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The goal of adoptive T cell therapy in cancer is to provide effective antitumor immunity by transfer of selected populations of tumor Ag-specific T cells. Transfer of T cells with high TCR avidity is critical for in vivo efficacy. In this study, we demonstrate that fluorescent peptide/MHC class I multimeric complexes incorporating mutations in the α3 domain (D227K/T228A) that abrogate binding to the CD8 coreceptor can be used to selectively isolate tumor Ag-specific T cells of high functional avidity from both in vitro expanded and ex vivo T cell populations. Sorting, cloning, and expansion of α3 domain mutant multimer-positive CD8 T cells enabled rapid selection of high avidity tumor-reactive T cell clones. Our results are relevant for ex vivo identification and isolation of T cells with potent antitumor activity for adoptive T cell therapy.
The identification of T cell-defined tumor Ags has set the stage for new cancer therapies based on Ag-specific immune responses. In the approach usually referred to as specific adoptive T cell therapy, patient-derived T cells of defined antigenic specificity are isolated, expanded in vitro, and reinfused to enlarge the pool of effector T cells in vivo (for review, see Ref. 1). Recent results of adoptive T cell therapy trials support the validity of this approach (2, 3). It is yet unclear what are the functional properties of the transferred T cells that contribute most to inhibition of tumor growth. Certainly, an important factor that influences their in vivo efficacy is the avidity of TCR interaction with its cognate peptide/MHC (pMHC)5 ligand (4, 5, 6, 7).
We asked whether fluorescent pMHC class I multimers unable to bind the CD8 coreceptor could identify high avidity T cells among heterogeneous populations. Binding of pMHC to CD8 involves mainly a conserved, negatively charged loop in the α3 domain of the MHC H chain (residues 223–229) (8, 9, 10). Accordingly, double amino acid substitutions such as D/T to K/A in positions 227/228 (thereafter 227,8KA) abrogate binding to CD8 (11, 12). Besides its role in signal transduction (12), CD8 augments the stability of TCR/pMHC interaction by binding cooperatively to the same pMHC molecule, which results in enhanced TCR avidity (11, 13, 14, 15). Previous reports suggest that CD8 dependence for multimer binding may vary according to TCR avidity (13, 16). In particular, when TCR avidity is relatively low, CD8 contributes significantly to pMHC binding. We reasoned that pMHC class I multimers containing 227,8KA-mutant H chain (thereafter peptide/227,8KA-A2 multimers) may fail to stain T cells with low avidity, but still bind to high avidity, tumor-reactive T cells. To address this hypothesis, we chose the repertoire of CD8 T cells reactive to the self-differentiation Ag Melan-A/MART-1 (thereafter Melan-A) (17, 18). We found that 227,8KA-A2 multimers selectively labeled high avidity tumor-reactive T cells and could be used to isolate such T cells even when present at a frequency as low as 1 in 10,000 CD8 T cells.
Materials and Methods
Cells and tissues
Cells were collected from HLA-A*0201 individuals, as identified by typing with PCR sequence-specific oligonucleotide probe. Melan-A/wt-A2 T cell clones were derived from PBMCs of a healthy individual (HD 009) and a melanoma patient (LAU 203), from a thymus tissue of a child that had undergone corrective cardiac surgery (T 12), from lymphocyte suspensions of vaccine-site sentinel lymph nodes (LNs) (LAU 371 and 445), and a tumor-infiltrated LN (LAU 392) from melanoma patients (19). A clone specific for the influenza matrix flu-MA58–66 peptide was derived from PBMCs of a healthy individual (HD 008). Clones were expanded by stimulation with 1 μg/ml PHA, 105 irradiated allogeneic feeders, and 100 U/ml IL-2. PBMCs from healthy individuals (HD 006, HD 009) and melanoma patients (LAU 155, 203, 233, 269, 337, 465, 567) and tumor-infiltrated LNs from melanoma patient LAU 352 were collected for ex vivo multimer staining.
pMHC multimers
PE-labeled HLA-A2 multimers were synthesized around the Melan-A26–35 A27L (ELAGIGILTV) peptide (20). The HLA-A2 H chain mutants D227K/T228A were produced by PCR mutagenesis using the QuickChange Mutagenesis Kit (Stratagene, La Jolla, CA). Mutant proteins were expressed in Escherichia coli, refolded in vitro, and purified as for the wild-type HLA-A2 molecule. ELISA using the w6/32 mAb were performed to ensure correct folding of MHC H chain/peptide/β2-microglobulin complexes.
Flow cytometry immunofluorescence analysis and sorting
CD8 T lymphocytes were negatively selected from PBMCs and tumor-infiltrated LNs using a CD8 T cell isolation kit (Miltenyi Biotec, Bergisch Gladbach, Germany). Cells were stained either with PE-labeled Melan-A/wt-A2 or Melan-A/227,8KA-A2 multimers for 1 h at room temperature, incubated with appropriate mAbs for 20 min at 4°C, washed, and immediately analyzed on a FACSVantage SE, using CellQuest software (BD Biosciences, San Jose, CA). Multimer staining at 4°C showed similar mean fluorescence intensities (MFIs), albeit values were in this case in general lower. Anti-CD3 FITC mAb and goat anti-rat IgG APC were purchased from BD Biosciences; anti-CD45RA-ECD mAb was from Immunotech (Nyon, Switzerland). Anti-CCR7 rat IgG mAb 3D12 was provided by M. Lipp (Max Delbrück Institute, Berlin, Germany). Melan-A/wt-A2 or Melan-A/227,8KA-A2 T cells were cloned by staining with Melan-A/wt-A2 and Melan-A/227,8KA-A2 multimers, respectively, and single cell sorting was performed using a FACSVantage SE.
Cytolytic activity
T cell clones were tested for their lytic activity in 4-h 51Cr release assays (20). The HLA-A,B-negative B cell line C1R expressing wild-type or mutant HLA-A2 molecules (provided by A. Sewell, Nuffield Department of Clinical Medicine, Oxford, U.K.), the melanoma cell lines Me 290 (Melan-A+/A2+), and Na 8 (Melan-A−/A2+) were used as target cells.
Selective staining of a high avidity, tumor-reactive Melan-A-specific CD8 T cell clone with Melan-A/227,8KA-A2 multimers
Initially, we selected two Melan-A-specific T cell clones (clones 1 and 5) that were derived from the same individual (T 12). Although both clones were efficiently stained with Melan-A/wt-A2 multimers (Fig. 1⇓a), clone 1 specifically lysed Melan-A-expressing tumor cells, whereas clone 5 did not (Fig. 1⇓b). With the latter, lysis was only observed when saturating doses of the Melan-A peptide were exogenously added. Functional avidity of Ag recognition defined as concentration of peptide required for 50% maximal cytolytic activity was assessed in a 51Cr release assay using C1R cells expressing wild-type HLA-A2 as target cells in the presence of serial dilutions of the Melan-A peptide (Fig. 1⇓c). Clone 1 efficiently recognized the Melan-A peptide (IC50, 10−11 M), whereas the functional avidity of clone 5 was significantly lower (IC50, 10−7 M). We tested whether the two clones could be distinguished based on the staining efficiency obtained with serial dilutions of multimers. However, the MFI of staining with Melan-A/wt-A2 multimers was similar and equally dose dependent for both clones (Fig. 1⇓e). To abrogate the participation of CD8 in multimer binding, we then produced multimers containing α3 domain mutants of the HLA-A2 H chain. Clone 1 was positively stained with Melan-A/227,8KA-A2 multimers, whereas the MFI of clone 5 was similar to that observed with an irrelevant influenza matrix flu-MA58–66 T cell clone (Fig. 1⇓, d and e).
High avidity, tumor-reactive Melan-A-specific CD8 T cell clones are selectively stained using Melan-A/227,8KA-A2 multimers and exhibit variable CD8 dependence
To further document these findings, we derived a series of Melan-A-specific T cell clones from healthy individuals and melanoma patients by Melan-A/wt-A2 multimer-assisted single cell sorting. We obtained 35 clones that were specifically stained with 300 ng/ml of Melan-A/wt-A2 multimers; this concentration did not result in staining above background of T cell clones with irrelevant specificity (Fig. 1⇑, d and e). All clones expressed comparable levels of TCR and CD8, as indicated by staining with specific mAbs (data not shown).
The Melan-A-specific T cell clones were further analyzed for tumor reactivity, as assessed by lysis of Me 290 (Melan-A+/A2+) or Na 8 (Melan-A−/A2+) melanoma tumor cells in 4-h 51Cr release assays (Fig. 2⇓a). A total of 16 of 35 clones efficiently killed Melan-A-expressing tumor cells (• and gray circles), whereas the 19 remaining clones did not (○). Functional avidity of Ag recognition was assessed on wild-type HLA-A2-transfected C1R cells as targets in the presence of serial dilutions of Melan-A peptide (Fig. 2⇓b). The functional avidity was extremely variable among the clones, with IC50 values varying between 10−13 and >10−6 M. As expected, tumor-reactive clones exhibited lower IC50 (median, 10−11 M) than nontumor-reactive clones (median, 10−8 M). All clones showed comparable intensity of staining with Melan-A/wt-A2 multimers. By contrast, when clones were incubated with Melan-A/227,8KA-A2 multimers (Fig. 2⇓c), none of the 19 nontumor-reactive clones were stained above background (group I, ○). These clones exhibited low functional avidity, but nevertheless could be stained with Melan-A/wt-A2 multimers. Among the tumor-reactive T cell clones, 9 of 16 were positively stained with Melan-A/227,8KA-A2 multimers (group II, •), whereas the 7 remaining clones were not (group III, gray circles). Accordingly, lack of multimer labeling in the latter group suggests that a significant fraction of T cells with high functional avidity was dependent on CD8/pMHC interaction. We used a blocking anti-CD8 mAb to confirm CD8-dependent binding of Melan-A/wt-A2 multimers to clones from group III. In contrast to clones from group II, multimer staining of clones from group III decreased when increasing doses of anti-CD8 mAb were added (data not shown).
To verify the requirement of CD8 in the recognition of APCs by the different Melan-A-specific T cell clones, C1R cells transfected with HLA-A2 D227K/T228A were used as target cells (Fig. 2⇑d). The majority of clones from group I failed to recognize the Melan-A peptide (median IC50, >10−6 M), underlining the essential contribution of CD8 to TCR/pMHC binding. In contrast, all clones from group II exhibited a low IC50 (median, 10−11 M), i.e., efficiently recognized peptide-loaded C1R targets with abrogated CD8/pMHC interaction. This confirms that the high avidity TCR/pMHC interaction of these clones was CD8 independent and enabled staining with Melan-A/227,8KA-A2 multimers. Finally, clones from group III exhibited a high IC50 (median, 10−6 M), stressing again the requirement of CD8 for TCR/pMHC binding for these clones. Nevertheless, a few clones from groups I and III displayed relatively high lytic activity against peptide-loaded D227K/T228A HLA-A2 mutant C1R cells, although they were not stained with Melan-A/227,8KA-A2 multimers. This suggests the possibility that accessory molecules other than CD8 expressed by these clones could compensate for CD8 in the functional Ag recognition assay. In addition, it is formally possible that D227K/T228A HLA-A2 mutant C1R cells, although lacking the D227 CD8 binding site, might still have served as CD8 avidity-enhanced accessory ligands (21).
Ex vivo identification, isolation, and characterization of Melan-A/227,8KA-A2 multimer+ cells in healthy individuals and melanoma patients
To evaluate the potential of Melan-A/227,8KA-A2 multimers for adoptive T cell therapy, we investigated whether tumor-reactive T cells could be directly identified and isolated from PBLs. To directly compare binding of Melan-A/227,8KA-A2 multimers with naive and Ag-experienced Melan-A-specific T cells, we used PBLs from healthy individuals (n = 2) as well as from melanoma patients with detectable Ag-experienced Melan-A-specific T cells (n = 8). We used both wild-type and mutated multimers in association with anti-CCR7 and anti-CD45RA mAbs (Fig. 3⇓, a–c). The frequency of Melan-A/227,8KA-A2 multimer+ cells in CD8 T cells was reduced as compared with that of Melan-A/wt-A2 multimer+ cells (mean ± SD, 0.3 ± 0.5% and 1.0 ± 1.3%, respectively), i.e., ∼30% of Melan-A-specific T cells stained positively with Melan-A/227,8KA-A2 multimers. These cells were detectable ex vivo in all 10 individuals analyzed. Detailed analysis revealed that ∼20% of naive Melan-A-specific T cells stained positively with Melan-A/227,8KA-A2 multimers, both in healthy individuals and melanoma patients (Fig. 3⇓c). Conversely, ∼40% of Ag-experienced Melan-A-specific T cells stained positively with Melan-A/227,8KA-A2 multimers (Fig. 3⇓c). It is likely that these cells exhibit a high avidity, CD8-independent TCR/pMHC interaction, similar to the clones from group II identified in Fig. 2⇑. The fraction of Ag-experienced Melan-A-specific T cells that were not stained with mutated multimers may resemble clones from group III in Fig. 2⇑, i.e., exhibit a high avidity, CD8-dependent TCR/pMHC interaction.
To substantiate the usefulness of MHC class I multimers with abrogated CD8 binding for the isolation of Ag-specific T cells with high functional avidity, we derived a series of clones from healthy individual HD 009 by single cell cloning ex vivo (Fig. 3⇑a) of either Melan-A/wt-A2 multimer+ (n = 10 clones) or Melan-A/227,8KA-A2 multimer+ CD8 T cells (n = 11 clones). All clones were specifically stained with wild-type multimers, whereas Melan-A/227,8KA-A2 multimers selectively stained Melan-A/227,8KA-A2 multimer+-sorted clones (data not shown). Tumor recognition was assessed on tumor target cells expressing or not Melan-A in a 4-h 51Cr release assay (Fig. 3⇑d). All 10 Melan-A/wt A2 multimer+ T cell clones failed to recognize Melan-A-expressing tumor cells. These clones exhibited a structural avidity sufficient to bind Melan-A/wt-A2 multimers, but a functional avidity too low for tumor reactivity. In contrast, 8 of 11 Melan-A/227,8KA-A2 multimer+ T cell clones exhibited potent and specific lysis of Melan-A-expressing tumor cells. Tumor reactivity of Melan-A/227,8KA-A2 multimer+ T cells was confirmed with T cell lines generated from three healthy individuals and two melanoma patients (data not shown).
The peripheral T cell repertoire specific for a particular pMHC ligand comprises a polyclonal population of T cells harboring TCRs of various avidities (22). As antitumor activity of T cells in cancer therapy is correlated to TCR avidity, the ability to rapidly identify specific T cells with high avidity is a critical requirement. In vitro stimulation with peptide-loaded APCs leads to expansion of Ag-specific T cells with a broad spectrum of avidities, of which very few are tumor reactive (6). Several approaches have been described to preferentially expand high avidity T cells. One approach that allows at least an enrichment of such T cells is based on the proposed inverse correlation between TCR avidity and concentration of antigenic peptide used during in vitro expansion (4, 5). Another approach relies on the isolation of strongly labeled T cells by flow cytometry cell sorting after staining with pMHC class I multimers, which is based on the assumption of a direct relationship between intensity of multimer labeling and TCR avidity (23). However, it has been recently noticed that other parameters such as experimental conditions, interaction with coreceptors, or integrity of lipid rafts may affect the staining efficiency of multimers (24). Recently, the measurement of TCR avidity that is based on the rate of decay of multimer binding has been suggested as a more reliable procedure (24). Because conventional multimers are heterogeneous mixtures of pMHC conjugates (16), the validity of this approach remains to be confirmed.
As a model, we investigated the T cell repertoire specific for the Melan-A26–35 tumor-antigenic peptide (17). We previously found that a large pool of Melan-A-specific CD8 T cells (∼10−3 CD8 T cells) is generated and maintained as a naive CCR7+CD45RAhigh population in healthy individuals (25). It is composed of T cells that exhibit a broad range of functional avidities of Ag recognition and significant differences in tumor reactivity (26). A fraction of these naive T cells may become activated and shift toward an Ag-experienced CCR7−CD45RAlow phenotype in ∼30% of melanoma patients (20, 27). During such an immune response, only T cells with high TCR avidity become part of the Ag-experienced T cell repertoire (A. Zippelius et al., manuscript in preparation). As wild-type multimers do not provide information on the functional avidity of T cell clones (this study and 24, 26), such reagents are ineffective to reliably isolate high avidity Melan-A-specific tumor-reactive CD8 T cells. We explored a novel approach by means of soluble pMHC multimers incorporating mutations in the α3 domain (D227K/T228A) that abrogate binding with the CD8 coreceptor. These reagents have been used recently to label low CD8-expressing Ag-specific T cells (28). As compared with wild-type multimers, α3 domain mutant multimers did not stain CD8 T cells with low functional avidity for Melan-A peptide. These findings corroborate previous observations that CD8 exerted a critical role in pMHC binding to low avidity TCR (13, 29). By contrast, a significant fraction of T cells with high functional avidity did not require CD8 for binding of pMHC multimers. Based on these novel insights into CD8 coreceptor participation of high avidity TCR/pMHC interactions, α3 domain mutant multimers allowed to selectively identify tumor-reactive T cells from freshly obtained lymphocytes of melanoma patients as well as healthy blood donors. We document a high sensitivity and specificity of such multimers, as the frequency (0.01%) of Melan-A/227,8KA-A2 multimer+ cells in CD8 T cells represented the lower limit of detection under the conditions used. The finding that a small fraction of naive tumor-reactive Melan-A-specific T cells could be stained with Melan-A/227,8KA-A2 multimers is compatible with the recent observation that high avidity T cells only compose a minority of the naive Melan-A-specific T cell pool. As the majority of melanoma patients do not exhibit Ag-experienced Melan-A-specific T cells in the peripheral blood (20, 27), α3 domain mutant multimers may open the avenue for generating tumor-reactive T cells from the vast majority of melanoma patients.
To our knowledge, we show for the first time that mutated pMHC class I multimers allow rapid identification and isolation of T cell clones with sufficient functional avidity for in vivo efficacy. The finding that tumor-reactive Melan-A-specific T cells can be stained with the mutated multimers may permit expeditious generation of T cell populations for adoptive transfer therapy in HLA-A2 melanoma patients.
The HLA-A2-transfected C1R cell lines and the anti-CCR7 mAb were generous gifts from Dr. A. K. Sewell (Nuffield Department of Clinical Medicine) and Dr. M. Lipp (Max Delbrück Institute), respectively. We thank Dr. H. R. MacDonald for critical reading of the manuscript, and Céline Barrofio, Andrée Porret, and Martine van Overloop for excellent technical assistance.
↵1 A.Z. was partly supported by a grant from the Deutsche Forschungsgemeinschaft.
↵2 M.J.P. and V.R.-G. equally contributed to this work.
↵3 Current address: Center for Molecular Imaging Research, Harvard Medical School, Charlestown, MA 02129.
↵4 Address correspondence and reprint requests to Dr. Alfred Zippelius at the current address: Department of Oncology, University Hospital Zurich, Rämistrasse 100, 8091 Zurich, Switzerland. E-mail address: alfred.zippelius{at}usz.ch; or Dr. Pedro Romero, Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Hôpital Orthopédique, Niveau 5, Aile est, Avenue Pierre Decker 4, 1005 Lausanne, Switzerland. E-mail address: pedro.romero{at}isrec.unil.ch
↵5 Abbreviations used in this paper: pMHC, peptide/MHC; LN, lymph node; MFI, mean fluorescence intensity.
- Received April 7, 2003.
- Accepted June 13, 2003.
- Copyright © 2003 by The American Association of Immunologists


