but that a proper balance of this chromosome is required for the full expression of immunogenicity in the TSTA system.


Influence of interferon on the synthesis of virus particles in oncornavirus carrier cell lines. III. Survey of effects on A-, B- and C-type oncornaviruses. The effect of interferon on the synthesis and release of A-, B- and C-type viruses by oncornavirus carrier lines was studied. Murine cell lines were selected which carry either of these viruses and are sensitive to the antiviral effect of interferon, as measured by inhibition of vesicular stomatitis virus. Release of C-type virus was found to be highly sensitive. Release of B-type virus, on the contrary, was only marginally inhibited. Synthesis of intracisternal A-type particles was finally not inhibited by interferon pretreatment. These differences between infectious C-type and non-infectious A- and B-type viruses may reflect fundamental differences in the synthesis of these viruses.


A fetuin-like antigen from human nephroblastoma. An antigen was detected in pooled human nephroblastomas using antiserum prepared in rabbits against an ethylemediaminetetra acetic acid (EDTA) extract of the tumors. This antigen was not found in normal human plasma or kidney extracts, and was not related to the ABO or Forssman blood groups. The antigen was detected in extracts of cultured nephroblastoma cells, but was not present in extracts of normal human fetal kidney cell cultures. The antigen is believed to be present at the cell surface, as cell viability was not significantly lowered during the extraction procedure. A reaction of complete identity was demonstrated by Ouchterlony double diffusion experiments with this antigen and purified bovine fetuin. The antigen was not found in extracts of human fetal spleen, thymus or kidney, nor in human fetal serum. Furthermore, the antigen does not possess determinants in common with the human alpha-fetoprotein of hepatomas, nor was it detected in human renal clear cell carcinoma. Initial characterization of the antigen showed it to be nondialysable, not sedimentable at 100,000 times g for 2 h, stable to repeated freeze-thawing and to incubation at 56 degrees C for 1 h, and water soluble over a wide pH range. The antigen was susceptible to digestion with pronase and trypsin and possibly hyaluronidase, but not to ribonuclease or neuraminidase. The protein portion is therefore of major importance to the structural integrity of this antigen. The relationship between this antigen and other abnormal materials reported previously in nephroblastoma patients is being studied.


Natural cytotoxic reactivity of mouse lymphoid cells against syngeneic acid allogeneic tumors. I. Distribution of reactivity and specificity. Lymphoid cells from many normal mice of a variety of inbred strains were found to have reactivity, in a 51Cr release cytotoxicity assay, against several syngeneic and allogeneic tumors. Very high reactivity was seen with effector cells from athymic nude mice, which was consistent with other evidence that the reactivity was not T-cell dependent. Target cells susceptible to lysis included tumors induced by oncogenic type-C viruses but also tumors induced by other means and expressing endogenous type-C viruses. The levels of natural reactivity were influenced by age, with highest cytotoxicity produced by cells from 5- to 8-week-old mice. Lymph-node cells, spleen cells, peritoneal exudate cells and peripheral blood lymphocytes all had cytotoxic reactivity. The specificity of the reactions was analyzed in detail by ana inhibition assay. Evidence was obtained for natural reactivty against several different antigens, each apparently associated with expression of murine endogenous type-C viruses.


Comparison of intracytoplasmic A particles and intracisternal A particles. Intracytoplasmic A particles and intracisternal A particles are associated with mouse tumors of various types, and both can coexist in the cytoplasm of the same cell. The designation of both as A particles is based on the recognition that they share morphological similarities. A comparison of purified isolates of these two particles reveals that the structural protein profiles are different, but that there is some antigenic cross-reaction as demonstrated by immunodiffusion and complement fixation. This result does not appear to involve the major structural protein of intracisternal A particles, but may reflect the presence of common antigenic determinants located on minor proteins.


Spleen-cell reactivity against transplanted neurogenic rat tumors induced by ethylnitrosourea: uncovering of tumor specificity after removal of complement-receptor-bearing lymphocytes. Spleen cells from BDIX-rats bearing either GVlAl-tumor (a syngeneic mixed glioma) or NVlAc-tumor (a cloned syngeneic neurinoma of the peripheral nervous system) were cytotoxic to both tumor cells in vitro. However, the tumors displayed individually distinct antigenic specificities by in vivo rejection tests. Their in vitro cross-reactivity disappeared when a particular subpopulation of the spleen cells was used. The procedure of lymphocyte purification included three consecutive steps: treatment with carbonyl iron and magnetism, passage through a nylon wool column, and finally removal of complement receptor-bearing cells present in the colum-excluded population. Cross-reactivity between the syngeneic tumors persisted after the first two steps of lymphocyte purification. In contrast, specific cytotoxic reactions were observed against each individual tumor subsequent to the removal of the remaining C3 receptor-positive but surface Ig-negative cells. While killer cells were present in normal spleen-cell populations, these were almost completely eliminated by passage through the nylon wool column.


