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expires=' + expires.toGMTString() + '; domain=.' + this.getMasterDomain() + ';'; }, /** * get the master domain based on business rules */ getMasterDomain : function () { // default the master domain var masterDomain = 'thermofisher.com'; // check for alternate domain portion of URL var regexMatch = document.location.hostname.match(/\.(thermofisher)\.(cn)/i); if (regexMatch !== null) { masterDomain = regexMatch[1] + '.' + regexMatch[2]; } return masterDomain; }, /** * Check if init has been initiated once * @type {Boolean} */ initted: false, /** * @return {Array} method returns an array of a given object's key properties */ init: function () { this.page.country = this.getCookie("CK_ISO_CODE") || 'us'; this.page.language = this.getCookie('CK_LANG_CODE') || 'en'; this.page.siteName = 'thermo'; //hard coded this.initMerchandisingData(); this.initUserData(); this.displayDigitalData(); this.formSubmitEvent(); }, //specific function to set type of the page // need specific function to keep page type separate from other page attributes setPageType: function (type) { this.setPageAttribute('type', type); 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class="nav-list-arrow">&rsaquo;</span></a></li><li class="item-no-children"><a class="" href="/sg/en/home/life-science/cell-analysis/cell-analysis-learning-center/immunology-at-work/inflammation-overview.html">Inflammation Overview <span class="nav-list-arrow">&rsaquo;</span></a></li><li class="item-no-children"><a class="" href="/sg/en/home/life-science/cell-analysis/cell-analysis-learning-center/immunology-at-work/cytokine-storm-overview.html">Cytokine Storm Overview <span class="nav-list-arrow">&rsaquo;</span></a></li></ul><script> if ( $('#navListActive').children('ul').length > 0 ) { var activeNav = $j('#navListActive'); $j('#navList').prepend(activeNav); } </script></div></div><div class="section"><div class="new"></div></div><div class="iparys_inherited"><div class="railInheritParsys iparsys parsys"></div></div></div><div class="railParsys parsys"></div></div></div><div class="span9 main" role="main"><div class="MainParsys parsys"><div class="heading section"><h2> What is a regulatory T cell (Treg)? </h2></div><div class="text parbase section"><p>Regulatory T cells (Treg) are critical to the maintenance of self-tolerance and immune cell homeostasis, which is demonstrated by the severe consequences of a lost or nonfunctional Treg population, as occurs in immune dysregulation polyendocrinopathy enteropathy X-linked syndrome (IPEX). Treg have also been found to play an important role in regulating immune responses responsible for autoimmune diseases, including type 1 diabetes, rheumatoid arthritis, multiple sclerosis, systemic lupus erythematosus (SLE), and myasthenia gravis.</p></div><div class="reference parbase section"><div class="cq-dd-paragraph"><div class="text_8fc4 text parbase"><div class="alert alert-info"><h4><a href="/sg/en/home/global/forms/life-science/help-flow-cytometry-immunophenotyping-panel.html">Request FREE Flow Cytometry Panel Design Help</a></h4></div></div></div></div><div class="well-cmp section"><div class="well "><div class="well-container" style=""><div class="well-content"><div class="parsys"><div class="heading section"><h3> On this page: </h3></div><div comp-order="bycol" class="parsys_column cq-colctrl-lt0"><div class="parsys_column cq-colctrl-lt0-c0"><div container-id="cq-colctrl-lt0" comp-id="A1"><div class="text parbase section"><ul><li><a href="#classification">Treg classification and nomenclature</a></li><li><a href="#disease">Tregs in disease</a></li><li><a href="#isolation">Isolation and sorting of viable Tregs</a></li></ul></div></div></div><div class="parsys_column cq-colctrl-lt0-c1"><div container-id="cq-colctrl-lt0" comp-id="B1"><div class="text parbase section"><ul><li><a href="#ex-vivo">Treg <i>ex vivo</i>: expansion, differentiation, and function</a></li><li><a href="#CytokineP">Cytokine profiling</a></li><li><a href="#references">References</a></li></ul></div></div></div></div><div style="clear:both"></div></div></div></div></div></div><div class="heading section"><h2 id="classification"><br>Regulatory T cell classification and nomenclature </h2></div><div class="textimage parbase textimage-right section"><div class="image" id="cq-textimage-jsp-cb55b0fa-3021-4418-b36c-884782fec6a0MainParsystextimage_21ee"><a class="image-enlarge-link" 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alt='Maturing