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Ambrose Rachier set 'unrealistic' target for Gor Mahia ahead of the new season
Kenyan champions Gor Mahia have set close to impossible target ahead of the upcoming season.
K'Ogalo are once again expected to have a busy season with at least six titles up for grab. Gor Mahia will be looking for their maiden Caf Champions League title when they kick off the continental campaign on Wednesday against Nyasa Big Bullets of Malawi.
Beside that, the record Kenyan champions will be up to defend the domestic league, the SportPesa Super Cup as well as reclaiming the Shield Cup title.
Gor Mahia, who might lift their maiden title when they take on Kariobangi Sharks in the Super Cup final on December 6, will also be taking part in the regional Cecafa Club competition-if the games will be there in the first place.
But despite the tedious task ahead, club chairman Ambrose Rachier says that K'Ogalo will be going for all silverware up for grab.
"Last season we set a higher standard by claiming the league title, winning the Super Cup. The dream has not changed. In fact, this season, we have even a bigger target of winning every competition we'll be taking part in," Rachier noted.
K'Ogalo will host Nyasa Big Bullet on Wednesday before traveling to Malawi for the return leg on December 5. After the away fixture, they will take on Kariobangi Sharks in the Super Cup final before kicking off the KPL season away in Mombasa, against Bandari on December 8.

