text:
|Publication number
|Publication type
|Application number
|Publication date
|Dec 16, 1980
|Filing date
|Apr 13, 1979
|Priority date
|Jun 30, 1973
|Publication number
|Dezso I. Bartos, Jerzy Rybczynski
|Original Assignee
|Dezso Istvan Bartos
|Export Citation
|BiBTeX, EndNote, RefMan
|Patent Citations (8), Non-Patent Citations (1), Referenced by (17), Classifications (23)
|External Links: USPTO, USPTO Assignment, Espacenet
This is a division of application Ser. No. 888,041 filed Mar. 16, 1978, now U.S. Pat. No. 4,162,003, which in turn is a continuation of Ser. Nos. 793,653 filed May 4, 1977; 693,801 filed June 7, 1976 and 483,659, filed June 27, 1974, all abandoned.
This invention relates to a ready to use rapid test package for carrying out serological investigations, in particular for carrying out the complement binding reaction.
When foreign substances are taken in parenterally, the body frequently produces a defence reaction in which so-called antibodies are formed. Substances which provoke the formation of antibodies are known as antigens. These are usually mixtures of proteins or their degradation products, lipoids and polysaccharides, for example bacteria, viruses, blood corpuscles and toxins. The antibodies formed, which are also termed immune bodies in cases where the antigens have a cell damaging action, bind the antigens by a highly specific reaction. The presence of antibodies therefore results in a specific resistance of the organism to the corresponding antigen.
Because of their high grade specificity, antibodies can be used to identify the corresponding antigens. If the presence of certain immune bodies can be demonstrated in an organism, it can be concluded with a high degree of certainty that the organism has been exposed to the corresponding antigen. The method of detecting the antibodies is, in principle, extremely simple and comprises bringing a body fluid, for example blood, into contact with antigen. If an antigen-antibody reaction takes place, then this is evidence for the presence of the antibody. It is particularly important that evidence for exposure to antigens can be obtained via the corresponding antibodies in cases where the antigens are no longer present in the organism.
The detection of antibodies which belong to the group of lysins is frequently carried out by the so-called complement binding reaction. Lysins are antibodies which have specific bacteriolytic, virucidal, cytolytic or haemolytic properties which are demonstrated when they are brought into contact with the corresponding antigens in the presence of a non-specific component present in every normal serum, the so-called complement. Without the complement, the specific antibody remains inactive. Since lysins must combine with two substances to manifest their action, the antigen and the complement, they are known as amboceptors.
The complement or complement system (abbreviated K or C') is a labile serum function which, as far as is known today, comprises at least 10 serum enzyme factors and which loses its activity when heated to 56� C. for half an hour or when left to stand at room temperature. The possibility of inactivation of the complement system C' by heating is an essential part of the method of carrying out serological reactions. When a patient's serum which contains antibodies has been heated to 56� C. for 30 minutes, it can be mixed with the corresponding antigen without a reaction taking place. The reaction between antigen and antibody (in this case lysin, in other words an amboceptor) then occurs only when a system which contains complement is added, for example fresh, unheated serum.
The dependence of the reactions between antigens and amboceptors on the presence of complement can be demonstrated particularly impressively in the so-called haemolytic system. By haemolytic system is meant a system of erythrocytes and the amboceptors which are specific for these erythrocytes. In practice, the amboceptor used is a serum which contains only the amboceptor component and not any of the other reactants necessary for the antigen/antibody/complement reaction, in other words a serum which has been heated to 56� C. for sufficiently long to inactivate the complement system C'. The erythrocytes used are generally sheeps' erythrocytes and the specific amboceptor a complement-inactivated rabbit serum which has been sensitised to sheeps' erythrocytes. An erythrocyte suspension is first added to an amboceptor solution to obtain a suspension of erythrocytes in serum. When this is added to a serum which contains complement, generally a fresh guinea pig serum free from blood corpuscles, haemolysis takes place, i.e. the membranes of the sheeps' erythrocytes are destroyed after the addition of complement and the blood pigment goes into solution.
The reaction between the haemolytic system and the complement (by complement is always meant in the following text a fresh serum which contains complement) is therefore a true colour reaction which indicates the presence of complement both quantitatively and qualitatively. It is only in the presence of complement (disregarding the cases of spontaneous haemolysis which may occasionally occur and which will be discussed hereinafter) that the originally colourless to pale yellow solution containing the erythrocytes takes up blood pigment. The intensity of the red colour depends, within certain limits, on the quantity of activity of the complement. If the complement is present in large quantities or is highly active, haemolysis or deep colouring of the solution above the blood corpuscles takes place within a short time whereas, if only a small quantity of complement is present or if it is less active, the same haemolytic system will undergo only a small amount of haemolysis in the same time.
The haemolytic system is therefore a suitable indicator for reactions which require complement, i.e. it can serve as an indicator system for detecting the reactions of other antigens with their corresponding antibodies, in particular those antigen/amboceptor/complement reactions which are not so obvious in their appearance as haemolysis. The principle is very simple. The serum which is to be tested for the presence of antibodies and in which the complement has been inactivated (usually referred to as test serum or patient's serum) is brought together with the corresponding antigen, and the complement is then added. If the test serum does not contain specific antibodies for the antigen used, then the complement added is not used up. The complement is therefore still available for other antigen/complement reactions. If, therefore, the test system or test serum and antigen also contains the haemolytic system (which is added to the other component after incubation), then haemolysis takes place and the solution becomes coloured. The occurrence of haemolysis is taken as a negative test result, i.e. as a sign that no specific antibodies for the given antigen are present. If, however, the test serum contains antibodies which react with the given antigen in the presence of complement, then little or no complement is available for haemolysis. Haemolysis is therefore absent or attenuated. The absence of haemolysis in a system composed of test serum, antigen, haemolytic system and complement is therefore taken as a positive test result, in other words an indication that the test serum contains the antibodies corresponding to the given antigen.
The test method described above has become established in serological laboratories as the `complement binding reaction` and is widely used to detect pathogens by way of their corresponding antibodies.
The complement binding reaction has become particularly important for the diagnosis of syphilis but it is also used to an almost similar extent to diagnose brucellosis, echinococcus, gonorrhoea, whooping-cough, leptospirosis, foot and mouth disease, mumps, pemphigus, rhinoscleroma, glanders, tuberculosis, toxoplasmosis, trichinosis and virus and other diseases.
Although the principle of the complement binding reaction described above is very simple, it is in practice very complicated to carry out as can be seen from the example of the syphilis test described below. This test generally requires five reactants or solutions of reactants which must be stored separately:
(1) The patient's serum (P) is obtained by running blood (generally venous blood) into a test tube, detaching the blood clot from the wall of the tube with a glass rod or a long platinum needle, and leaving the glass tube to stand overnight at a temperature of about 2� to 5� C. The serum usually separates as a clear fluid which can easily be pipetted off. Freshly drawn blood must not be immediately vigorously cooled and centrifuged because the serum is then liable to manifest non-specific inhibition of haemolysis. The serum which has been pipetted off is heated to about 56� C. for half an hour to inactivate the complement. It is generally diluted with five times its volume of 0.9% sodium chloride solution which should as far as possible be sterile.
(2) Sheeps' blood erythrocytes (E) are generally used as a 2 to 4% suspension of washed erythrocytes in physiological saline solution. Instead of the physiological saline solution, which is also used for washing, similar solutions may be used such as Kolmer of Osler-Strauβ-Meier solution. The concentration of the suspension may be adjusted by various methods, in particular by the method of haematocrit determination.
(3) The haemolytic amboceptor (A) used to complete the haemolytic system (EA) is rabbit serum which has previously been exposed to sheeps corpuscles. The serum is complement-inactivated as described under (1). The haemolytic titre of the amboceptor should be at least 1:1000. By haemolytic titre is meant that dilution of guinea pig serum at which a given quantity of sheeps' blood corpuscles will be just completely haemolysed under certain standard conditions in the presence of a sufficient or excess quantity of complement. The titre is generally determined on a system of 0.5 ml of amboceptor dilution (serial dilution), 0.5 ml of complement dilution, 0.5 ml of 2% sheeps' erythrocyte suspension and 1.5 ml of diluent (physiological saline solution, Kolmer salt solution, Eagle solution or Veronal buffer solution) incubated at 37� C. for 60 minutes. The amboceptor should preferably have a titre of at least 1:4000, in other words it should still bring about complete haemolysis under standard conditions when diluted to 1:4000. The quantity of amboceptor used for the test is four times the quantity of amboceptor which causes solution after 1 hour.
(4) The complement (K) is fresh guinea pig serum. It is diluted immediately before use, e.g. with physiological saline solution. The quantity of complement required is determined by a method of serial dilutions similar to that indicated under (3), in which varying quantities of complement are added to a test system of test tubes containing suitably adjusted quantities of sheeps' erythrocytes (E) and haemolytic amboceptor (A), and the contents of each test tube are diluted to the same total volume (generally 3 ml) with diluent and incubated at 37� C. for 60 minutes. This method determines what is the least quantity of complement required to bring about complete haemolysis. The quantity used in practice is generally about twice this quantity.
(5) The antigen (Ag) used for the complement binding reaction for syphilis is generally cardiolipin antigen or Reiters antigen. Cardiolipin is a phospholipid obtained from ox heart, which forms an antigen-antibody complex with the antibodies produced in response to the syphilis pathogen. The possibility of using cardiolipin is based on the fact that the antibodies produced in response to a given antigen will undergo an antibody-antigen reaction not only with the original virulent antigen but also with certain other antigens which are structurally extremely similar, for example antigens whose virulent action has been attenuated by certain chemical reactions. The specificity of the antibody is to that extent not absolute. This is of great importance in practice since it makes it possible for the presence of antibodies against highly dangerous or difficultly obtainable antigens to be tested by using substitute antigens.