Identification of an antigen associated with malignant melanoma. Xeno-antisera, prepared by immunization with soluble membrane material obtained from melanoma tumours and partially purified by column chromatography, were found to detect an antigenic specificity common to some tumour extracts and sera of melanoma patients. The presence of this antigen in patients' sera allows speculation for its possible use for early diagnosis of metastases and its role in the blocking of the immunological defences of the melanoma patients. Preparation of monospecific antisera by immunization with insoluble serum-antigen/antibody complexes has been successfully undertaken.


Penicillin immunogenicity in the presence of different pharmaceutical adjuvants. Preliminary results in the variations of the penicillin immunogenicity in the presence of aminosugars, polysaccharides and terpenic substances are presented and the decreasings of the potential antigenicity of the penicillin molecule appeared in the presence of hexosamine molecules are estimated. It might be possible as a consequence of modifications in the early immunogenic interactions between antibiotic and serum proteins of the hipersensitive animals.


Neuroichthyosis with hypogonadism (Rud's syndrome). Rud's syndrome is a neuroichthyosis with hypogonadism, associated with mental deficiency and epilepsy. Short stature is a frequent component of the syndrome. The primary genetic defect and the pattern of inheritance have not yet been determined. A typical patient is presented, with mental deficiency, short stature, pypoacusia, muscular atrophy, tylosis, pseudoacanthosis nigricans and endocrine disturbances. The neuroichthyosis with hypogonadism must be considered Rud's syndrome. A classification of neuroichthyosis is proposed. In a first group is neuroichthyosis with hypogonadism, in the second group is neuroichthyosis with spasticity and in the third group, neuroichthyosis without hypogonadism or spasticity.


Localization of acetylcholinesterase in the rabbit cornea by light and electron microscopy. Acetylcholinesterase (AChE) has been localized in the rabbit cornea by light and electron microscopy histochemical techniques. In the stroma, the enzyme is concentrated in nerves. In the epithelium, the enzyme is concentrated in intercellular spaces devoid of nerves. The morphologic appearance of the enzyme staining by light and electron microscopy in the epithelium is similar; consequently, the staining demonstrated with light microscopy examination does not always represent epithelial nerves. A significant portion of corneal acetylcholinesterase therefore appears unrelated to nerves. Considerably smaller deposits of enzyme reaction product were present in cells in every layer of the cornea, using electron microscopy histochemistry; they were not identified by light microscopy.


Storage granules of thyroid C cells in the dog: a cytochemical and ultrastructural study, in relation to the masked metachromasia reaction. Masked metachromasia can be demonstrated in thyroid C cells, and other cells of the APUD series, by staining with a metachromatic basic dye after hydrolysis of suitably fixed tissue. The reaction is thought to be due to the presence of polypeptides with a high concentration of side-chain acidic groups. Since most APUD cells possess storage granules, presumed to contain a polypeptide hormone, it has been assumed that the masked metachromasia reaction gives information concerning the contents of these granules. However, there has been an increasing suspicion that the reaction might actually be due to the membrane bounding these granules, rather than to the contents. We have examined, cytochemically and ultrastructurally, dog thyroid tissue which has been subjected to fixation and hydrolysis as in the usual method for masked metachromasia. We found that the membrane surrounding the C cell granules is removed by hydrolysis, confirming the hypothesis that the reaction is due to the contents (hormone and/or matrix)rather than to the membrane. Tissues were fixed in an aqueous mixture containing glutaraldehyde (6 25% v/v), picric acid (three-quarters saturation) and sodium acetate (I% W/V)adjusted to PH 7 with sodium hydroxide. This was found to be a very satisfactory fixative for electron microscopy Some morphological details of C cells were noted, such as the richness of desmosomes between C cells in this species, and frequent direct contact with the colloid.


Technical aspects of the Haematoxylin--Basic Fuchsin--picric acid (HBFP) stain applied to skeletal muscle. The Haematoxylin-Basic Fuchsin-picric acid (HBFP) stain, a new non-enzymatic histochemical technique described previously to detect early myocardial ischemia, was applied to skeletal muscle. Several factors were found which have an important effect on HBFP positivity including ageing in room air of unstained tissue sections, and the precise timing of the differentiation step of this stain. Using carefully standardized techniques, repeatable staining was obtained and a high level of interobserver consistency in the interpretation of staining results was achieved. Although the technical requirements of this new stain are rigorous, it offers promise and deserves further evaluation in the study of skeletal as well as cardiac muscle disorders. The histological advantages include vivid contrasts and the ability to use the stain on formalin-fixed paraffin-embedded muscle tissue.


An autoradiographic study of human blood basophils. The aim of this paper is the selective visualization of human blood basophils in autoradiographs. [(3)H]Thymidine-labelled basophils in buffy coat smears were fixed in methanol-formaldehyde follwed by 5-aminoacridine hydrochloride and stained with basic Aldehyde Fuchsin prior to autoradiographic processing. The technique described represents a simple method for quantitative autoradiographic studies on basophil kinetics and the interaction of these cells with IgE-mediated atopias.