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Jones Bridge Back to Front
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Please join me for a hike through the beautiful trails of the Jones Bridge unit of the Chattahoochee River National Recreation Area.
We will be hiking at a steady but moderate pace only stopping to hydrate. There are a couple of hills but otherwise the trail is flat along the Chattahoochee River. The trail has rocks and roots so hiking poles might be helpful.
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NOTE.....We will start the hike at the Wine Store on the corner of Holcomb Bridge Road and Barnwell. We will walk down Barnwell into Rosemont Park and then into Jones Bridge.
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By Nigel Clark
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Start Date: Friday 1st February 2019
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Post: Midday Supervisor
Start date: March 2019
Salary: £8.55 per hour
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Burton End Primary Academy is part of the Unity Schools Partnership. To support the running of the school, we are seeking to recruit a dedicated individual into our team of midday supervisors to support our pupils over the lunchtime period. You will be understanding and supportive of our pupils and supervise them while eating lunch and socialising during the lunch break.
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We are committed to safeguarding and promoting the welfare of children, young people and vulnerable adults and expect all staff and volunteers to share this commitment. DBS checks required for relevant posts.
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DESCRIPTIONAustralia's biggest telecommunications company is investing in customer driven models include lean and agile practices to transform their world of work.

 1. TELSTRATEMPLATE4X3BLUEBETA|TELPPTV4TELSTRATEMPLATE4X3BLUEBETA|TELPPTV4 ENTERPRISE AGILE NBN - NETWORK INVENTORY AGILE DELIVERY PROGRAM STEPHANIE BYSOUTH LEAN AGILE COACH @innovatebysouth www.innovationbysouth.com.au

