Use my Search Websuite to scan PubMed, PMCentral, Journal Hosts and Journal Archives, FullText. Kick-your-searchterm to multiple Engines kick-your-query now !>

A dictionary by aggregated review articles of nephrology, medicine and the life sciences Your one-stop-run pathway from word to the immediate pdf of peer-reviewed on-topic knowledge.

10.3389/fimmu.2018.01378 http://scihub22266oqcxt.onion/10.3389/fimmu.2018.01378 C6034003!6034003!30008714     free     free     free Deprecated: Implicit conversion from float 231.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 231.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 231.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 231.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 231.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 231.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534       Front+Immunol 2018 ; 9 (ä): ä Nephropedia Template TP {{tp|p=30008714|t=2018. Optimized Peptide?MHC Multimer Protocols for Detection and Isolation of Autoimmune T-Cells |pdf=|usr=}}{{30008714}} gab.com Text Optimized Peptide MHC Multimer Protocols for Detection and Isolation of Autoimmune T-Cells JO: Front Immunol http://www.kidney.de/pget.php?&tw=1&pmid=30008714 #FOAvip Peptide?MHC (pMHC) multimers have become the ?gold standard? for the detection and isolation of antigen-specific T-cells but recent evidence shows that normal use of these reagents can miss fully functional T-cells that bear T-cell receptors (TCRs) with low affinity for cognate antigen. This issue is particularly pronounced for anticancer and autoimmune T-cells as self-reactive T-cell populations are enriched for low-affinity TCRs due to the removal of cells with higher affinity receptors by immune tolerance mechanisms. Here, we stained a wide variety of self-reactive human T-cells using regular pMHC staining and an optimized technique that included: (i) protein kinase inhibitor (PKI), to prevent TCR triggering and internalization, and (ii) anti-fluorochrome antibody, to reduce reagent dissociation during washing steps. Lymphocytes derived from the peripheral blood of type 1 diabetes patients were stained with pMHC multimers made with epitopes from preproinsulin (PPI), insulin-? chain, glutamic acid decarboxylase 65 (GAD65), or glucose-6-phospate catalytic subunit-related protein (IGRP) presented by disease-risk allelles HLA A*02:01 or HLA*24:02. Samples from ankylosing spondylitis patients were stained with a multimerized epitope from vasoactive intestinal polypeptide receptor 1 (VIPR1) presented by HLA B*27:05. Optimized procedures stained an average of 40.5-fold (p?=?0.01, range between 1.4 and 198) more cells than could be detected without the inclusion of PKI and cross-linking anti-fluorochrome antibody. Higher order pMHC dextramers recovered more cells than pMHC tetramers in parallel assays, and standard staining protocols with pMHC tetramers routinely recovered less cells than functional assays. HLA A*02:01-restricted PPI-specific and HLA B*27:05-restricted VIPR1-specific T-cell clones generated using the optimized procedure could not be stained by standard pMHC tetramer staining. However, these clones responded well to exogenously supplied peptide and endogenously processed and presented epitopes. We also showed that anti-fluorochrome antibody-conjugated magnetic beads enhanced staining of self-reactive T-cells that could not be stained using standard protocols, thus enabling rapid ex vivo isolation of autoimmune T-cells. We, therefore, conclude that regular pMHC tetramer staining is generally unsuitable for recovering self-reactive T-cells from clinical samples and recommend the use of the optimized protocols described herein. Twit Text FOAVip Optimized Peptide MHC Multimer Protocols for Detection and Isolation of Autoimmune T-Cells ????Front Immunol http://www.kidney.de/pget.php?&tw=1&pmid=30008714 #FOAvip Twit Text # Free Paper on # # # # # LINK http://www.kidney.de/pget.php?&tw=1&pmid=30008714 #FOAvip English Wikipedia <ref>{{Cite pmid|30008714}}</ref> Optimized Peptide?MHC Multimer Protocols for Detection and Isolation of Autoimmune T-Cells #MMPMID30008714 Dolton G; Zervoudi E; Rius C; Wall A; Thomas HL; Fuller A; Yeo L; Legut M; Wheeler S; Attaf M; Chudakov DM; Choy E; Peakman M; Sewell AK Front Immunol 2018[]; 9 (ä): ä PMID30008714show ga Peptide?MHC (pMHC) multimers have become the ?gold standard? for the detection and isolation of antigen-specific T-cells but recent evidence shows that normal use of these reagents can miss fully functional T-cells that bear T-cell receptors (TCRs) with low affinity for cognate antigen. This issue is particularly pronounced for anticancer and autoimmune T-cells as self-reactive T-cell populations are enriched for low-affinity TCRs due to the removal of cells with higher affinity receptors by immune tolerance mechanisms. Here, we stained a wide variety of self-reactive human T-cells using regular pMHC staining and an optimized technique that included: (i) protein kinase inhibitor (PKI), to prevent TCR triggering and internalization, and (ii) anti-fluorochrome antibody, to reduce reagent dissociation during washing steps. Lymphocytes derived from the peripheral blood of type 1 diabetes patients were stained with pMHC multimers made with epitopes from preproinsulin (PPI), insulin-? chain, glutamic acid decarboxylase 65 (GAD65), or glucose-6-phospate catalytic subunit-related protein (IGRP) presented by disease-risk allelles HLA A*02:01 or HLA*24:02. Samples from ankylosing spondylitis patients were stained with a multimerized epitope from vasoactive intestinal polypeptide receptor 1 (VIPR1) presented by HLA B*27:05. Optimized procedures stained an average of 40.5-fold (p?=?0.01, range between 1.4 and 198) more cells than could be detected without the inclusion of PKI and cross-linking anti-fluorochrome antibody. Higher order pMHC dextramers recovered more cells than pMHC tetramers in parallel assays, and standard staining protocols with pMHC tetramers routinely recovered less cells than functional assays. HLA A*02:01-restricted PPI-specific and HLA B*27:05-restricted VIPR1-specific T-cell clones generated using the optimized procedure could not be stained by standard pMHC tetramer staining. However, these clones responded well to exogenously supplied peptide and endogenously processed and presented epitopes. We also showed that anti-fluorochrome antibody-conjugated magnetic beads enhanced staining of self-reactive T-cells that could not be stained using standard protocols, thus enabling rapid ex vivo isolation of autoimmune T-cells. We, therefore, conclude that regular pMHC tetramer staining is generally unsuitable for recovering self-reactive T-cells from clinical samples and recommend the use of the optimized protocols described herein. äOptimized Peptide?MHC Multimer Protocols for Detection and Isolation o(epmc).pdf DeepDyve Pubget Overpricing