Staining of sulphatides in metachromatic leukodystrophy with Alcian blue at high salt concentrations. Alcian blue combines with purified sulphatide in 1.OM magnesium chloride. In tissue sections from patients with metachromatic leukodystrophy, sulphatide is stained by Alcian blue in 0.8 M magnesium chloride, and the staining can be abolished by prior treatment with chloroform and methanol. The simplicity of the technique, its specificity and ease of interpretation recommend Alcian blue staining at high salt concentrations as a routine method in the diagnosis of metachromatic leukodystrophy.


Background staining and sensitivity of the unlabelled antibody-enzyme (PAP) method. Comparison with the peroxidase labelled antibody sandwich method using formalin fixed paraffin embedded material. The PAP procedure was compared with the peroxidase labelled antibody sandwich method and was found to be at least 20 times more sensitive. Background staining was reduced by the addition of normal swine serum to all the immune sera or by pretreating sections with it at the beginning of either method. The PAP procedure could be effectively reduced to a period of 1 hour or less.


Aminoglycoside 3'-phosphotransferases I and II in Pseudomonas aeruginosa. Aminoglycoside 3'-phosphotransferases I and II in three strains of Pseudomonas aeruginosa were studied in comparison with those in two strains of R factor-carrying Escherichia coli. The strain TI-13 of P. aeruginosa produced the former and strain H-9 the latter. Strain B-13 produced the both enzymes. The 3'-phosphotransferases of type I in P. aeruginosa TI-13, B-13 and E. coli K12 J5 R11-2 were different from each other in chromatographic behavior, molecular weight, pH optimum, and Ii. The 3'-phosphotransferase of type II in P. aeruginosa H-9 and E. coli JR66/W677 showed the same behavior.


5"-Amino-3',4',5"-trideoxybutirosin A, a new semisynthetic aminoglycoside antibiotic. 5"-Amino-3',4',5"-trideoxybutirosin A (IX) was synthesized through a reaction series starting from 5"-amino-5"-deoxybutirosin A (Ic), the key step being the treatment of its tetra-O-acetylpentakis-N-((phenylemthoxy)carbonyl)-3',4'-bis-O-(methylsulfonyl) derivative (VI) with zince-sodium iodide. Compound IX inhibits enhanced antibacterial activities, including strains of Pseudomonas aeruginosa and Escherichia coli which are highly resistant to Ic, butirosin or gentamicin.


Synthesis of aminotrideoxybutirosin A, a chemically modified antibiotic active against butirosin-resistant bacteria. 5"-Amino-3',4'5"-trideoxybutirosin A (4) was synthesized by two routes starting from the known tri-O-acetyl-tetra-N-benzyloxycarbonyl-3",5"-O-cyclohexylidene-3',4'-di-O-mesylbutirosin A (5). Introduction of amino function at C-5" was carried out by displacement of 5"-tosyloxy group with sodium azide either before or after 3',4'-deoxygenation. Compound 4 shows outstanding activities against strains including Pseudomonas aeruginosa and Escherichia coli which are highly resistant to butirosin and 5"-amino-5"-deoxybutirosin A (2).


Characterization of bleomycin action on DNA. Cleavage of SV40 DNA by bleomycin was assayed quantitatively in vitro in the presence of various ppolynucleotides. SV40 DNA was protected from bleomycin-  duced cleavage by native or denatured DNA of other origins, oly dG-C.poly dG-C, poly dA-T.poly dA-T and poly dA-T (denatured) but not by tRNA of E. coli, apurinic acid, poly dA, poly dT and various deoxyribooligonucleotides. Various bleomycins and their derivatives and various fragments of bleomycin were tested for possible activity in cleaving SV40 DNA and from the results some structure-activity relationships for the action of bleomycin to act on DNA were outlined. Actinomycin D stimulated bleomycin action while ethidium bromide inhibited it.


Residues in broiler chick tissues from low level feedings of seven chlorinated hydrocarbon insecticides. Two chlorinated insecticide feeding studies from 1967-1968, using broiler chicks, have been completed. In Study I, lindane, heptachlor epoxide, dieldrin, endrin, methoxychlor, and DDT were fed in combination at 3 levels: 0.05, 0.15, and 0.45 ppm. Data show that, in fat tissues, heptachlor epoxide attained a level approximately 20 times the respective levels in the feed; dieldrin 15 times; endrin 10 times; p,p'-DDT 9 times; lindane 3 times; and o.p'-DDT less than the feeding levels. Of the DDT metabolites, p.p'-DDE and p,p'-DDD, only p,p'-DDE was significant at 3 times the 0.45 ppm feeding level. Endrin ketone, a metabolite of endrin, reached plateau levels approximately equal to feeding levels. All residue levels in liver tissues were less than 0.02 ppm. In Study II, technical chlordane was fed singly at the 0.05, 0.15, and 0.45 ppm levels. Results are tabulated for both total chlordane and for 6 identifiable isomers. Total chlordane in fat tissues attained plateau levels 3-5 times the respective feeding levels. Total chlordane levels in liver and breast tissues were all less than 0.01 ppm.