 2. VALUES . The Eight Fundamental Truths of being an enterprise lean agile coach to remember. 1. People and companies can be more effective than they are today. 2. We know you can make a difference. 3. Opportunity comes to those who create it. 4. People, Community and Culture are the heart of all success. 5. Systems, processes and delivery are all created by people. 6. Innovation is available to every person, process and product. 7. Coaching is the competitive leverage your neighbor wishes they had. 8. We love what we do and do it simply because we can make a difference. 3. ESTABLISH A FOUNDATION Appreciate Engage Participate 4. APPRECIATE THE ENTERPRISE 4 5. APPRECIATE THE ENTERPRISE; UNDERSTAND THE ENVIRONMENT ON THE GROUND 1. People - capacity to change, rate of learning 2. Culture is THE undocumented influencing & controlling system 3. System infrastructure, architecture, constraints and alternative success flows 4. Tools in play for the enterprise vs. for the team to create clarity and simplicity 5. Practices designed by others, failed by users vs. local, feasible, applicable, doable 6. COMPLEX INTEGRATED APPLICATION ENVIRONMENT us. 7. ENTERPRISE PROGRAM - OUR PROGRAM COMPLEX INTEGRATIONS 8. OUR PEOPLE us. Vendor Management * varied Project Manager & Iteration Manager Product Owner Stakeholders * 6 4 * BA 2 * Design 12 * Dev 5 * Release & Test Vendor Management * varied Project Manager & Iteration Manager Product Owner Stakeholders * 6 4 * BA 2 * Design 10 * Dev 5 * Release & Test Program Management * 3 Solution, Application, Release & Test Management * 3 Local Program Management * 2 2 * Architect 9. C R C R C R PROD PRODCAT PRODCAT PRODCAT LAYERED INFRASTRUCTURE, BRANCHING, MANUAL 9 TEAM A stream 1205 NETWORK INTEGRATIONS stream CAT PRODintegration PRODintegration stream stream stream C R C R C R PROGRAM LOCAL- DEV DEV System Test integrati on BAU Converging environment demands across multiple streams of delivery BAU Corrupting true Capacity + administration overload to ensure baseline integrity + cost of context switching HIGH COST TO REGRESSION + SYSTEM TESTING = TIME + DEFECT RATE 10. APPRECIATE - FLOW, SYSTEM CONSTRAINTS People Culture Process ToolsPractices HR Finance Delivery 11. ENGAGE LEADERS EARLY 1 12. Program & Release Planning Team Customer capture Validate Prioritise MeasureValidate3 tiered MVP FUNNEL FOCUS COLLABORATE AUTOMATE CONTINUOUS INTEGRATION GIVE UNDERSTANDING & SHARE VALUE DRIVEN ROADMAP 13. INVEST FOR STRATEGIC LEVERAGE Teach, and provide insight on how the enterprise will rapidly (1-2yr) advance 14. PARTICIPATE BUSINESS / CUSTOMER COLLABORATION VALUABLE & EASY 15. CO-LOCATE EVERYONE (AT LEAST) IN DISCOVERY 16. THE PEOPLE. Appreciate Empathy Participate 17. STOP WHAT NOTS WORKING! Really? 30 people expected to do a 15 minute stand-up? - always ask WHY is this valuable? 18. APPRECIATE - COMPLEX TO ACCOUNTABLE VISUALIZE ALL DATA FROM PROGRAM, PORTFOLIO TO TEAMS AND ENVIRONMENTS 19. ELIMINATE PROBLEMSBUILD ACHIEVEMENT & CREATE CAPACITY FOR AGILE CHANGE TO COME IN Kaizen 1. Problem 2. Action 3. Outcome 20. CREATE CORE TEAM TO INTERNALISE ASSETS & IP SUSTAIN THE CORE AS LONG TERM AND POPULATE NEW TEAMS AS VARIABILITY DEMANDS 21. BREAK DOWN ORGANISATIONAL BARRIERS WITH HUMAN CONNECTIONS 22. PARTICIPATE - PROTECT & GUIDE CREATE CLARITY, ACCOUNTABILITY & FOLLOW THROUGH 23. THE PRACTICES. Accountability Evolve Participate 24. PARTICIPATE CONTINUOUS PLANNING 25. OUTCOME CONSTRAINT MAPPING LEVERAGE VARIETY OF MODALITIES TO CREATE VISIBILITY OF THE SYSTEM 26. ACCOUNTABILITY - LEVERAGING KANBAN 27. EVOLVE THE TEAM FROM WORK TO INNOVATION 28. LOOKING BACK 12 MONTHS 2 F T P D V D Program b branching Program A branching integration Production VMs 29. AUTOMATED TESTING Manual Testing Sanity testing : 2-3 hrs 1 person Unit testing : Not considered as part of this BC Acceptance testing Approx 50 Test Case 3weeks (4-5 resources) Regression testing Limited : Time/resource constraints Approx 10 TC 1-2 resource 1 wk Automated Testing Sanity testing 4 TC 15min (4* more than normal) Unit testing : Not considered as part of this BC Acceptance testing 2 VM approx 250 TC 9 hrs execution time 2 hr / Dev review ( 3Dev) Regression testing 50 TC 2.5hrs execution time VS 30. THE BOTTOM LINE 71% 29% 2012 Weekends worked ON OFF 0% 100% 2013 ON OFF $0 $100,000 $200,000 $300,000 $400,000 $500,000 $600,000 2012 2013 Automation & Continuous Integration Savings / deployment Merge, Test Integration Deployment Production Deployment Testing Application Solution Integrity sustained Standard way of work vs. an expensive option Team members now mentor other new agile teams Foundation / evidence for business case to continue adoption of agile tech practices 31. THANKS. Stephanie BySouth www.innovationbysouth.com.au @innovatebysouth #agile #coaching

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Exploring Enterprise Agile Transformation Strategies Mike Cottmeyer, Enterprise Agile Coach LeadingAgile, LLC


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The Priesthood of Melchizedek is the overseeing function of all Light Workers to assist each Light Worker to stand in the truth of their Divinity with Lord Melchizedek as a guide and mentor for the New Earth.
This is one who works with the Higher Forces of Light to fully acknowledge the remembrance of the True Self to become manifest within the physical creation. To be able to hold that Light first within the physical creation, and change all aspects of the personality of the Self is to become integrated with the I AM Presence, the ultimate form of Light in the Creation of Oneness under the complete direction and guidance of the Unified Whole Command within the 144th dimension of reality.
This Light will infuse within the physical vehicle to assist in clearing the debris of the past. The Etheric Body can be healed once and for all to access the Higher Mind into the Lower Mind; to remove the Lower Ego and access the Higher Ego for the good of humanity and the Creation of Light within the planetary forces guided by the Spiritual Hierarchy of the Brotherhood and Sisterhood of White Light under the auspices of Lord Sanat Kumara, Father of the Brotherhood of Light and Lord Buddha, our Planetary Logos.
Lord Melchizedek holds the Priesthood of Light for all Initiates on the Pathway of Mastery.
©2012-2014 Walking Terra Christa Academy of New Earth Mastery / by Rev. Christine Meleriessee and Rev. Michael Aranathanara. Permission to repost this information electronically on your non-commercial website or blog is granted as long as you include this copyright statement and the following referral links. Use for commercial websites, blogs and printed or written reproduction requires written permission.
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At Mischief Managed, we are open to all styles of roleplay, as well as all experience levels.
Whether we're your first roleplay experience or you para-RP, your style and contribution are welcome here.
That being said, there are a few basic courtesies that we do ask you to observe. " Brenna Morrigan rubs her tummy as she hears the dinner bell chime in the Great Hall.
There are also some terms thrown around that, to a new roleplayer, might be unfamiliar. Brenna Morrigan looks around as she walks up to the Hogwarts Express and spots her friend. Once arriving, she walks to her house table and sits down, leaning forward to grab a roll off the table.
You do NOT need to roleplay after every professor post. It's also courteous to post no larger than a solid paragraph when roleplaying in class. This is when you are talking over and over again before giving anyone a chance to respond.
Since there may be 10 people all trying to get the most out of the class, shortening your standard 3-paragraph RPs to just 1 is a nice thing to do. You should post one, and only one, post before waiting for your next turn. A courtesy to RPers around you is to follow a simple order.
It's easier for the professor and your classmates to engage with and ensures you get your post out in time to be relevant. There should always be a short action in your roleplay when one-line roleplaying. [] Brenna Morrigan waves as she sees her group of friends. Of course this usually does not apply in classes, for example.
You do not have to shorten your posts; but, do not expect others to wait for you or the professor to acknowledge you if your posts are consistently behind those of everyone else. If wishing to join a group of RPers that are already immersed, it never hurts to IM them and politely ask the order of the RP and where you can come in.
Chat boxing is when you just talk in local without using any sort of emotion. If someone RPs in a style that you do not like, you can simply walk away and not chose to RP with them, or you can adjust your style however you feel fit to meet theirs.
This is entirely up to the player — no one is forced to Roleplay a certain style, and anyone found attempting to change the way someone roleplays will face OOC consequences.