The cardiolipin antigen ready for use may comprise e.g. a 0.0175% alcoholic solution of cardiolipin containing 0.0875% of lecithin and 0.3% of cholesterol, and is available commercially. It is generally used in a dilution of from about 1:100 to about 1:200. Other preparations of the cardiolipin antigen are also possible.
Reiter's antigen comprises preserved spirochaetes grown on culture medium and activated by ultra high-frequency sound. It is also available commercially and used in dilutions of between 1:2 and 1:8.
(6) Lastly, a stock of diluent must be kept in the serological laboratory. As already mentioned above, physiological saline solution may be used (0.9% or if sheeps' erythrocytes with high resistance to haemolysis are used then 0.85%). Instead of physiological saline solution, one may also use Kolmer solution which contains magnesium sulphate in addition to sodium chloride, or Eagles buffer solution, which contains a K/Na phosphate buffer in addition to sodium chloride, or an Osler-Strauβ-Meier solution or similar solution. It is generally advantageous to use solutions which contain magnesium and/or calcium ions because the enzyme reactions of the complement binding reaction require divalent cations as co-factors. The diluted solutions generally contain additions of phenol (e.g. 0.3% by weight) to stabilise them.
The serological tests using the complement binding reaction are extremely difficult to carry out in practice, firstly because the reactants vary considerably in their activity from one batch to another and lose their activity with varying rapidity in storage and secondly because the activities of the reactants must be adjusted to each other. It is obvious that such adjustment of activities is necessary. If, for example, the test system contained considerably more complement than is required for releasing the reaction between the patient's antibodies and the antigen, then rapid haemolysis would take place in every case. The test would therefore be less sensitive or even meaningless. Conversely, in the presence of insufficient complement the test becomes excessively sensitive so that false positive results may be obtained. The same applies to the adjustment of concentrations of the two components of the haemolytic system as to the adjustment of concentrations of complement to haemolytic system. The erythrocyte suspension must also be adjusted to the amboceptor dilution. The amboceptor solution used for the test is usually adjusted so that it has about twice the minimum concentration which is required for complete haemolysis when an excess of complement is present. In the description given below it should always be assumed, unless expressly stated otherwise, that the reactants are brought together in adjusted quantities as described above. Mixtures of the reactants are expressed for the sake of simplicity in terms of the abbreviations of their components indicated above, for example "EAK" represents a mixture of sheeps' erythrocytes (E), amboceptor solution (A) which has been sensitised against sheeps' erythrocytes, and complement (K) in suitably adjusted concentrations. The solutions E, A, K, P and Ag usually amount to about 0.25 ml.
To carry out the main test, i.e. the investigation of the patient's serum, the system PKAg is prepared in a test tube and incubated at 37� C. for 30 minutes. This first step of the main test is generally referred to as "binding". The haemolytic system EA is then added and the resulting PEAKAg system again incubated at 37� C. The time of the second incubation is generally also about 30 minutes but varies according to the exact concentration ratios. In cases of doubt, the incubation time required for the second step of the main test can be determined by a comparison experiment carried out on the serum of a healthy subject which is not auto-inhibiting. The haemolytic system (EA) is sensitised by incubation at 37� C. before it is added to the PKAg system.
Another test carried out in the same way as the main test but without antigen is used as so-called serum control. The serum control carried out on the PEAK system tests the patient's serum for any tendency to auto-inhibition since it is only when a serum with auto-inhibition is used that haemolysis fails or is attenuated even when no antigen is present. Additional preliminary incubation tests should be carried out to determine whether the systems (E) and (EA) show signs of spontaneous haemolysis, which would make them unusable for the complement binding reaction. A so-called system control should also be carried out to ensure that the EAK system undergoes haemolysis.
Determination of the degree of haemolysis is carried out visually or photometrically in known manner.
The necessity to prepare the experimental system from a plurality of solutions (6 solutions for the main test, 5 solutions for the serum control test) makes the complement binding reaction even more complicated to carry out then it would in any case be, because of the need to adjust the concentrations and activities of the various reactants to each other. It is well known that a large number of individual pipetting operations carried out in a test leads to experimental errors.
A simplification of the methods for carrying out the complement binding reaction has not previously been possible because the maintests and control tests (PEAKAg, PEAK, EAK, EA, E) in any case necessitate separate delivery of the reactants. There are, however, also other reasons why it was not possible to supply several of the reactants in one combined solution. Since the reactants differ considerably from each other in their stability, correct adjustment of the activities of the different reactants in a combined solution is maintained for only a limited period. Moreover, the reactants react with each other when present in one solution.
The problem therefore arose of providing the clinician or serological laboratory with reliable and simple means of carrying out the complement binding reaction without the complicated adjustments of the activities of the different reactants.
This invention relates to a ready-for-use rapid test package for carrying out serological investigations in which at least two of the following reactants:
1. patient's serum,
2. erythrocytes,
3. amboceptors specific for the erythrocytes,
4. complement and
5. antigen, or antibodies not belonging to (3),
are reacted with each other. The rapid test package according to the invention contains at least two of the reactants 3 to 5 in a solid and preserved form and in quantities suitably adjusted to each other for carrying out the test.
It has now been found that large quantities of haemolytic amboceptor, complement and antigen (optionally antibody) adjusted to each other in their concentrations or activities can be stored in portions in rapid test packages preserved by deep-freezing or freeze drying without the reactants in the various rapid test packages undergoing any significant relative changes in activity in storage. It was also found that the loss of activity of the preserved reactants in a rapid test package is slight.
The development of a new process for carrying out the complement binding reaction is particularly important in this connection in this process at least two of the reactants 3 to 5 are preserved in a solid form and used together in suitably adjusted quantities. The process is preferably carried out as follows: A mixture containing all the reactants required for the reaction in suitably adjusted quantities with the exception of the erythrocytes is incubated for the purpose of the main tests and comparison tests of the complement binding reaction. A quantity of erythrocytes adjusted in known manner to the quantity of the other reactants is then added to the mixture and the mixture is again incubated. The incubation time is in each case about 10 minutes to 2 hours, preferably 25 to 65 minutes.
The main difference between the old process and the new process in the main test is that in the new process it is not the PKAg system but the PAKAg system which is subjected to the first incubation (`binding`). The sensitisation of the EA system by incubation is therefore omitted. The main difference between the old process and the new process in the serum control test is that in the new process the PAK system is subjected to the first incubation but not the PK system. The process described above may, of course, also be carried out with conventionally stored or freshly prepared reactants.
The rapid test package according to the invention is particularly suitable for carrying out serological tests, in particular by the new process.
The rapid test package according to the invention for reactants from groups 3 to 5 generally comprises a container and a sealing cap, both of which may have various forms. The container and preferably also the sealing cap, however, should be chemically inert towards the reactant solutions used. For example, glass containers sealed with the usual plastics stoppers or closure caps used for pharmaceutical packages are suitable. The container may also have several internal cavities for storing the reactants separately.
According to a preferred embodiment of the new rapid test package, it is designed as a disposable package which is used only once and serves also as reaction vessel for carrying out the test. In this case, the new rapid test package is preferably in the form of the test tubes commonly used for carrying out the complement binding reaction and is made of glass or some other transparent material, and it contains the reactants in the quantities suitable for carrying out an individual test.
The reactants in the new rapid test package are preserved in the solid form, in particular by freeze drying or deep freezing at temperatures below -21� C. Freeze-dried reactants are particularly suitable as are also deep frozen solutions of the reactants which are rapidly frozen at temperatures below -30� C., preferably below -55� C. After they have been frozen, the reactant solutions should be stored at temperatures below -21� C., preferably below -40� C. and in particular below -55� C.
If the reactants in the new rapid test package have been preserved by deep freezing, they can be melted before use, e.g. by leaving them at room temperature, and then used in the same way as solutions prepared in the ordinary way, by adding the still missing solutions to them. If the reactants in the new rapid test package have been preserved by freeze-drying, they are brought into solution by the addition of solvents, for example physiological saline solution, or by the addition of other reactants in an excess quantity of solvent.
When individual reactants are freeze-dried, it may be advisable to add an inert carrier, for example human albumen. When individual reactants have been freeze dried, it is not necessary to use a solution with as large a quantity of solvent as would be required when the complement binding reaction is carried out in the conventional manner. In many cases, more concentrated solutions of the reactants can be used, thereby achieving more rational drying. In that case, the freeze-dried reactants, i.e. the residue obtained when a concentrated solution of reactants is freeze-dried, has a lower content of accompanying substances such as inorganic salts, for example, than would be required to prepare a ready-for-use solution of reactants by the addition of distilled water. When solvents are added to freeze-dried reactants it is therefore necessary to take into account the amount of accompanying substances present with these reactants, in particular the amount of inorganic salts. If all the reactants in the new rapid test package have been preserved in the freeze-dried form, it may be advisable, when delivering the rapid test package, also to include a diluent which has been adjusted in its concentration to the proportion of accompanying substances present with the reactants.
The reactants present in a rapid test package according to the invention are generally either all freeze-dried or all deep frozen but there is in principle no objection to using mixed forms of packages in which part of the reactants are deep-frozen and others freeze-dried.
In the case of freeze-drying it is generally desirable to introduce the salts required for the solution into the solvents supplied for carrying out the test because it has been observed that insufficient stability of freeze-dried reactants for the complement binding reaction is, to a large extent, due to the presence of salts and particularly to the presence of hygroscopic alkaline earth metal salts. If these salts are substantially eliminated from the freeze-dried reactants, the stability of the reactants is greatly increased.
In cases where the reactants are preserved by freeze-drying, it is advisable to adjust the individual solutions so that when they are melted and mixed, the resulting solution has a relatively low sodium chloride content, for example 0.85% by weight. In that case, increased resistance of the erythrocytes to haemolysis can to a certain extent be compensated without having to discard the erythrocyte suspension. When using erythrocytes with a normal sensitivity to haemolysis, it is generally desirable to have a total sodium chloride concentration of 0.90% by weight and, if the deep-frozen solution has a lower sodium chloride concentration, it can be adjusted to this value by adding sodium chloride concentrate.