of T cells to multiple types of regulatory t cell subsets depending on where the cell is located and it’s activated state.' title='Artistic illustration of regulatory t cell subsets'></picture></div><style> #cq-textimage-jsp-cb55b0fa-3021-4418-b36c-884782fec6a0MainParsystextimage_21ee { max-width: 100%; } #cq-textimage-jsp-cb55b0fa-3021-4418-b36c-884782fec6a0MainParsystextimage_21ee img { width: 100%; max-width: 750px; } </style><script type="text/javascript"> $( document ).ready(function() { if(!!window.picturefill){ window.picturefill(); } }); </script></a><div class="caption"><small><span class=" icon-zoom-in"> </span>Click image to enlarge</small></div></div><div class="seo-event-text" data-seo-schema='' data-seo-element='description'><div class="text"><p>Classification and nomenclature of Treg have been standardized and include three main Treg populations: (1) Thymus-derived Treg (tTreg), formerly referred to as natural Treg, differentiate from two CD4 single-positive progenitor subsets that are characterized as CD25<sup>+</sup> Foxp3<sup>–</sup> or CD25<sup>–</sup> Foxp3<sup>low</sup>. (2) Peripherally derived Treg (pTreg) are induced following antigen stimulation of naive T cells in peripheral lymphoid tissues, such as gut-associated lymphoid tissues. (3) <i>In vitro</i>–generated Treg (iTreg) are derived from naive CD4<sup>+</sup> T cells stimulated with anti-CD3 in the presence of IL-2 and TGFβ. While no single marker distinguishes between these three subsets, the expression of Foxp3 is essential for their suppressive function.</p><p>The identification and characterization of Treg are complicated by their ability to take on phenotypic and transcriptional characteristics of effector T cells. For example, Treg that must control immune responses in the gut will express RORγt and CD196 (CCR6), in addition to Foxp3 and CD25, while Treg in visceral adipose tissue will express Foxp3 and PPARγ. Such plasticity is driven by cytokines such as IL-6, IL-1, TNFα, an IL-23, and it allows Treg to respond to tissue-specific chemoattractants, migrate to the active site, and regulate the corresponding immune response. Much of what we know about the phenotype and gene signatures of Treg come from studies in mouse models. Studies of Treg in humans are further complicated by the fact that the expression of Foxp3 and other surface markers of Treg can be transiently induced at low levels on conventional T cells upon stimulation of the T cell receptor (TCR). Studies of epigenetic modifications found that CpG islands located in the Treg-specific demethylation region (TSDR) of the Foxp3 locus were demethylated in tTreg and pTreg but methylated in iTreg and conventional T cells with low levels or no expression of Foxp3. Thus, demethylation of the TSDR is considered the bona fide method for identifying tTreg & pTreg; the search for surface markers that will enable sorting of specific, live Treg continues.</p></div></div><div class="clearfix"></div></div><div class="text parbase section"><h4>Table 1. Properties of Treg cells</h4><div class="table-component-wrapper"><table class="rte-table" id="table-743037225"><tbody><tr><th>Property</th><th style="width: 28%;">Thymus-derived Treg (tTreg)</th><th style="width: 28%;">Peripherally derived Treg (pTreg)</th><th style="width: 28%;"><i>In vitro</i>–generated Treg (iTreg)</th></tr><tr><td><b>Development</b></td><td>Thymus</td><td>Periphery (primarily gut)</td><td><i>In vitro</i></td></tr><tr><td><b>Progenitor cell</b></td><td><a href="/antibody/primary/query/CD4">CD4</a> single-positive cells<br><a href="/antibody/primary/query/CD25">CD25</a><sup>+</sup><a href="/antibody/primary/query/Foxp3">Foxp3</a><sup>–</sup> or <a href="/antibody/primary/query/CD25">CD25</a><sup>–</sup><a href="/antibody/primary/query/Foxp3">Foxp3</a><sup>low</sup></td><td>Naive <a href="/antibody/primary/query/CD4">CD4</a><sup>+</sup> cells</td><td>Naive <a href="/antibody/primary/query/CD4">CD4</a><sup>+</sup> cells</td></tr><tr><td><b>Surface phenotype</b></td><td><a href="/antibody/primary/query/CD4">CD4</a><sup>+</sup><a href="/antibody/primary/query/CD25">CD25</a><sup>hi</sup> CD127<sup>low</sup><a href="/antibody/primary/query/CD62L">CD62L</a><sup>hi</sup><a href="/antibody/primary/query/CD44">CD44</a><sup>low</sup><a href="/antibody/primary/query/CD45RB">CD45RB</a><sup>low</sup> (mice) <a href="/antibody/primary/query/CD4">CD4</a><sup>+</sup><a href="/antibody/primary/query/CD25">CD25</a><sup>hi</sup><a