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Saving money on healthy meal ingredients can be a challenge. Coupons are few and far between on things like meat, cheese and veggies. So how do you maximize your budget without resorting to beans and rice? Here are my top tips.
Be Loyal to Stores That Give Consistently Good Deals
Find stores that offer good prices on most everything (even the stuff that isn't on sale), and give them most of your business. Stores like Kroger that offer huge savings on a few items, and grossly overprice the rest will end up saving you less – even with all the extra effort of coupon clipping, and planning meals around the sales. I've tried it! For the long haul, it is best to reward the stores with the best pricing plans with the majority of your business.
Visit the Farmers Market
Buying in-season produce straight from the source is often the best way to score a deal. My biggest savings has come when I spend some time getting to know my favorite vendors, and ask about peak-season bulk discounts. If you're willing to do some canning or freezing, this relationship will pay off for years to come!
Reduce Food Waste
I've written a lot lately about food waste, mostly because it is the biggest money drainer on the American grocery budget. According to the EPA, we waste as much as 20% of what we bring home from the grocery store. So being mindful of what we have, and when it expires is not something I want to take lightly. Follow a meal plan that includes snacks and beverages to shrink your waste. Be sure to include leftovers into the plan when appropriate.
Make More Items From Scratch
Did you know a loaf of bread costs around $0.60 to make? Or that you can use dried beans and get 6 cups for $1? Did you know you can substitute expensive pre-made cereal for oats at around one-third the cost? Get into the habit of making a few frequently eaten things from scratch, and save as much as $10 per week or more!
Ditch the Drinks
I am a sucker for a sweet beverage. Especially with my meals. But I have seen such a huge savings by opting for water – not to mention the calories I save! When I do have a hankering for something sweet, I go for something homemade and affordable, like a big batch of sweet tea!
Let Your Family In On The Plan
I can't tell you how many times I have attempted to make a meal, and then discovered I am missing ingredients because some hungry teenager decided to eat them as a snack! Posting my meal plan in an obvious place, and letting my family know they are accountable to it has helped save money and my sanity!
How about you? What routines help you save money on food?


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Mission Possible
Westminster | Website
This event occurred in the past.
This special date night for families celebrates National Family Day with a spaghetti dinner & family activities. After the meal, a break out session for parents & older students will be presented by the Carroll State's Attorney's Office with the movie "Heroin Still Kills." Younger children will enjoy crafts and activities during the breakout session. There's no cost, but registration is required by Sept. 17. A free-will offering will be taken.


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City of God print by Cheba
City of God print by Cheba
from 85.00
New print by street artist Cheba.
Available in 2 sizes 50 x 50 or 90 x 90 CM
Each size edition of 11
Signed and numbered
Hand finished with pearlescent and gold acrylics.
Printed on Hahnemuhle German 310 gsm high quality fine art white textured paper
Prints are made to order and may take a few days longer to arrive
Add To Cart


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HomeEducation & MeetingsKidney Week
November 03, 2017 | 10:00 AM - 10:00 AM
Click an icon below to load this item into your calendar. Please note that times are exported as Coordinated Universal Time (UTC). Time zone help.
Maintenance of homogenous kidney tubular structures is critical to retain kidney shape and function, and in accordance aberrations in their structures are manifested in disease conditions. Lack of in vitro kidney tubule models that are homogeneous in tissue geometry structures and also emulating aberrations in disease conditions have limited the understanding of their pathology and the therapeutics development.
Homogenous tubular tissue geometry micropatterns were imprinted in different extracellular matrix combinations as an array using a combination of photolithography and soft lithography techniques. Tubular kidney epithelial cells were then cultured in the3D micro-molded extracellular matrix array and were evaluated for the morphogenic outcomes. Cystic kidney disease emulation was achieved by treatments with cAMP-elevating agents, while for acute kidney injury emulation cisplatin drug treatment was used.
Both mouse and human kidney epithelial cells formed homogeneous tubular structures as defined by the tissue geometry yielding kidney tubular arrays. The tubule features in the array were validated based on the characteristic distributionof actin (f-actin), cilia (acetylated tubulin),tight junction (ZO-1), and Na+K+-ATPasepump. Further, the disease emulations of cystic kidney disease and acute kidney injury were confirmed for tubule to cyst transformation and Kidney injury molecule (Kim-1) expression respectively.
The tubule array yielded organotypic homogenous tubules with in vivo- scale dimensions, which can be utilized for assessing nephrotoxicity of drugs and the mechanistic studies of various tubular kidney disease, making it an alternative to animal studies.
© American Society of Nephrology


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The Global Epidemic of TB
India 2013
June 21, 2013
Also published by Scientific American
A few weeks back, I shared with you of the first episode of a series of videos about the persistent and rising problem of the global epidemic of TB. Since then, the three final episodes have been released.
In February, I was in India with the International Reporting Project to look at issues of child survival, and saw, first hand, issues related to TB in many slum and settlement dwellers, where TB is estimated to affect 40% of the Indian population, most of the cases being latent, not active. This image on the right shows the microscope and staining reagents seen in the ever present small TB testing storefronts that are scattered throughout India. Anyone may stop by and submit a sample of sputum to have it visually assessed for the presence of Mycobacterium tuberculosis for a small fee.
EXPOSED is a four-part series of short films that tells the story of the deadly global epidemic of tuberculosis. The series focuses on current efforts to halt this airborne disease, which is growing more difficult to address, as well as the urgent movement to develop new tools to prevent it. By telling the stories of four inspiring individuals interspersed with expert commentary from some of the world's top TB physicians, scientists, advocates and policymakers, EXPOSED brings viewers to the forefront of the race against tuberculosis. For more information, go to aeras.org/exposed. EXPOSED is produced by Aeras, a nonprofit biotech advancing tuberculosis vaccines for the world. The series is developed in partnership with filmmaker Mo Scarpelli of RAKE FILMS with support from GATHER.
Chapter 2: The Rise of a Superbug
The second video in the EXPOSED series takes us to Delhi, India where we meet a doctor who treats TB in his practice. It then introduces us to a DOT center (Directly Observed Therapy, short-course).
From the video description: "Dr. Jayant Banavaliker, a leading TB doctor in Delhi, chronicles his daily struggle to save patients using today's antiquated tools: diagnostics barely more predictable than chance, drugs that can be toxic and weak against the pathogen, and a vaccine with limited protection. We also travel to South Africa, where Phumeza Tisile tells the harrowing story of her near-death experience with extensively drug-resistant TB."
Mumbai physician with mHealth device to assist with drug compliance in TB patients.
As seen in the above video, compliance with the extensive drug treatment for ever-growing-drug-resistant TB is a constant struggle. Fortunately, technology may be able to assist with compliance. With the penetrance of mobile technology being greater than 50% in India (with expected variability between cities, slums, settlements and rural areas), many health practitioners are employing various versions of mHealth (mobile health), which is the practice of medicine and public health, supported by mobile devices, to manage long term treatment, where multiple visits to physicians can be a hardship or inconvenience. Examples of mHealth use in India include sending mothers-to-be reminders of important check-ups and diet and care tips during pregnancy and after delivery, reminding mothers to bring children in for scheduled vaccinations, managing diabetes care and enforcing TB drug compliance. The image to the right shows a clinic in Mumbai and the physician with his device that sends reminders and receives data of medication taken daily for drug treatment by the TB patients in his care, saving them additional trips to the DOT center.
Chapter 3: The Innovation Movement
"Unathi Gwintsa lives with her daughter in Worcester, South Africa, a community devastated by TB. But she's not sitting back and waiting for the answer. She has volunteered in a clinical trial of one of a dozen TB vaccine candidates in clinical development. We follow her story along with others working tirelessly to rid their communities and the planet of TB."
Chapter 4: The Last Mile
"Professor Helen McShane has dedicated the last 12 years of her life to developing a new tuberculosis vaccine. We follow her team's unprecedented work at Oxford University and explore why now is a pivotal moment in history to save millions of lives and end the TB epidemic."
Joanne Manaster reported on health in India with the International Reporting Project. Manaster is a university level cell and molecular biology lecturer with an insatiable passion for science outreach to all ages. Enjoy her quirky videos at Joanne Loves Science and follow her on Twitter and Facebook.
More from this Reporter
- Can Science and Engineering Change Sanitation in India?
- A Visit to an India Full of Science and Engineering
- Childhood Malnutrition in India
- Childhood Vaccines in India
- Pediatric Infectious Diseases in India
- Why Am I Going to India?
From Other Reporters in This Country
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I have a new hampton bay sprigview ceiling fan that will not work after I replace the capacitor. The UPC is 718212149225. The fan ran for about 2 minutes and stops after I replaced the capacitor. Could it be the speed switch that needs to be replaced?
Hi stjunko, welcome to the community and glad you joined. Sorry to hear you're still having trouble with your fan after replacing the capacitor. Lets see how we can get your fan up and running again.
Since you have your UPC code, you can place a call to the Hampton Bay Customer Service Team and they will help you figure out the problem and how to fix it.
You can also order replacement parts while you're on the phone with them.
Here is the Hampton Bay Customer Service phone number. Hope this helps.
HI stjunko and thanks for your question. There is all kinds of things that could be wrong with your fan. The reversing switch, the speed control switch, the mutiprong connector, a wire loose at the ceiling and the motor is bad. You stated that it is a new fan. How new? Does the motor spin freely? Does the light work? You can get tech support at 1-877-527-0313. I hope this helps. Stukas
I am a master trades specialist with the Home Depot