The reactants are preferably kept separate from each other in the new rapid test package. This can be achieved, for example, by deep-freezing or freeze-drying the solutions of reactants in different areas of the rapid test package but it can also be achieved by first freezing one solution of reactant, covering it, for example with a physiological saline solution, freezing this separating layer and finally covering this frozen layer with another reactant solution. This should be carried out in such a way that the reactants do not mix to any significant extent. Separate storage of the various reactants can also be achieved by preserving part of the reactants in the sealing cap of the rapid test package (the term preserving will be used hereinafter to mean either freeze-drying or deep-freezing) and then preserving the remainder of reactants required in the test package, for example at the bottom of the package. There are other possible methods of keeping the reactants separate.
Separate storage of the individual reactants is important if the test package is required to remain suitable for use over a long period. If the rapid test package is to be used within a relatively short time, for example within a few days or up to one week after preparation separate storage is generally unnecessary. The length of time for which a rapid test package remains suitable for use when the reactants are all stored together should be determined in the individual case.
Although it is in most cases advantageous to store the individual reactants in the test package separately from each other because no reactions can then take place during storage, there is no objection to mixing freeze-dried reactants, especially if care has been taken to ensure that the residual moisture content of the freeze-dried substances is low. The residual moisture should generally be less than 1% by weight and preferably less than 0.5% by weight. When reactants are freeze-dried individually, it is also advantageous to store them at temperatures below room temperature, preferably below about 5� C. It is particularly advantageous to store even the freeze-dried products at temperatures below about -21� C., especially if they are intended to be stored for several years.
The concentrations or activities of the individual reactants in the new rapid test package are adjusted to each other in principle (qualitatively) in the same way as has been described above for the conventional process for carrying out the complement binding reaction. As regards the quantitative adjustment of the titre of the reactants, however, proportions different from those employed for the conventional process are particularly preferred. The amount of amboceptor used for the test is preferably at least about 25% higher than the usual amount. It is generally 5 to 16 times, and preferably about 6 to 10-times the quantity of amboceptor which will just cause solution if incubation is carried out for 60 minutes at 37� C. The complement may be used in quantities only about 10 to 25% higher than the complement unit. If desired, the process may also be carried out with the usual proportions of reactants (test quantities) employed for the conventional process, in particular with incubation times of about 30 to 100 minutes. The optimum incubation time can easily be determined.
The new rapid test package may contain the combination of reactants AK, KAg, AAg and AKAg. As already mentioned above, it is in most cases advantageous to store the reactants separately. This applies especially to the storage of K and Ag which have a particular tendency to undergo non-specific reactions when stored together. The reactants in the combination AK and AAg, on the other hand, can generally be stored quite satisfactorily together even when deep-frozen, provided they are not kept for more than about 9 months. If reactants are stored together, however, their keeping quality should be tested in the individual case.
Rapid test packages according to the invention which contain the combination of reactants AKAg (or AK antibody) are particularly important, i.e. combinations which contain all the reactants required for carrying out the complement binding reaction with the exception of the erythrocytes and the patient's serum. There are various possibilities of storing the combination AKAg, namely separate storage of all the reactants A/K/Ag, and partly separate storage in which one of the components is separated from the other two. For partial separation of the reactants in storage, the arrangements AK/Ag and AAg/K are suitable. The arrangement KAg/A is generally not possible owing to non-specific reactions between K and Ag. It may, of course, be carried out in individual cases if suitable preliminary tests in the case of a particular antigen have shown that K and Ag are compatible with each other in storage. The same applies to the combined storage of all three components, which is usually not possible. Moreover, as already mentioned above, reactants which are incompatible with each other can be stored together after they have been freeze-dried separately. Among the various arrangements for storage in the preferred embodiment of the invention which contains the combination of reactants AKAg, the arrangement AKAg and the arrangement A/K/Ag are particularly important.
In practice, the user of the rapid test packages is generally provided with two variations of the rapid test package, namely one for carrying out the main test, which contains the reactants A, K and Ag, and another for carrying out comparison tests, which contains only two of the reactants. The reactants in the two types of rapid test packages are preferably taken from the same batch. Here again the embodiment of the invention in which components A and K are contained in a test tube is very advantageous. Preparation of the package is greatly simplified by the fact that only one type of test tube containing reactants is necessary. This test tube, which contains reactants A and K, is either sealed with a cap containing antigen to produce a test package for the main test or with an empty cap to produce a test package for the comparison test. The test package which contains all three reactants, the antigen being in the cap, may, of course, also be used for carrying out the comparison tests but this would entail unnecessary loss of antigen. When manufacturing the rapid test packages, it may be advisable to prepare caps with antigen for 40% of the test tubes which contain A and K.
If the serum of several patients is to be tested at the same time, several test packages for the main test and serum control may be set up pairwise in a test tube stand in which the various test systems are prepared and incubated. The test packages, may, of course, be supplied in stands for several parallel tests or the individual test systems for carrying out several parallel tests may be rigidly connected together, e.g. in the form of a spot plate. In that case, some of the spot recesses may contain the reactants A, K and Ag and others only A and K. Numerous variations of these arrangements are possible.
The description has so far been restricted to the use of the new test package for detecting certain antigens by the presence of the corresponding antibodies in the patient's serum. The basic principle of detecting the formation of specific antigen-antibody complexes by the consumption of complement can, of course, be used for the reverse process, that is to say the antigen itself can be detected in the patient's serum if a specific antibody for this antigen is available. This method is particularly applicable to serum hepatitis (Australia antigen). The individual test package in this case contains test antibodies instead of the test antigen, for example an amboceptor specific for the Australia antigen in addition to the haemolytic amboceptor.
The rapid test package according to the invention may also be designed with several chambers under one sealing cap, one of which chambers, for example, may be used for carrying out the main test and another for carrying out the serum control. In that case the chambers may, for example, each contain an individual portion of the reactants required for carrying out the test, and the reactants of each individual portion should, of course, preferably be kept separate.
The test package according to the invention may contain an antigen combination instead of a single antigen. This provides a simple method of carrying out a so-called screening test on several pathogens.
For carrying out comparison tests, it may also be advantageous to use test packages according to the invention which, in addition to the components already mentioned above, contain, stored separately, preserved test positive patient's serum, weakly-positive patient's serum or test negative patient's serum.
The rapid test package according to the invention constitutes an exceptional advance in the field of applied serology. It makes an extraordinary large number of serological investigations available to a wide circle of users for the first time. When using the new rapid test package, tests based on the complement binding reaction are no more difficult to carry out than most of the present-day laboratory tests carried out in general medical practice. Screening, that is to say the precautionary routine testing of a large number of patients, therefore becomes possible on a scale which could hardly have been envisaged up to now because of the complexity of the measures required. In particular, the rapid test package according to the invention can improve the diagnostic facilities in areas which have no easy access to large diagnostic laboratories. But, even for the diagnostic laboratory itself the invention provides an important advance since the difficult and time-consuming operations of measuring out the reactants and adjusting their activity can be carried out mechanically by the manufacturer so that the consumer, that is to say the serological laboratory, is relieved of the burden of this very time-consuming labour. It is particularly for laboratories which carry out serological work that the grouping of several test packages in one unit which enables several patients' serum to be tested side by side is of great importance.
Typical examples of embodiments of the rapid test packages according to the invention are illustrated in the accompanying drawings.
FIG. 1 is a cross-section through a rapid test package consisting of a glass container 1 of about 10 cc capacity sealed with a plastics cap 2. The rapid test package of FIG. 1 contains an amboceptor 3 which is specific for sheeps' erythrocytes and a guinea pig complement 4, both deep-frozen. Amboceptor 3 and complement 4 are kept separate from each other by holding the test package at a slant when introducing and deep-freezing the complement and then slanting it in the opposite direction when introducing and deep-freezing the amboceptor. The haemolytic amboceptor is rabbit serum which has been sensitised to sheeps' blood corpuscles in known manner and then complement-inactivated. The amboceptor (0.1 ml) was diluted by 1:250 and had a titre of 1:4000. The guinea pig complement (also 0.1 ml) was diluted by 1:9 and had a titre of 1:90.
Since the complement had a titre of 1:90 (determined on 0.5 ml samples, using 2% erythrocyte suspensions), 0.1 ml of complement diluted to 1:9 correspond to two complement units, that is to say twice the minimum quantity necessary for a complete solution. Since an amboceptor with a titre of 1:4000 was used (the titre being based on 0.25 ml of solution, which together with 0.25 ml of erythrocyte suspension makes up the 0.5 ml of haemolytic system used for standardisation), 0.1 ml of amboceptor diluted to 1:250 correspond to about 6.4 amboceptor units, that is to say 6.4 times the dose of amboceptor which will just cause solution when incubated at 37� C. for one hour.
FIG. 2 shows a rapid test package which, like the package of FIG. 1 consists of a container 1 sealed with stopper 2 but which, in addition to amboceptor 3 and complement 4, contains antigen 5. The antigen 5 is kept separate from the other reactants 3 and 4 in a deep-frozen form in a cavity in the sealing cap 2 which is open to the interior of the package. Amboceptor 3 and complement 4 are kept separate from each other in the same way as in FIG. 1. The quantities of amboceptor and complement and their ratios of concentration and activity correspond to those of FIG. 1. Antigen 5 (0.5 ml) was a commercial cardiolipin antigen 1:150.
Another example of the rapid test package according to the invention is similar in arrangement to that of FIG. 2 but, instead of 0.1 ml of amboceptor diluted as indicated above, 0.1 ml of the same amboceptor solution diluted by 1:6.5 with 0.55 ml of 0.85% physiological saline solution was used.
FIG. 3 shows a rapid test package according to the invention in the form of a reaction vessel for carrying out the test. It consists of a glass test tube 6 with cap 7 basically the same as cap 2 with a similar cavity. The test tube contains 0.1 ml of complement (dilution and titre as indicated above) frozen into the bottom of the test tube at -35� C. The complement is covered with 0.55 ml of 0.85% physiological saline solution 8. The physiological saline solution in turn is covered with 0.1 ml of amboceptor (dilution and titre as indicated above). 0.5 ml of antigen (Reiters antigen 1:8) are frozen in the cavity of cap 7. The rapid test package contains nitrogen as protective gas.
FIG. 4 shows a rapid test package similar to that of FIG. 2 but with the reactants at the bottom of the package separated by a partition 9. Reference numerals 3 to 5 have the same meaning as in FIGS. 1 to 3.
FIG. 5 shows a rapid test package according to the invention in the form of an ampoule. It contains a mixture 10 of 0.1 ml of amboceptor 1:50 and 0.1 ml of complement 1:9 with 0.8 ml of 0.85% physiological saline solution and a solution of 0.5 ml of cardiolipin antigen 1:150 and 0.8 ml of 0.85% physiological saline solution.
FIG. 6 shows a rapid test package according to the invention in which two reactants are stored separately from each other in the cavity of the sealing cap, namely a deep-frozen solution 11 of 0.1 ml of amboceptor 1:50 and 0.4 ml of 0.85% physiological saline solution and another deep-frozen solution of 0.1 ml of complement 1:9 and 0.4 ml of 0.85% physiological saline solution. The test tube 6 contains a solution of 0.5 ml of commercial Reiter antigen 1:8 and 0.8 ml of 0.85% saline solution.
FIG. 7 shows rapid test packages according to the invention combined in a spot plate. Each of the bores 13 in a polymethacrylate plate 14 constitutes a rapid test package according to the invention. Each bore contains separately stored reactants arranged as in FIG. 1.
FIG. 8 shows a rapid test package according to the invention which is similar to that of FIG. 4 but contains freeze-dried amboceptor 15 and freeze-dried complement 16. Freeze-drying was carried out by taking up 0.1 ml of complement (dilution and titre as above) with a solution of 0.0025 g of human albumen in 0.5 ml of water and introducing it into one of the chambers formed by partition 9 inside the package. The method was repeated with 0.1 ml of amboceptor solution (dilution and titre as above), using the other chamber. Freeze-drying itself was carried out in known manner at an initial temperature of about -50� C.
The rapid test packages according to the invention have been illustrated above with the quantitites of reactants which are normally preferred for carrying out an individual test. In the complement binding reaction, however, the absolute quantities of the reactants are not important, provided that the quantities of the individual reactants are adjusted to each other. An individual test can therefore in principle be carried out with any multiple of all the reactants. Rapid test packages for micro tests (up to 0.1 ml total volume of all the reactants) are therefore in principle the same as those of FIGS. 1 to 8 but contain, for example, one twentieth of the quantities of reactants. A flat watch glass shaped dish as shown in FIG. 9, for example, is particularly suitable for carrying out tests on a micro scale. The dish 17 has a diameter of about 2 cm and contains amboceptor complement and antigen as deep-frozen drops 18 to 20 of their solutions in quantities suitably adjusted to each other for carrying out the complement binding reaction. The dish is covered with a piece of plastics adhesive tape 21. Spot plates are also very suitable for this purpose. Just as the quantities of reactants in the rapid test package can be reduced in scale, so also larger quantities may be used in a package.
FIG. 10 shows a rapid test package which contains 50 ml of amboceptor 22 (dilution 1:1250, titre 1:4000) and 50 ml of complement 23 (dilution 1:45, titre 1:90) in a glass reagent bottle 25 sealed with a plastics stopper 24.
All the packages of the examples with the exception of the package shown in FIG. 8 contain the reactants in a deep-frozen form.
The new process for carrying out the complement binding reaction and method of handling the rapid test package are explained by the following examples with reference to the rapid test package shown in FIG. 3.
To carry out the main test, a test package according to the invention is taken from the deep-freezer and thawed by immersing it in a water bath at 37� C. Thawing forms the solution AKAg to which 0.25 ml of patient's serum (method of obtaining see above) are added. The resulting solution of PAKAg is then incubated at 37� C. for half an hour. A binding reaction takes place if the patient's serum contains antibodies, in this case syphilis antibodies. The complement is used up with formation of antigen-antibody complex. This step of the process differs from the corresponding step in the conventional method of carrying out the complement binding reaction in that the haemolytic amboceptor is already present during the binding reaction.
After the half hour's incubation period, 0.25 ml of the 2% sheep's erythrocyte suspension are added to the system and the system is again incubated at 37� C. The incubation time in this second step is also half an hour but depends on the exact adjustment of the reactants to each other and since this is carried out by the supplier of the individual package, directions should be supplied with the package as to the correct incubation time. The time is generally 30 minutes but can be determined by a comparison test with non-inhibiting serum from healthy subjects.
After the second incubation period, the result, i.e. the degree of haemolysis of the preparation, is assessed visually or photometrically in known manner.
Comparison tests are virtually essential for carrying out serological investigations. The comparison tests described above can be carried out very elegantly with the individual test package according to the invention. The control tests are also carried out by the new process as follows.
To carry out the serum control (auto-inhibition test) an individual test package is taken from the deep freezer and the sealing cap which contains the antigen is removed. The contents of the test tube are then melted in a water bath at 37� C. to form liquid solution AK. The same quantity of patient's serum as in the main test is added to this solution and the solution is madeup to the same volume as in the main test by the addition of physiological saline solution. To keep the reaction conditions as close as possible to those of the main test, the resulting system of PAK is then incubated at 37� C. for half an hour and then incubated for a second time after addition of the erythrocyte suspension in the same way as in the main test. The interpretation of the result is carried out in the same way as in the conventional serum control test.
System control in system EAK is carried out on the same principle as the serum control. The cap containing the antigen is removed but no patient's serum is added.
There remain the control tests in system EA and E. The erythrocyte suspension E can, of course, quite easily be tested for spontaneous haemolysis under incubating conditions but the individual test package according to the invention is not necessary for this. To test the system EA which must not show any signs of haemolysis under incubating conditions, the sealing cap which contains the antigen is removed and the contents of the test tube are melted in a water bath at 56� C. and kept at this temperature for 20 minutes. The remaining solution A is cooled to room temperature, erythrocyte suspension is added, and the system is then incubated at 37� C. for about 30 minutes. The test on system EA can generally be omitted because spontaneous haemolysis, that is to say haemolysis in the absence of complement, is extremely rare in this system. A test carried out on system EA to test the amboceptor for the presence of traces of complement is unnecessary because the amboceptor has, of course, been tested by the manufacturer.
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|Miles Lab
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|Merck & Co Inc
|Method of producing a multicomponent lyophilized product
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|K Bagshawe
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|Citing Patent
|Filing date
|Publication date
|Apr 2, 1982
|Sep 11, 1984
|The Board Of Trustees Of The Leland Stanford Junior University
|Method for HLA-DR typing of total human lymphocyte sample
|Apr 5, 1982
|Jan 8, 1985
|Duke University
|Complement assay method
|Oct 1, 1985
|Oct 14, 1986
|Bert Keenan
|Petroleum alcohol test kit and method of testing petroleum for alcohol content
|Jun 4, 1984
|Apr 14, 1987
|The Board Of Trustees Of The Leland Stanford Junior University
|Method for B cell typing of total human lymphocyte sample
|Dec 8, 1986
|Mar 6, 1990
|Novo Industri A/S
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|Serres Pierre F
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|Novo Industri A/S
|Methods of treating disorders which cause conglutinin deficiency
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|Zenith Electronics Corporation
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|Oct 2, 2003
|Mar 4, 2014
|Novartis Ag
|Polypeptide-vaccines for broad protection against hypervirulent meningococcal lineages
|Oct 2, 2003
|Aug 3, 2006
|Chiron Spa
|Polypeptide-vaccines for broad protetion against hypervirulent meningococcal lineages
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|Neisserial vaccine compositions comprising a combination of antigens
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|Veterinary-medical kit for a rapid test to determine the calcium content of blood
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|Calgon Corporation
|Method and device for enumerating microorganisms
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|Indicia Diagnostics
|Method and apparatus for the determination of immunologically reactive substances of clinical interest
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|Miles Inc.
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|Sep 16, 1997
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|U.S. Classification
|435/5, 435/7.25, 436/522, 435/7.36, 435/7.22, 422/913, 436/808, 435/7.32, 435/7.1, 435/7.2, 422/942
|International Classification
|Cooperative Classification
|European Classification


text:
Vita Brevis
Vita brevis,
Ars longa,
Occasio praeceps,
Eperimentum periculosum,
Iudicium difficile.
Life is short,
the art of healing is long,
opportunity fleeting,
experience perilous,
and decision difficult.
4 Years Old
"Mommy isn't feeling well today, Katniss." Daddy tells tiny, brown-haired Katniss Everdeen as he helps her into her red coat. She loves her red coat and she loves it when Daddy takes her someplace special, just the two of them. "We're going to go to the zoo so she can sleep."
The innocent, little girl smiles up at Daddy as he takes her hand, leading her to his old truck, the one that still has a cassette player in it. He always lets her choose which tape they listen to, and she always chooses the one with a funny looking skull holding a rose. He likes to sing to that one and she likes hearing Daddy sing.
"Is Mommy sick?" Katniss asks as he buckles her into the car in her special seat.
"Mommy is just very tired today," he answers, brushing a hand over her dark hair. He tried his best to braid this morning, but Daddy can't do it like Mommy does so it's very messy. It's been messy every day for a very long time now.