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California Original, 1994 Chevrolet G-30, 1ton, 100% Rust Free, Needs TLC
- Condition: Used
- Make: Chevrolet
- Model: G30 Sport Van
- SubModel: No Reserve, G-30,100% Rust Free (310)259-5383
- Type: 12 Passenger Van
- Trim: No Reserve, Needs TLC,100% Rust Free (310)259-5383
- Year: 1994
- Mileage: 128146
- Color: White
- Engine size: 5.7 liter
- Number of cylinders: 8
- Fuel: Gasoline
- Transmission: Automatic
- Drive type: RWD
- Interior color: Brown
- Vehicle Title: Clear Want to buy? Contact seller!
1994 Chevrolet G30 Sport Van No Reserve, Needs TLC,100% Rust Free (310)259-5383
This is a California original, 1994 Chevrolet G-30, 1 ton, Sport Van with 128,146 original miles.
This van comes with a clean Auto check history, no known accidents, actual miles vehicle. The average Auto check score for similar vehicles is 39-58. This particular vehicle scored a high 66!(report available upon request)
The van would make a great project and needs work to make it turn key. Due to the high volume of restoration projects we have we will not be able to put anytime or effort into this one.
The van has been sitting for a number of years. The engine does not turn and will not start
The structural points and integrity of the van are A+.
-The entire undercarriage is dry and solid
-Nice glass all around
-Nice seating surfaces, carpets, headliner, door panels and headliner(small rip)
The exterior finish appears to be original. It still retains a nice luster, with various imperfections chips, nicks, dents, creases, scuffs, etc..
We were unable to test any of the other mechanical or electrical systems do to its non running status so bid accordingly.
Pictures speak a 1,000 words so what you see is what you get, nothing more, nothing less.
This 12 passenger van is definitely worth some time and effort due to it's beautiful rust free body.
Due to the non-driving status the van will NOT be offered to any California buyers. Out of state or country buyers only. We priority/overnight titles to out of state or country buyers titles immediately after payment is cleared.
This vehicle is also available for sale at other venues and is subject to removal from the auction at anytime should it be sold. If you have any questions call Manny 310-259-5383 or e-mail. We can arrange shipping and it can be expedited quickly and inexpensively to your door worldwide.
We do not offer the opportunity for inspections after the auction closes. All inspections are to be done during the auction. No exceptions. Thank you!


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A City Court judge Tuesday more than doubled bail for a 34-year-old city man accused of shooting two police officers and stabbing and shooting the mother of his two children.
Adam J. Hamilton, who police said also was wounded in the early morning shootout on South Avenue earlier this month, has been released from Erie County Medical Center and is now in the Niagara County Jail.
Hamilton's bail was increased as Officer Walter Nichols Jr., an 18-year veteran of the Niagara Falls Police Department, was released Tuesday from ECMC, where he underwent surgery to remove shotgun pellets from near his heart and stomach shortly after the incident.
Police have said Hamilton shot Nichols and Officer Michael D. Bird with a 12-gauge shotgun as they responded to a domestic violence call at about 1:15 a.m. Feb. 7.
Police have charged Hamilton with the shootings, and also with stabbing and shooting Stephanie C. Turk, 27, the mother of his two sons, before officers arrived on the scene.
City Court Judge Angelo J. Morinello increased Hamilton's bail to $500,000 cash or $1 million bond. Bail had been set at $200,000 cash or a $400,000 bond.
Hamilton has pleaded not guilty to two counts of first-degree attempted murder and one count of second-degree attempted murder.
He was out on bail on charges of third-degree assault and unlawful imprisonment when the shooting occurred. According to court records, Hamilton was accused of punching Turk on Jan. 20 and refusing to let her leave the 22nd Street apartment they had shared.
At the time, Turk told police she was staying with her mother because of trouble in her relationship with Hamilton and had returned to the apartment to get some of her belongings.
Turk told police that she signaled for help to her sister, who was waiting in a car outside, by pulling down the living room blinds during the Jan. 20 incident.
Morinello Tuesday also raised the bail for the assault charge, from $1,500 to $10,000.
Hamilton was waiting for Turk on Feb. 7 at the South Avenue home where she was staying, police said, and arrived there armed with the shotgun, a .22-caliber rifle, a knife and handcuffs.
He told police at ECMC after the shooting that he got the guns from "his old home" and that he did not know who owned them, according to court records.
Hamilton told investigators he wanted to marry Turk "to give her a better life," but when she left him he "could not take it anymore," court records said.
Turk, who was stabbed in the stomach and shot in the head, is still in ECMC, but has been moved out of intensive care.
Bird, who remains on leave, still has a shotgun pellet lodged in his face and is seeking medical treatment for it, Police Superintendent John R. Chella said.
Hamilton is scheduled to appear again in City Court on Feb. 27.
e-mail: firstname.lastname@example.com


text: GreggStar 1 Posted November 8, 2015 One of Secondlife's premier and competative priced real estate realtor, Established in November 2011 by the Hearts as a hobby which quickly grew into what we are today. We're proud to offer quality land at affordable prices, without cutting corners. Legally Licenced to trade as a business and registered with our local tax office.Registered Business No #813407400022Minnesota U.S.AWe have a number of Full Prim Regions, Homesteads and Openspace Regions around the Secondlife Grid, We also offer a selection of Mainland Parcels at a competative rate where you do not need a premium account to rent with us.Various lands to suit everyone's budget and style, from Residential Island plots with protected community water ways, to Commercial, Breedables and Club allowed regions. Weekly or Monthly payments or Paypal if you prefer, just speak with a member of our staff about how to pay via paypal. We also offer renaming and moving of our regions (if you have the full) at a extra cost (LL Charges).*~*~*~*~*~*~*~*~*~*~*~*~*~*~*~*~*~*~*~*~*~*~*~*~* About Our Land :-No Premium account is needed to rent / own with Stargazer Estates.Stand alone Full prim regions and Homesteads with full rights to your parcel (or EM rights if you take the compleate region)Relax with piece of mind from our fully trained staff and outstanding management team and support that actually cares about thier tenants.24/7 real time monitoring of all Stargazer Estates regions, powered by our very own scanner (devoloped by Damienthor) Visit our Offices for many more plots of all sizeshttp://maps.secondlife.com/secondlife/Beregovoi/191/50/402 Features :-No Premium account is needed to rent / own with Stargazer Estates.All our regions are updated at time of purchase and constantly updated for the newest server software in Secondlife.Easy to use billing system via our ATM, payments avalible in L$ or USD (Paypal) if you prefer.24/7 Real time region monitoring (if any region is offline for more than 10 minuets our system automaticly emails Linden Labs support.Stargazer Estates is a company that is quickly growing with the intentions of being one of the biggest companies in Second Life.Quality out of world training for all staff is provided in efforts to guarantee that all staff are able to assist tenants quickly and professionally without need of waiting for a manager or owner, on most problems that arise. Constant in game training to further staffs knowledge to provide even better customer service with tenants in order to provide a better place to call home.At Stargazer Estates we have extended concierge support through Linden Labs for quick turn around on problems outside of our control, i.e. server changes, tenants paid name changes for full regions, and land purchases. This concierge support allows for all cases to be responded to within a few hours or even immediately in cases of emergencies during normal operating hours.Stargazer Estates holds a celebration every year with all profits going to the Relay for Life organization in Second Life and all extra proceeds out of game go to a different charity per month. Contact :-Website: - http://www.stargazer-estates.comLive Land Book : http://www.stargazer-estates.com/secondlife/vacant_all.phpLooking for a certain cut? visit to our office we may be able to help you out.********************************************************************************************************************Darkness Rises#02Full PrimADULT750032768 Sqm9500 L$VisitZanzo#01Full PrimGENERAL375016384 Sqm4500 L$VisitConstellation Valley#02Full PrimMODERATE375016384 Sqm4500 L$VisitDragovar Dynasty Estates#06Full PrimMODERATE18758192 Sqm2495 L$VisitStargazer Bay#03Full PrimMODERATE18758192 Sqm2495 L$VisitPortscatho#03 (Corner)Full PrimMODERATE18758192 Sqm2595 L$VisitDragovar Dynasty Estates#06aFull PrimMODERATE18758192 Sqm2495 L$VisitConstellation Valley#03Full PrimMODERATE18758192 Sqm2495 L$VisitConstellation Valley#04 (Corner)Full PrimMODERATE18758192 Sqm2595 L$VisitIsola Gavia#07Full PrimMODERATE12305376 Sqm1599 L$VisitRavens Ridge#11Full PrimMODERATE9374096 Sqm1395 L$VisitRavens Ridge#08Full PrimMODERATE9374096 Sqm1395 L$VisitStargazer Fields#04Full PrimMODERATE9374096 Sqm1395 L$Visit * Powered by stargazer enterprises parcel managment systemView our land book for more lands updated every 30 mins: http://www.stargazer-estates.com/secondlife/vacant_all.php Visit our Officeshttp://maps.secondlife.com/secondlife/Beregovoi/191/50/402More sizes and quality vacant plots up for rent.Specific Cuts/ Prims are a possability, speak to any member of our team. ******************************************************************************************************************** Stargazer Etates is proud to be featured in the Secondlife Destination Guide. Testermonials :- Stargazer Estates are the best people in the world to work with! I had rented off of one that simply sent my meeroos back to me without warning. Stargazers welcomes breedables and anything wrong they are easy to get a hold of and very willing to help! 5 *****s to the Stargazer team !- LaineyStickley ******************************************Stargazer Estates is a great place to live. The managers of the Estates are great and keep you very informed on what is going on with the sim you live on. I would definitely recommend Stargazer Estates to anyone who is looking for a nice place to live.- Hermione Lane ****************************************************************************************************************** Quote Share this post Link to post Share on other sites