Katniss nods solemnly in understanding. Mommy gets very tired sometimes and doesn't come out of her room for a many, many days. Daddy talks to Mommy very quietly when she gets tired like this, but Mommy never answers. Katniss sometimes sneaks into her room and sits next to her. She likes to brush her hand through her mother's pretty yellow hair. Mommy doesn't talk to her either, but sometimes Katniss likes to pretend what she might say when she tells her stories about the day.
Katniss picks the music and Daddy sings the entire way to the zoo. His voice is so very pretty that she hardly notices the hum of the truck's old engine as his deep, baritone voice fills the cab. When Daddy parks the car, Katniss unbuckles her seat all by herself while he walks around to open her door.
Putting his hands on her hips, he prevents her from jumping out of the truck for a minute. "You know I love you very much, right Katniss? And Mommy loves you so, so much too."
"I love you too, Daddy," Katniss giggles, reaching up and wrapping her arms around his neck. He lifts her out of the car and carries her through the parking lot. Mommy would say she's too big for him to carry, but today it's just Daddy and her, so he does it anyway.
"You want to go see the reptiles first, right Katniss?" Daddy asks her with a grin that makes the dimples in his cheeks appear and the corners of his eyes crinkle.
She wrinkles her little nose at the suggestion. "No!"
She laughs and he does too. He knows that the snakes and lizards aren't her favorite animals to see at the zoo. She knows he doesn't like them best either. Both Everdeens have always had a fascination with the big cats- the tigers and leopards and lions and cougars.
"Oh, well, I guess we could go visit the snow leopards this morning, if you really want," Daddy shrugs, bouncing Katniss on his hip playfully as he heads in the direction of the Big Cats exhibit.
Katniss loves spending days like this with Daddy. He works a lot but he always tries to make time for her on the weekends, which is why, even at the tender age of four, the weekend is already her favorite part of the week.
They spend the morning watching the lazy cats wander around their pens, rolling in the early sunlight and chewing on the bones of their breakfast. Katniss holds Daddy's hand the entire time, not because she's afraid of getting lost because she has always been independent for a child of her age, but because she misses the gentle, warm feel of his skin against hers when he's not there.
By the time they get home, the sun has already started its descent in the sky. Katniss can barely keep her eyes open and Daddy tucks her under a blanket on the couch, whispering against her hair. "Take a nap, Katniss. I'll be here when you wake up."
She drifts off quickly, the sound of his soft footsteps lulling her into a sweet, dreamless sleep as he leaves the room.
Through a sleepy haze, Katniss wonders why Daddy is shouting. He is never one to shout, even when she gets in trouble. She buries her face against the soft cushion of the couch, not ready yet to wake up after the exertion of the morning.
"Lily! What did you do, sweetheart?"
Her eyes crack open against the late evening light at the frantic tone of her father's voice. Katniss doesn't think Daddy has ever sounded quite so afraid before, his voice cracking and shaking as he continues to speak more quietly to Mommy, but it doesn't stop his voice from reaching her downstairs.
She crawls off the couch, carrying the blanket with her up the stairs to stand outside her parents' bedroom.
"Lily, how many did you take?" Katniss watches as Daddy fumbles around on the bedside table, his trembling hands lifting the little orange bottles one after another until he finds an empty one. "Lily, how many were left, dear?"
Mommy doesn't answer him, staring blankly into the space between them instead. She looks strange; her lips are a funny color. Daddy looks so scared as he shakes her shoulders that it makes Katniss scared too. She doesn't understand why she feels so cold all of the sudden. She doesn't know why she can't get her legs to move her tiny body into the room to stand next to him, to pat his hand like she usually does when he's upset.
So instead, she does the only thing she can think to do- the very thing she is best at- she stays very quiet and huddles down against the doorframe, wrapping the blanket around her in hopes that it might help warm her. She doesn't want to distract Daddy, but she doesn't want to be alone anymore either.
Katniss wishes very much that she could be braver; that she could help her daddy and her mommy, but she's just so very afraid. She doesn't understand why Mommy is so quiet tonight. Even on her worst nights she will sigh or look at Daddy. She wishes that Mommy would say something, anything, to him so he might not be so scared anymore.
"Just remember that Mommy is sick right now, Katniss, so she might not want to talk very much, okay?" Katniss nods, swinging her hand in her father's as they walk through the long hallways of the hospital, at least Daddy told her this is a hospital. She's not quite sure since this place doesn't look much like hospitals she sees in her picture books.
It's been a long time since that day at the zoo; long enough that Katniss had to turn the calendar that hangs above the phone, the one that Mommy uses to write appointments on. The men in the ambulance came and took Mommy away that day in a flurry of voices and flashing lights. Katniss has noticed how sad Daddy has been since then and she hopes Mommy will come home soon.
A big man unlocks the door and lets them into a giant living room. Katniss spots Mommy right away, sitting on a couch with her legs tucked under her. She tugs nervously at one of her dark, messy braids that Daddy fixed her hair in, suddenly shy at the sight of her mother. Mommy is wearing light blue pajamas that Katniss doesn't recognize but she doesn't think Mommy has ever looked prettier.
"Hello, Katniss," Mommy smiles, holding out her arms so she can wrap her in a hug. Her tummy flips happily. She hadn't realized just how much she missed Mommy and her voice and her touch.
"Hi, Mommy," she grins back toothily. Mommy doesn't look very sick anymore. Maybe she'll be home again soon.
"How was your day?" Mommy asks, nestling Katniss to her side.
She immediately begins to tell her mother about how Daddy took her to the park where there is an ice skating rink right now. She likes the way Mommy smiles easily. She missed her mommy's smile and she missed the way Mommy is holding her now. Katniss makes a silent promise that she will try extra hard to be good so as not to make Mommy sad so she won't leave again. She wraps her tiny arms tighter around Mommy's waist in hopes that this might be able to keep her with them forever.
"You're quieter than usual today, Katniss," Thom says as she passes by him on the way out of the med room, "What's eating you?"
"It's more of a problem with who isn't eating her if you ask me," Johanna offers offhandedly, never looking away from the charting on the screen before her.
"Thanks for the input, Jo," Katniss spits back, hoping Thom doesn't pick up on the blush quickly rising to her cheeks.
Johanna always assumes everything revolves around sex, which for the older woman, it usually does. Katniss sometimes wishes her life could be that simple. This morning, she got a call from Greenfields, which always throws her off, but Johanna is at least correct in her assertion that Katniss isn't bedding anyone regularly these days. Honestly, she has more important things to be thinking about than plotting ways to keep a guy between her thighs.
"Just thinking about stuff, Thom. No big deal," Katniss says with a passable smile. "I put a page out for Doctor Aurelius, so if he calls back, forward him to my phone," she adds, heading for the room where the antibiotic in her hands is supposed to be hung up.
"Good morning, ladies… and Thom," Dr. Odair greets the trio as he walks off the elevator next to the nurses' station, flashing a bright smile at Katniss as she eyes him curiously, pausing on her way to see her patient to hear why in the world Dr. Finnick Odair is visiting the floor today.
"Haven't seen you up here in a while, Doctor," Johanna comments from her seat in front of the computer as Katniss fiddles with the tubing of the antibiotic.
"That would be because I prefer spending my time with the pediatric patients downstairs," the young doctor admits, a sparkle in his eye, "They understand me. Plus, they share their toys with me."
"So what brings you up here today then?" Katniss asks, flicking at a bubble in the line. "We still don't have any toys."
He laughs, pulling a hand through his bronze hair, leaving it an unruly mess, "I'm covering for Dr. Aurelius today until his new hospitalist can get in and one of you put a page out for him. So here I am."
Katniss recalls hearing something about the new hospitalist a few weeks ago. Dr. Aurelius is one of the best doctors in the city, and lately he's been swamped trying to keep up with appointments at his office on top of doing rounds at the hospital. Hiring a hospitalist was the obvious solution since it will free up more of his time to be at the office. The new hospitalist will take over Dr. Aurelius's physician rounds, taking care of any patients that he is normally the primary physician for. It has become an increasingly popular option for physicians at Capitol Memorial to hire hospitalists, or at least nurse practioners, to help with their patient loads.
"You're making trips to every floor that pages you personally?" Katniss arches an eyebrow curiously at the young man.
"Only for the lovely nurses like yourself, Everdeen," he quips, shooting a suggestive wink her way. She lets out a snort in spite of herself. Dr. Odair has been a flirt since the day he was hired at Capitol Memorial, but she's come to recognize he's harmless, especially when compared with some of her patients. Somehow he always manages to make her smile, even on days like this when she's so preoccupied with everything going on in her personal life.
"So were you the one to page me?" he asks in his most seductive tone.
"Forget it!" Katniss throws her hands up, the tubing of the antibiotic swinging haphazardly between them as she laughs, backing away from the bronze-haired doctor, "I'll just write orders and lose my license for it. I can't work with you today, Doctor."
"Oh, come on now, Kit Kat," Dr. Odair chuckles playfully after her. "Your colleagues won't tell on us if they discover the true relationship between us. I'm sick of pretending your disdain for my affections is real."
Katniss scurries away from the nurses' station, making a gagging noise as she hurries off to hang the antibiotic before the dose will be considered late. When she returns several minutes later, Dr. Odair is still at the nurses' station, leaning lazily against the counter as he chats with Johanna and Thom.
"So fill us in then, Odair," Johanna turns in her chair with a smirk, "What's the new doc like?"
"A completely pompous asshole," Finnick answers easily, folding the papers in his hand with a grim look, "Thinks he knows everything, looks at nurses and everyone else as his underlings, expects everyone to fall all over themselves to keep him happy-"
"So, basically, he's you, right Doc?" Thom retorts boldly from where he's sitting on hold with the local Walgreens, trying to get information on a newly-admitted patient's home medications.
"That's offensive, Thom!" Finnick shouts over the laughter that breaks out from everyone at the nurses' station although he's biting back a smile himself, "Everybody knows I'm the best-looking and most lovable doctor around. You'll all love me even more once you meet the new one."