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We are a small team of experienced stylists and assistants who provide all aspects of hair styling for all ages at very competitive prices. If you are looking for a ladies hair salon in Aberdeen thenwe believe you have come to the right place! Our small, friendly salon offers 15% OAP discount days and is easily accessible with ample free parking. Book your appointment with one of our stylists today and receive a 15% discount and a free Joico Conditioning Treatment (new customers only).
Open 5 days a week, Heaven Hair and Beauty is a warm, friendly Salon to come and have all your Hair, Nail and Beauty Treatments taken care of by a staff of talented professional angels. Situated inthe Woodend area of Aberdeen, you'll find we have unrivalled facilities, in fact clients often tell us that Heaven Hair and Beauty is a city salon without the city costs and parking. You'll also be pleased to know we also have free parking right at the door for our clients which is another bonus.
Step inside our Mid Stocket Road location and you will discover our beautiful and unique hair and beauty salon. Our carefully selected team specialise in cutting edge hairdressing and beautytreatments, and ensure your experience starts with us the moment you walk through the doors. Utopia first opened it's doors in 1997 and we've been welcoming clients old and new ever since.
Ishoka Hairdressing and Beauty epitomises and encapsulates high quality service and customer care. The salon was opened in 1986 by business partners and hair experts, Philip Bell, Malcolm MacNeil andKathryn Longmuir with one aim; to bring London quality hairdressing to Aberdeen. The salon's friendly team and warm atmosphere ensure there is a personal relationship and connection with each and every client.
Cach-ca Hair Studio is a fabulously modern hairdresser based in Rosemount Aberdeen catering for both Male & Female clients. Our aim is simple - to make your visit to the salon an experience to lookforward to. We want you to leave totally satisfied with your treatments, which have been carried out in a professional and friendly manner.
I have been fortunate to work with several mobile wedding hairdressers in the Aberdeen area and hairstylists in Westhill, Kingswells and Stonehaven. Often travelling between Aberdeen, Inverurie,Westhill and Stonehaven, I have discovered plenty of inspirational people who are employed within the industry.
Looking for a hairdresser based in Aberdeen, Scotland, look no further than Vanilla Style. A unisex hairdressing salon that offers a professional service in a friendly relaxed atmosphere, just 5minutes walk from Aberdeen city centre. Our professional, vibrant stylists will consult with you to provide a high standard of styling and colouring to your requirements.
Ellee's Hair Design was established on 27th October 2009 By the Mckay Family. Over the past 4 years the team & the salon have evolved into a strong market leader for hair & beauty within Aberdeen &have achieved National Awards In Scotland & in London. The Ellee's Hair Design team created the Customer Charter which ensures, that every client will recieve the best possible customer care experience within their salon experience.
Our Salon is in the heart of the West end of Aberdeen. We moved to new premises at 15 Thistle Street in 2006. This was to allow us to have a larger floor space, beauty room and automated spray tanningfacilities. The salon is independently owned by Claire she has built it for the 11 years.
At Beautylicious Hair and Beauty Salon Aberdeen our goal is to pamper you from head to toe and to bring out your individual, natural beauty. The experienced and talented staff at Beautylicious areamong the most qualified in the industry. Our Hair and Beauty Salon is Located just up from His Majesty's Theatre in Aberdeen City Centre, close to all Shopping Centre car parks and the Denburn car park and 10 minutes from Aberdeen Train Station for those travelling from Aberdeenshire and further afield.
We are a exceptionally friendly salon with a relaxed atmosphere, specialising in all areas of Hairdressing including Hair Extensions and Weaves. Our Beauty Boutique offers a full range of treatments.


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With that in mind, here is the insight you need into making the decision about what style of eyewear to choose…
Round Face
When selecting your eyewear avoid round frames, instead try on angular shapes to lend definition to your face.
Oval Face
Focus on frames that enhance your features, and avoid thin frames that may elongate your face.
Heart Shaped Face
Draw attention to your eyes by balancing a pointy chin with frames that are thicker on the bottom, to add the appearance of width.
Hair + Eye + Wardrobe
Color is a matter of style of course but there is always a color that can compliment your hair color, your eyes and your wardrobe. Do not be afraid to try different colors and hues to influence the style you want to project.
By measuring you can make sure the glasses you choose will fit. Looking to transform your look, try on different sizes to see how it transforms your face and your overall look.
Will change, whether you are going for a more casual look or something a little more sophisticated take time, have fun, and experiment with something new.
If you are in search of a new pair of specs and need assistance finding a pair that fits you, we are here to help! Our team will help make your experience easy and seamless!


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12 Twine Court Port Geographe
Live the Marina Lifestyle!
A wonderful opportunity to create the home of your dreams in a superb waterside location! Create your contemporary masterpiece and enjoy the best of the South West in the celebrated Port Geographe Marina.
This generous north-facing block takes in sweeping views of the canals, with a beautiful water vista that makes every day a pleasure. It's superbly situated in a quiet cul de sac location with flexible building covenants, and easy access to private jetties and canal entrance.
There are restaurants, cafes, beaches and lush nature reserves all within walking distance, and it's a short ride in your tender to the main pens and open water. The popular Port Geographe estate is just 8 minutes from Bursselton's main street and famous jetty.
Busselton is the gateway to the beautiful South West, and offers a world class lifestyle of wineries, galleries, surf and stunning countryside. Enjoy the many international festivals, countless social events, sports clubs and laid-back lifestyle of this wonderful area.
Whether you're keen to build your own family masterpiece or the perfect retirement/holiday home, this is the opportunity you've been waiting for!
For more information
0438 166 894


text: I had a machine with this card as well. At the time I looked, the documentation on xfree86.org said that this card is supported in the developement version of X 4.?.?, but not currently supported in the official release, but will be in the next official release. Gerry On Tue, 21 Nov 2000, Daniel Borgmann wrote: > Hello list! > > i recently helped my father to install debian (potato r0) on his box. > everything was fine, but X is a problem. i have to say this was ever a > problem since he upgraded his graphic card to a hercules 3d prophet which is > using the geforce MX chipset. > he tried mandrake before but when it comes to auto detection, it damages the > display (not physically of course) so i can't even use the textmode anymore. > this is really ugly... > now with debian there is a similar problem. i used XF86Setup. in "cards", i > used the details, not one of the list and left everything on probing. > after starting the x server, everything was foobared again... > i know, that the geforce MX ist NOT supported with xfree 3 but i still think > it should be possible to run x in a kind of compatiblity mode isn't it?? > i don't care about all the fancy geforce features (in fact my father's > favorite game is nethack, so he's not in a hurry to install x, but it would > be really nice to show him the great new kde2 desktop!), but it should be > able to render 1024*768 with 32 bit colors in a reasonable speed. > what does i have to do exactly to make this work? > btw, it already worked using the VGA16 server and a generic vga driver. > but 640*480 is not a sollution ;) > i hope i can show him now how great the debian support is.. :)) > > -Daniel > > > -- > Unsubscribe? mail -s unsubscribe email@example.com < /dev/null >


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Social Scene 1.1
Social Scene lets you spend less time bouncing to different social networks, and more time socializing by putting Facebook and Twitter in a helpful sidebar, built right into an ultra-fast web browser!
With Social Scene you get quick and straightforward access to your networks without any hassle, while also getting the freedom to browse the internet - all on one screen. With everything in one app, you can get info posted quickly, while browsing!
Info updated on:


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MONTEBELLO – As of Monday, drivers who run the red light at the Montebello and Paramount boulevards intersection will get a $381 citation in the mail.
Enforcement of a new red light camera program began at 6 a.m. at the intersection, following a two-month grace period in which violators received warnings, said retired Montebello Police Department Sgt. Craig Powers, who is charge of the program.
"The intersection is one of the top three intersections that studies show has the highest rates of accidents," he said.
For more on this story, pick up Tuesday's Whittier Daily News.


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Vitamin C – Not just for the common cold
It is a widely accepted belief that when you feel the start of a cold coming on it is advisable to start popping Vitamin C to help nip it in the bud. Dr. John Ely, a researcher at the University of Washington says, "Ten grams of Vitamin C on the first day of a cold always works, after that it never works." I have never personally tried this dosage however, many people swear by its efficacy. Vitamin C is exceptionally easy to absorb, and since it is water soluble the kidneys eliminate what we don't need, therefore it can safely be taken in high doses.
Looking beyond curtailing the common cold, Vitamin C is indispensable in the body. Its primary function is the manufacture of collagen – the main protein of our body. Collagen is found in our skin, providing a youthful firmness. Unfortunately, as we age our body simply does not produce the amount of collagen it once did back in our twenties. I am reminded of this whenever my beautiful teenage daughter and I stand in front of a mirror together. Vitamin C not only supports firm skin, it works to build cartilage, connective tissue, ligaments, and tendons, which keep our bodies pliable and joints supple with movement.
We already know that vitamin C supports our immune system, but to what degree may be underestimated. Back in the 1980s, Dr. Linus Pauling brought fame and distinction to Vitamin C by keeping terminal cancer patients alive for an extended period of time with twenty grams of the vitamin per day. Vitamin C is vital for the functioning of our white blood cells fighting bacteria, viruses, cardio vascular disease, and may even reduce the risk of developing cataracts.
As a powerful antioxidant, Vitamin C protects us against pollution, cigarette smoke and carcinogens. It supports the liver in the clearance of toxins in the bloodstream. Dr. Walter Crinnion, author of Clean, Green and Lean, recommends city dwellers take at least 3,000 milligrams of Vitamin C each day to guard against pollution.
A few things to look out for as you strive to preserve your Vitamin C levels.
During times of physical, emotional and chemical stress the excretion of Vitamin C is increased through the urine. Remember to increase your Vitamin C consumption during times of high stress. Examples of chemical stresses are pollution and cigarette smoke.
Vitamin C is not only destroyed by heat but also diminished when exposed to air. Although a salad bar is a healthy lunch choice, its Vitamin C content is only a fraction of what it would be if the salad was made fresh and consumed after preparation. Consuming a portion of raw vegetables at each meal ensures that you are getting all the goodness of the vitamin content.
Alcohol, excessive sugar and many common drugs including aspirin and oral contraceptives reduce Vitamin C levels in the body.
Great Sources of Vitamin C
Citrus fruits typically come to mind as the best source of Vitamin C, but many vegetables and a few fruits outrank citrus in content.
In order of Vitamin C content:
- papayas
- bell peppers
- broccoli
- Brussels sprouts
- strawberries
- pineapple
- oranges
A simple way to boost your Vitamin C level each day is to add a few slices of lemon or strawberries to your drinking water.
In addition to my whole foods diet, I take 2000+ milligrams (mg) of Vitamin C daily and have successfully weathered our Northern California winter rains this season with barely a sniffle. My favorite supplement is from Thorne Research available on my site under resources. It is an effective, easy-to-use Vitamin C powder, buffered with calcium, magnesium, and potassium, making it gentle on the digestive system. I simply add a teaspoon of powder to a glass of lemon water or a smoothie and I am good to go!
- Clean, Green and Lean by Walter Crinnion
- The Encylopedia of Healing Foods by Michael Murray
- The World's Healthiest Foods website


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A 70-year-old man was face down in the water and unconscious Tuesday morning when lifeguards pulled him out of Mission Bay following an apparent boating accident, authorities said.
Support Times of San Diego's growth
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The man suffered a head injury a little before 8 a.m. and was face down in the water near Fiesta Island "for a few minutes," San Diego Fire-Rescue Department spokesman Jose Ysea said.
Lifeguards sped to the scene in a boat and pulled the man from the water, then helped stabilize him aboard the private boat involved in the accident, Ysea said. The man was unconscious but breathing when pulled from the water, and initial reports said he regain consciousness and was speaking with his rescuers a few minutes later.
The lifeguards took him to shore on Fiesta Island, where paramedics transported him to UCSD Medical Center, Ysea said. The extent of his injuries were not immediately known, and lifeguards were investigating how he was injured.
—City News Service
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John Deere Plow. $39, USD Livingston, MT. Updated 3 Days Ago 8 Bottom, Pull Type, Hydraulic Variable Bottoms, Coulters. More. Because it's a pull behind plow, a 3 point hitch tractor is not required. A hydraulic cylinder is It is a John Deere number 55A-B-H, 3 bottom plow. The front.
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Design examples (scroll down) include magazine layouts.
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Campus, Fall 2015
Sunset on Stacks, Fall 2015
University Reacts to Pro-Trump Chalkings, Spring 2016
Awarded SPJ Mark of Excellence for Breaking News Photography (Region 3)


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Bedroom Classics, Vol. 1
5.0 1 5 1
USD 4.99$4.99 View All Available Formats & Editions
Few singers could have kicked off Rhino's Bedroom Classics series as fittingly as Teddy Pendergrass, but this is the umpteenth time he has been anthologized; it's not as if the planet has been starving for a strong summation of his baby-making material. That said, a good deal of care was used in putting this particular compilation together. While not as thorough as 2003's Anthology (The Right Stuff), it's a fine introduction -- despite the fact that it's built around a theme. (After all, the bedroom was Pendergrass' theme.) Plenty of enduring songs are here, including "Close the Door," "Joy," "It Should've Been You," "Hold Me," and "This Is My Last Time."
Product Details
|Release Date:
|Rhino Flashback
Album Credits
Performance CreditsTeddy Pendergrass Primary Artist
Technical CreditsMiles Jaye Producer
Gerald Levert Composer,Producer
Teddy Pendergrass Composer,Producer
Luther Vandross Composer,Producer
David Gates Composer
Gerry Goffin Composer
Gabriel Hardeman Composer
Reggie Calloway Composer,Producer
Vincent Calloway Composer,Producer
James Carter Composer,Producer
Linda Creed Composer,Producer
Joel Davis Composer
Kenny Gamble Composer,Producer
Annette Hardeman Composer
Leon Huff Composer,Producer
Stephanie Huff Composer
Nathaniel Lee Composer
Nick Martinelli Producer
Michael Masser Composer,Producer
Dennis Matkosky Composer,Producer
Marcus Miller Composer
Bill Neale Composer,Producer
Cynthia Weil Composer
Terry Price Composer,Producer
Barry Benson Producer
Linda C. Cobb Art Direction
Tracey New Liner Notes
Kevin J. Askins Composer
Marvin Hammett Composer
Edwin "Tony" Nicholas Composer,Producer
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