"Oh, please, Finn," a deep voice calls, edged with humor, from just beyond the nurses' station. Katniss turns just in time to see a broad-shouldered, blonde-haired man step off the elevator. "Don't convince them that they hate me before we even meet."
Finnick's face breaks into an even brighter grin as he turns to greet the hospital's newest physician, extending a slender hand, "Peeta! You're here earlier than Dr. Aurelius thought you would be."
"It was a pretty quick stop by his office and I didn't see the point in waiting to come in," the new doctor says with a shrug and a shy smile. "I thought it might be nice to meet some of the staff I'd be working with."
Finnick nods at the group of nurses before him, "This motley crew is the group of nurses you'll be relying on not to kill your patients on the sixth floor." Finnick points a thumb towards the handsome man beside him. "Everyone, this is Dr. Aurelius's new hospitalist Dr. Peeta Mellark."
Peeta nods at the group before him, giving an awkward wave. "Nice to meet you all."
"Here's your list for the day, Peeta," Finnick says, gladly shoving the papers he printed only moments before into the blonde man's hands. "I'm going back to pediatrics where I fit in. Everyone up here is old and grumpy."
"At least the patients up here can tell me what's wrong," Peeta chuckles, clapping the other man on the back as he looks down at the names on the sheet in his hand.
Finnick shrugs, knowing that the blonde man has a point. "Katniss is the one who paged me up here, so you'll probably need to talk with her first. She's a hard shell to crack though, Peeta, so don't beat yourself up too hard if she doesn't like you." Finnick flashes her a grin. "I won her over with my stunning good looks, but not everyone is so lucky."
Dr. Mellark glances in the direction that Finnick is looking and a blush rises along his neck as his eyes meet the curious grey ones a few feet away. Katniss, for her part, maintains a neutral appearance even though her heart is suddenly pounding against her chest. She doesn't think she has ever met a person with eyes quite so blue, the color rich enough to get lost in for hours.
Finnick says his goodbyes, waving as he heads for the elevator, but Katniss has a difficult time tearing her gaze away from the beautiful man before her. She's not sure when he moves, but all at once he's standing directly in front of her, his head tilted down so their gazes still meet.
Up close, she realizes just how big Dr. Peeta Mellark is. He's not so much tall as he is stocky, muscular, wide. He looks like the type of man who might play football, hockey- some sport where there's enough contact that his size would be beneficial to throw around. Katniss wonders how in the world a man of this size ended up going through medical school. Almost every other doctor she knows is barely a few inches taller than her at best and slightly built.
It's not until he clears his throat that Katniss realizes, a bit belatedly, that she's been staring in silence for an awkwardly long time. She sticks out a hand, realizing once she does that it's sweaty and probably not going to give the best impression when she's hoping to convince this man that she's a confident nurse.
"Nice to meet you, Dr. Mellark," she says, trying to offer a pleasant smile, but she's afraid it comes across as more of a grimace. Katniss has never been good at pleasantries or introductions.
He doesn't seem to notice, or if he does, he's kind enough not to point it out as he takes her hand, his own large hand engulfing hers. His hand is also dry and just that comfortable temperature that isn't too cold. Her eyes narrow of their own accord, wondering how on earth this man can be so freaking perfect and how his grip sends electric tingles traveling up her arm.
"Nice to meet you as well… Katniss? Is that the name Finn used?" Dr. Mellark squints at the name badge hanging from the collar of her navy scrubs.
She nods, "Yes. We're a family full of flowers." She cringes. Why the hell did she just tell him that?
Dr. Mellark's lips break into an easy smile though as a sweet chuckle escapes from deep in his throat, drawing a shy smile from Katniss in spite of her embarrassment.
"Good to know," he nods amiably as he reaches into the breast pocket of his white medical coat, pulling out a pair of black-framed glasses and placing them on his nose. They somehow seem to enhance the color of his eyes and Katniss finds herself mesmerized, "So Finn said you paged for Dr. Aurelius?"
Right. Work.
She gives her head a little shake and looks down at her nails, "Yes, I got in report this morning that Mr. Strider was supposed to be going home today, so I was hoping to get orders and reconcile medications."
Dr. Mellark gives an easy smile. "Oh, sure. I'll just have a look at him first and then come back out here to do the paperwork. How's Mr. Strider doing today?" he asks, pulling a sheet of hand-written notes from a pocket of his coat and looking it over briefly, the frames of his glasses sliding down the bridge of his nose ever so slightly.
"He's good," she replies dumbly, tugging anxiously at the end of her braid. She's not sure why her nerves are so raw right now. It's not as though this man is threatening or intimidating, at least not in the way most physicians are. Physically he's pretty massive, but she can already tell he's kind and easygoing in a way most doctors aren't with nurses.
A soft chuckle catches her attention, causing her to look up again and meet those stunning eyes. Dr. Mellark is smiling at her, his eyes crinkling at the corners, "I'm going to need a little bit more to go on before I'm ready to discharge him."
Blood rushes to her cheeks and she ducks her gaze back to her hands. How embarrassing! And he's still smiling at her like she hasn't just made a complete fool of herself. She wishes she could just crawl under the desk at the nurses' station and hide there until Dr. Peeta Mellark leaves for the day.
"How are his lungs sounding?" he prompts, raising a pale eyebrow over the dark frame of his glasses.
Her heart begins to race again and Katniss tries to remember if she put a nicotine patch on a patient without using gloves this morning. The last time she did that she thought she might be having a heart attack. It felt a lot like she's feeling right now as her eyes find his vibrant ones again.
"Pretty shitty-" Katniss stops, her eyes widening before she corrects herself. "Or- uh, coarse. They sound coarse, I mean. They are improving though."
Why wouldn't the floor just open up and swallow her whole? Katniss hears Johanna's amused snort from behind her and it really must be as embarrassing as Katniss thinks if Jo's laughing about it.
Dr. Mellark laughs loudly though, apparently oblivious to her lack of professionalism, "I'll bet they don't sound so good. I looked at the x-rays from yesterday." He gives her a conspiratorial wink before folding the papers up and slipping them back into his coat. Maybe he's not so oblivious after all. Not that that's any comfort at this point. "How are his sats this morning?"
"Ninety-six on two liters." Katniss answers more readily this time, clearing her throat in an attempt to keep her voice from shaking now that she's thoroughly embarrassed herself. "They had to bump him up to three overnight but I weaned him down this morning, and he's been okay with that."
"Have you tried taking him off the oxygen completely yet?" Dr. Mellark questions, rubbing a hand over the back of his neck.
"Not yet. He was working with physical therapy earlier so I didn't want to do it then." Katniss shuffles awkwardly from one foot to the other.
"Okay," he nods, his blonde hair capturing the light as he moves in the direction of Mr. Strider's room, "Why don't you give it a try sometime today before he leaves and if he desats below ninety let me know? I'll write the discharge orders and do the meds before I leave, but we can keep him another day if he needs the oxygen. Otherwise I'll switch him over to an oral antibiotic and have him follow-up with Dr. Aurelius in two weeks."
"Sure," Katniss says softly, doing her best to smile confidently at him but she's not sure it's very convincing. Judging by the amused look he gives her, she figures he agrees.
"So you have the hots for Dr. Blondie," Johanna says, collapsing into the seat next to Katniss at the break room table.
"What the hell are you talking about, Jo?" Katniss mumbles through a mouthful of sandwich, rolling her eyes at the stunning, dark-haired woman beside her.
"I'm talking about the way you turned into a stuttering, idiotic, puddle of mush as soon as that broad-shouldered god started talking to you," she fixes the younger woman with a shit-eating grin. "Even you aren't that socially stunted with the average person, Katniss."
"And you're fishing for gossip," Katniss replies, shaking her head as she stuffs more sandwich into her mouth.
"Okay, if you want to go for denial that's fine," Johanna shrugs. "But that man is ridiculously attractive and you haven't had a good fuck in ages-"
"Just saying!" She holds her hands up in defense with a little cackle. "I would just go for it. You're young and unattached and you deserve some happiness in your life, Katniss. I don't think people tell you that enough so I'm saying it for them."
"I just have a lot going on right now and the last thing I need is to get involved with a physician," Katniss mutters, picking at the crust of her sandwich.
Johanna gives her a knowing glance. "How's she doing?"
Katniss looks up in surprise but Jo simply shoots her a wry smile.
"You always get quiet when she gets bad again. You might not say anything about it, but you're not that hard to read, Katniss," Johanna shrugs. "Is there anything I can do?"
"No," Katniss admits, her voice barely louder than a whisper, "I'll be okay."
Both girls share a look, one that says they both hope her words are true. Katniss's nerves are wearing thin lately and she doesn't know how much longer she can keep doing this. But for now she just has to buck up and make do with what she has to work with.
Directly after Katniss finishes her shift at the hospital, she jumps into her old beat-up sedan and makes the drive out to Greenfields. She can't help but think that the name is an apt description of the establishment as she turns down the familiar drive surrounded by the large open areas of what would be manicured, green grass and trees if it weren't the dead of winter.
She finds her way to the unit easily, the path ingrained deeply in her memory at this point as she passes through the hallways washed in soothing colors with tranquil paintings hanging upon them.
"Hi there, Katniss." Cinna greets her warmly when he opens the heavy, metal door after she rings the bell.
She returns his gentle smile as she steps through the doorway and follows him down the short corridor. Cinna swings his badge around his finger carelessly as they approach a black keypad that he waves his ID in front of before a high-pitched beep sounds. A loud click echoes and Cinna pushes the door open, following Katniss through it as she walks onto the unit.
"How is she doing?" She asks softly, rubbing a hand nervously over the sleeve of her jacket.
"Better since we called you this morning," Cinna nods smoothly as he lets them through the door and locks it behind them once again. "The Haldol calmed her down. She's just really quiet right now like she always is after an episode."
"Is she still talking about leaving?" Katniss tugs at the end of her braid.
Cinna looks at her sympathetically, "She's still mumbling to herself about breaking out of here. She thinks the government is holding her here to keep her quiet. So pretty much the usual."
Katniss sighs heavily, pressing her fingers against her eyes. It's the same cycle, but after this many years it's really starting to wear on her. Cinna places a gentle hand to her back. "She's in the rec room."
"Thanks, Cinna." She gives a halfhearted smile, "I'll talk to you before I leave for the night, okay?"
The dark-skinned man nods gently, returning to the nurses' station and settling down at a computer to chart. Katniss waves at Portia, who is in the med room pulling out evening medications and settling the tiny paper cups onto a tray. Katniss makes her way to the rec room, which is separated from the main visiting area by a small half-wall. It only takes a moment to spot the head of blonde hair sitting just beyond a table where three patients are quietly working on a puzzle.
"Hi, Mama," Katniss greets quietly as she walks up behind the rocking chair that's facing the window, looking out at the dark courtyard of the psychiatric facility.
She moves around the chair and settles herself carefully on the windowsill, looking up at the woman who raised her. Her mother's eyes flit to her face for a short moment before returning their focus to the darkness outside. Observing the tired face that seems to have aged twenty years practically overnight, Katniss can't help but watch the older woman with a wistful look.
Katniss always found her mother beautiful, but her illness has started to take its toll of late, really ever since Daddy passed. He was always the string that held her to reality and grounded her, but when he died, she spiraled out of control in a way that neither of her daughters seemed able to help.
The depression was bad enough; Katniss lived in constant fear that she would come home from her shift one evening to her mother and a handful of empty pill bottles. But when the manic phase took hold, her mother became dangerous. She wasn't just a hazard to herself anymore when she felt invincible, and she was liable to ruin the household finances as well.
When Katniss came home one day to find a towel on fire on the stove it was the last straw. Mama told her she had been cooking a feast but got distracted by the news report. There was a message hidden in the words, she said. The government was trying to communicate with her.
That first time after Daddy passed, it was impossible to convince her mother that the hospital was what she needed, that they would give her medicines that would make her feel better. The court only took ten minutes to decide that she was a danger to herself and society if she wasn't in a locked facility. Since that first time, Katniss has become quite at home at Greenfields Psychiatric Facility. She tries to make it out here at least once a week when her mother is here.
"Mama?" Katniss repeats after a few minutes of silence, "Will you talk to me?"
There's a long pause before Lily turns to look at her eldest daughter, "You have to get me out of here!" she demands, her wide, blue eyes frantically searching Katniss's face.
"They're giving me drugs to keep me quiet because of what I know-"
"Mama, stop it-"
"You can't leave me here, Katniss, they'll kill me." The words tumble out of her mother's mouth so quickly that Katniss doesn't have a chance to stop her. "We can escape! I have a plan that won't fail!" Her mother reaches for her hands, clasping them desperately, bouncing her heel quickly against the floor, unable to remain still as the mania overtakes her again.
Katniss sighs, "Mama, no one is trying to hurt you. You're in the hospital. They're trying to make you feel better."
"I feel fine!" she argues fervently. "Or I did until they gave me that shot today. Look at it." She raises the sleeve of her t-shirt to show the swollen area on her upper arm where the needle had bruised the tissue when they had forcibly administered the shot of antipsychotic earlier this morning.
"Don't you see?" she pleads, moving to stand, wrenching her hands through her stringy, blonde locks. "They're putting trackers in me. If we don't get out of here soon-"
"Mama, stop it!" Katniss shouts, clasping her shaking hands together as she stands. "I'm not taking you out of here. You need to be here. Don't you remember what happened last time?"
"I was fine before I came here!" the older woman maintains but Katniss is shaking her head even before she can finish.
"Mama, you almost drove your car off a bridge-"
"I did not!" she hisses, her offense hard to miss. "I was trying to deliver an important piece of research and the cops were chasing me."
Katniss shakes her head, taking a step back. Talking to her mother when she's like this is pointless. No matter how logically Katniss tries to argue with her, her mother will never be convinced that her delusions are just that- delusions.
"Why are you shaking your head like that? You know I'm right, Katniss!" Lily shouts angrily.
"Lily, here's your medicines for the night." Portia holds out the little, white paper cup.
"I don't want those!" She swats a frustrated hand in the air, nearly knocking the cup from the other woman's hand before turning back to Katniss, fixing her with a pointed glare. "You see. They're trying to drug me."
"You need to take those pills, Mama," Katniss warns, trying her best to soothe the older woman. "Those pills help you."
"I won't take them!" she shouts, backing away, a look of betrayal plastered on her face, "And you can just leave if you're going to take their side, Katniss. I don't need you here! I don't want you here, so just go!"
"Get out! Leave me alone!" she screams as two nurses' aids suddenly appear, guiding her away down a hallway that leads to her room. The shouting doesn't stop as she walks away though. Her words quickly revert back to accusations about the government, about escaping this place, about having an important message to deliver.
Katniss lets her body sag, settling into the rocking chair where her mother was only a handful of minutes before. Her head settles back against the headrest and she closes her eyes, barely hearing Cinna's quiet tread as he walks up to join her.
"Hopefully tomorrow will be better," he offers. Katniss opens her eyes to see him shoving his hands into his pockets, a grim look.
"But it probably won't, right?" Katniss asks, already knowing the answer. They've been through this before after all.
"Probably not." Cinna sighs, reaching out to cover one of her hands with his.
"When will the judge be here again?" she asks, "So I can switch shifts if I need to."
"Thursday," he replies gently. "You know that your sister could be a family witness as well. It doesn't have to be you every time, Katniss. That would be a lot to ask of anyone."
She shakes her head, letting her forehead fall into her hands. "Prim doesn't need this stress. Besides, it's harder for her to get home now that she's started med school rotations."
"Okay," Cinna nods understandingly, "we'll get an appointment for Thursday set up then. I'll call you with a time once we get one."
"Thanks, Cinna." Katniss shoots him a weak smile before standing, cracking her back tiredly. Seeing her mother like this always ages her about ten years. "I guess I'll see you Thursday then."
They walk back the way she came and Cinna lets them through both locked doors, giving her a smile as she steps out into the main hallway of the facility.
"Take care, Katniss." Cinna waves goodbye. "We'll be in contact sometime tomorrow."
"Thanks," Katniss waves, letting herself out of the building and hurrying through the cold night to her car. She lets her forehead fall against the steering wheel, taking several deep breaths to steady her nerves before the drive home. It's times like these that Katniss misses her father more than anything. He always knew how to talk to her mother when she got like this. Sometimes she just doesn't know what to do anymore. Sometimes Katniss just wishes there was someone else there who could share this burden. She just feels so lost, so small, that she's not sure how much longer she can keep this up.
AN: Thank you all so very much for reading! Please feel free to let me know what you thought! I love hearing from you. An extra big thanks to Court81981 for beta-ing this chapter and whipping into readable shape for you all.

subset: commoncrawl


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On The Road With Buddy 19Favorites [+]
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Running Time: 97 minutes
Released: 01/2005
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Series: Erotic Street Life
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Russia–Seychelles relations
Russia–Seychelles relations (Russian: Российско-сейшельские отношения) is the bilateral relations of Russia and Seychelles. Diplomatic relations between Seychelles and the Soviet Union were established on 30 June 1976, a day after the island nation gained its independence from the United Kingdom.[1] Russia has an embassy in Victoria.[2] Seychelles is represented in Russia through its embassy in Paris (France) and two honorary consulates (in Saint Petersburg and Yekaterinburg).
In February 2009, Alexander Vladimirov, Russia's Ambassador to Seychelles paid a final visit to President James Michel and Vice-President Joseph Belmont at the State House.[3]
Soviet-era relations[edit]
On 15 February 1980, the Soviet Union and Seychelles signed the Agreement on merchant navigation in Victoria.[4] The government of Seychellois President France-Albert René supported the Soviet invasion of Afghanistan.[5]
In 1987 The Sunday Times, quoting unnamed US intelligence officers, reported that the Soviet Union had landed 50 naval infantry troops in the Seychelles after making landfall on the Ivan Rogov in October 1986; a month after a foiled assassination attempt on René.[6]
Economic relations[edit]
Bilateral trade between Russia and the Seychelles in 2008 reached a total of US$6.23 million. Russia exported some US$4.54 million worth of goods and services to the Seychelles, including mineral oils, machinery and equipment. The Seychelles exports to Russia totalled US$1.69 million made up of fish, seafood and spices.[7]
In 1999 an agreement on co-operation in the field of tourism was concluded between the two nations, and in 2008 some 6,400 Russian tourists visited the Seychelles.[7] Burgeoning Russian tourism to the Seychelles led the Seychellois national airline Air Seychelles to launch a weekly flight to Moscow's Vnukovo International Airport in March 2009, with the expectation of the addition of another weekly flight once its presence in the Russian market has been consolidated.[8]
See also[edit]
- ^ Ginsburgs, George (1987). "1976". A calendar of Soviet treaties, 1974-1980. BRILL. p. 163. ISBN 90-247-3628-5. Retrieved 2009-05-27.
- ^ Посольство в Виктории (in Russian). Ministry of Foreign Affairs of Russia. Retrieved 2009-05-27.
- ^ "Seychelles-Russia relations now 'at their best'". Seychelles Nation. 11 February 2009. Retrieved 2009-05-27.
- ^ "Agreement on merchant navigation. Signed at Victoria on 15 February 1980" (PDF). Government of the Soviet Union and Government of the Seychelles. 15 February 1980. Retrieved 2009-05-26.
- ^ "Catholic church says SPPF not telling the truth about past relationship". Seychelles Weekly. 7 July 2006. Retrieved 2009-05-26.
- ^ "Soviet base reported in Seychelles". London: The Spokesman Review. 9 November 1987. Retrieved 2009-05-26.
- ^ a b Российско-сейшельские отношения (in Russian). Ministry of Foreign Affairs of Russia. 16 February 2009. Retrieved 2009-05-26.
- ^ "Air Seychelles officially launches flight linking Seychelles-Moscow". Seychelles.travel. 24 March 2009. Retrieved 2009-05-26.
External links[edit]
|Wikimedia Commons has media related to Russia–Seychelles relations.


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Batteries made with chemicals that we mine from rhubarb plants, rather than expensive metals taken out of the earth. Magnets that clip onto electrical lines and reroute electrons. Solar panels that work efficiently even when they get hot or are shaded. These are just a few of the advanced energy research projects in the works that could bring the promise of a future electricity grid that is decentralized, resilient, and less carbon-heavy closer to a reality.
You've probably heard of DARPA, the Defense Advanced Research Projects Agency, or as Fast Company's Jessica Leber elaborates, "the military's research agency that funds the development of mind-reading science, cyborgs, and deadly robots." But have you heard of ARPA-E? As you might can guess from the title of this article, the "E" stands for Energy. Indeed, ARPA-E is the U.S. Department of Energy's analog to DARPA, and it's only five years old.
DARPA got its start in response to the Soviet Union's launching of the world's first artificial satellite, Sputnik 1. According to "DARPA: Bridging The Gap Powered By Ideas" (2005), "DARPA's original mission, established in 1958, was to prevent technological surprise like the launch of Sputnik, which signaled that the Soviets had beaten the U.S. into space. The mission statement has evolved over time. Today, DARPA's mission is still to prevent technological surprise to the US, but also to create technological surprise for our enemies."
There are many calls these days for another "moon shot" to restore America as the lead in the clean energy race and to combat the twin evils of our fossil foolishness, global warming and ocean acidification. That's where ARPA-E comes in!
According to ARPA-E's web site, "Who We Are,"
The Advanced Research Projects Agency-Energy (ARPA-E) advances high-potential, high-impact energy technologies that are too early for private-sector investment. ARPA-E awardees are unique because they are developing entirely new ways to generate, store, and use energy.
And under "What We Do,"
ARPA-E projects have the potential to radically improve U.S. economic prosperity, national security, and environmental well being. We focus on transformational energy projects that can be meaningfully advanced with a small investment over a defined period of time. Our streamlined awards process enables us to act quickly and catalyze cutting-edge areas of energy research.
Here's ARPA-E Acting Director Dr. Cheryl Martin discussing the future of innovation for the electric grid on NPR's Science Friday:
Image source: sciencefriday.com
"We're trying to take really crazy ideas… and we get it from the impossible to the plausible. That's an important place. Once people think it's plausible, then they can decide they want to step up and put some money to do a demonstration." ~ARPA-E Acting Director Dr. Cheryl Martin
Kyle G. Crider (MPA, LEED AP ND) is a professional science and sustainability "story teller." In his spare time he is pursuing his Ph.D. in Interdisciplinary (Environmental Health) Engineering and traveling the highways and by-ways of home state with his wife Beverly in search of fact, fiction, and folklore for Strange Alabama.


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I found some of Dagmar's classes on you tube about 2 years ago and they became part of my regular practice. When a trip to Costa Rica came up it was no question where I would spend my time. Montezuma Yoga has exceeded my expectations by far. The setting is beautiful, the open air studio is surrounded by mango trees, overlooking the ocean. All the teachers are world class, very experienced, helpful and friendly. I'm looking forward to class every morning, such a nurturing practice and beautiful energy. And it's not just the teachers, meeting like minded people from all around the globe has been such a nice experience too. Hours go by with inspiring conversations over breakfast each moning. I highly recommend Dagmar's studio and would come back here anytime.
Own or manage this property? Claim your listing for free to respond to reviews, update your profile and much more.


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login with
Prayers and hugs please
I got a call early this morning that my step dad had a heart attack. We saw him last night and he seemed fine. He had to have triple bypass surgery today so I was running back and forth to the hospital all day. He came out of surgery fine but there was a lot of damage to his heart tissue. They have to keep him intibated and sedated all night. Hopefully he will be able to wake up tomorrow
He is an amazing man who has been a godsend to my family. He helped my mother work through her issues, which made it possible for me to have a relationship with her. He has to be ok. He just has to.
my prayers your and your step dad's way!!! Wish him a speedy recovery!
praying for you and your family.
thinking of you. Hope he recovers okay.
So sorry Taylor. Hugs for you and your family.
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About Taylor
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Birth: July 26
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I'm the proud mama of my daughter Avery, born on June 6, 2013. I'm 26 years old, I work from home as a graphic designer, I will be testing for my 5th degree black belt in Tae Kwon Do in 2015. My husband, Derek, and I have been happily married for 3 years.


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If you are conducting some Stone family history research then you can view the Stone coat of arms and the name meaning and origin of the name as well as the meaning of symbols below. This is a great place to start when tracing your genealogy and Stone surname history and will help you answer the question 'what is the meaning of my name?'
The name Stone in Ireland is usually
of immigrant origin having been
introduced into the Province of
Ulster by settlers from England
and Scotland, especially during
the seventeenth century.
Stone has also been used as an
anglicized form of the Gaelic
O'Clochartaigh and O'Maolchluiche
sept names of Connemara and
Munster. These names were more often
changed to Clogherty and Muckley.
A Sept is an ancient form of a Clan and is a group of people whose ancestors all bore the same surname and inhabited the same locality.
|Represents Loyalty and Truth.
|'The Martyr's colour', signifies Military Fortitude and Magnanimity.
|Represents Generosity.
|Signifies Hope and Joy
|Symbol of Power and Strength
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From Wikipedia, the free encyclopedia
|This article does not cite any sources. (December 2009) (Learn how and when to remove this template message)
Kibon is a Brazilian-Argentine ice cream producer, now owned by Unilever. The logo that it uses is the same Heartbrand logo that Wall's ice cream, Good Humor, Streets and Selecta use in the United Kingdom, United States, Australia and Philippines respectively, also owned by Unilever.
- Kibon was founded in 1941 by Americans U.S. Harkson and John Kent Lutey. Lutey was the on-site Director and President. The two originally worked together in Shanghai, China. The original corporate name was U.S. Harkson do Brasil, which was later changed to Cia Harkson Industria e Comercio Kibon. Lutey was Director-President until 1962.
- In 1942, Kibon introduced its first ice cream products to Brazil: Eskibon (chocolate-covered ice cream) and Chicabon (chocolate and malt popsicle) with 50 refrigerated pushcarts on the streets of Rio de Janeiro. They were an enormous success and the company grew quickly, introducing classic and tropical flavors. The company also produced dried eggs for food industries. Kibon later introduced a variety of candies, chocolates and gum. By 1960, Kibon was available through more than 3,000 pushcarts as well as retail stores throughout the industrial south of Brazil.
- In 1957, Kibon entered into a joint venture with the General Foods Corporation to manufacture and market several of its brands, including Kool-Aid and breakfast cereals. The joint venture was named Industrias Alimenticias Gerais, S.A., a Brazilian corporation.
- In 1960, the General Foods Corporation bought Kibon and Kibon's share of the joint venture.
- In 1985, the firm was bought by Philip Morris.
- By the 1990s, Kibon was made by the Swiss company Mövenpick.
- In 1995, Kibon bought Brazilian chocolate company Lacta.
- In 1997, the Anglo-Dutch company Unilever bought Kibon.
- In fall 2013, Kibon will be sold in the Falkland Islands.
External links[edit]
- (in Portuguese) Kibon website
|This Brazilian corporation or company article is a stub. You can help Wikipedia by expanding it.
|This Brazilian cuisine-related article is a stub. You can help Wikipedia by expanding it.


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Man becomes 16th farm fatality of the year
A man has died following a farm incident in Carlow.
The ma, who is in his 60s, died following an accident on a farm yesterday, Gardai said. A Garda spokesperson confirmed that the accident happened on a farm near Hacketstown.
Gardaí at Tinnehely attended the scene where the accident happened, just after 4pm on Saturday.
A man in his 60s was fatally injured while carrying out work on farmland. He was pronounced dead at scene and conveyed to Naas Hospital Mortuary.
The Health and Safety Authority has been notified and a post mortem will take place.
This is the 16th farm fatality to happen on Irish farms this year. Two farmers in Co Kerry died in separate farm accidents in recent weeks.
Farmers are eight times more likely to die in a farm accident than any other occupation and figures from the Health and Safety Authority show that working with machinery and animals are the two main causes of deaths on Irish farms.
On average 19 people are killed on Irish farms every year, making farms the most dangerous work places in Ireland.
There have been almost 200 farm fatalities in the last decade.
Margaret Donnelly


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2D-Sliding Mesh Problem
my name is Tim. I´m a german mechanical engineering student. I have been reading this website for quite some time. But now I have encountered difficulties and need help :).
I have tried to get the sliding mesh problem working several times. But I always get the following result:
Is there anyone who could give me some advice? Is this a known issue? The sliding mesh-interface seems to have a problem.
Would it be helpful to post the Ansys-Workbench project on the internet?
Thanks in advance
hi tim,
I can't really see what is going on in your picture.
Also I use Fluent directly and Gambit for the meshing but it should be similar in workbench.
So here are a few remarks:
If you want to move e.g. a box through a rectangle domain you can use layering.
the sides of the rectangle need to be specified as stationary, the box and the fluid must be specified as moving. for the movement you need to provide a UDF or *.profile file. also on the stationary sides of the rectangle you need to specify how cells are created and destroyed.
Without more information it is not really possible to help, so please provide a detailed problem description
Hi Martin,
I just found out, that I had to switch the Contour-Plot to stationary frame.
Now everything works fine.
Thank you very much for your reply!
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