href="/antibody/primary/query/CD127">CD127</a><sup>low</sup> or<br><a href="/antibody/primary/query/CD4">CD4</a><sup>+</sup><a href="/antibody/primary/query/CD25">CD25</a><sup>hi</sup><a href="/antibody/primary/query/CD45RO">CD45RO</a><sup>hi</sup> (human)</td><td><a href="/antibody/primary/query/CD4">CD4</a><sup>+</sup><a href="/antibody/primary/query/CD25">CD25</a><sup>+</sup><a href="/antibody/primary/query/CD62L">CD62L</a><sup>low</sup><a href="/antibody/primary/query/CD44">CD44</a><sup>hi</sup></td><td><a href="/antibody/primary/query/CD4">CD4</a><sup>+</sup><a href="/antibody/primary/query/CD25">CD25</a><sup>+</sup><a href="/antibody/primary/query/CTLA-4">CTLA-4</a><sup>+</sup><a href="/antibody/primary/query/CD62L">CD62L</a><sup>low</sup></td></tr><tr><td><b>TSDR methylation status</b></td><td><a href="/order/catalog/product/A43675">Demethylated</a></td><td><a href="/order/catalog/product/A43675">Demethylated</a></td><td><a href="/order/catalog/product/A43675">Methylated</a></td></tr><tr><td><b>Transcription factors</b></td><td><a href="/antibody/primary/query/Foxp3">Foxp3</a><br><a href="/antibody/primary/query/c-Rel">c-Rel</a><br><a href="/antibody/primary/query/EOS">EOS</a><br><a href="/antibody/primary/query/Helios">Helios</a><br><a href="/antibody/primary/query/STAT5">STAT5</a></td><td><a href="/antibody/primary/query/Foxp3">Foxp3</a><br><a href="/antibody/primary/query/GATA-3">GATA-3</a> (subset)<br><a href="/antibody/primary/query/Helios">Helios</a> (subset)<br><a href="/antibody/primary/query/IRF4">IRF4</a> (subset)<br><a href="/antibody/primary/query/pparg">PPARγ</a> (subset)<br><a href="/antibody/primary/query/rorg">RORγt</a> (subset)<br><a href="/antibody/primary/query/STAT5">STAT5</a><br><a href="/antibody/primary/query/T-bet">T-bet</a> (subset)</td><td><a href="/antibody/primary/query/Foxp3">Foxp3</a><br><a href="/antibody/primary/query/Helios">Helios</a> (subset)<br><a href="/antibody/primary/query/STAT5">STAT5</a></td></tr><tr><td><b>Other associated surface markers</b></td><td><a href="/antibody/primary/query/CD31">CD31</a> (subset)<br><a href="/antibody/primary/query/CD45RA">CD45RA</a> (subset)<br><a href="/antibody/primary/query/cd49d">CD49d</a><br><a href="/antibody/primary/query/cd101">CD101</a><br><a href="/antibody/primary/query/cd103">CD103</a><br><a href="/antibody/primary/query/cd121a">CD121a</a> (activated)<br><a href="/antibody/primary/query/cd121b">CD121b</a> (activated)<br><a href="/antibody/primary/query/cd134">CD134</a> (OX-40)<br><a href="/antibody/primary/query/cd137">CD137</a> (4-1BB)<br><a href="/antibody/primary/query/cd152">CD152</a> (CTLA-4)<br><a href="/antibody/primary/query/cd197">CD197</a> (CCR7)<br><a href="/antibody/primary/query/cd304">CD304</a> (NRP1)<br><a href="/antibody/primary/query/cd357">CD357</a> (GITR)<br><a href="/antibody/primary/query/FR4">FR4</a><br><a href="/antibody/primary/query/GARP">GARP</a> (activated)<br><a href="/antibody/primary/query/LAP">LAP</a> (activated)<br><a href="/antibody/primary/query/TIGIT">TIGIT</a></td><td><a href="/antibody/primary/query/CD45RA">CD45RA</a> (subset)<br><a href="/antibody/primary/query/cd49d">CD49b</a><br><a href="/antibody/primary/query/CD194">CD194</a> (CCR4; subset)<br><a href="/antibody/primary/query/CD196">CD196</a> (CCR6; subset)<br><a href="/antibody/primary/query/CD183">CD183</a> (CXCR3; subset)<br><a href="/antibody/primary/query/CD223">CD223</a> (LAG3)<br><a href="/antibody/primary/query/CD226">CD226</a><br><a href="/antibody/primary/query/CD278">CD278</a> (ICOS)<br><a href="/antibody/primary/query/CD279">CD279</a> (PD-1)<br><a href="/antibody/primary/query/GARP">GARP</a> (activated)<br><a href="/antibody/primary/query/LAP">LAP</a> (activated)</td><td><a href="/antibody/primary/query/CD152">CD152</a> (CTLA-4)<br><a href="/antibody/primary/query/GARP">GARP</a> (activated)<br><a href="/antibody/primary/query/LAP">LAP</a> (activated)</td></tr><tr><td><b><i>Ex vivo</i> expansion</b></td><td><a href="/antibody/product/CD3-Antibody-clone-OKT3-Monoclonal/16-0037-81">Anti-CD3</a> and <a href="/order/catalog/product/PHC0021#/PHC0021">IL-2</a></td><td>n/a</td><td>n/a</td></tr><tr><td><b><i>In vitro</i> differentiation</b></td><td>n/a</td><td>n/a</td><td><a href="/antibody/product/CD3-Antibody-clone-OKT3-Monoclonal/16-0037-81">Anti-CD3</a>, <a href="/antibody/product/CD28-Antibody-clone-CD28-2-Monoclonal/16-0289-81">anti-CD28</a>, <a href="/order/catalog/product/PHC0021#/PHC0021">IL-2</a>, and <a href="/order/catalog/product/PHG9214">TGFβ</a></td></tr><tr><td colspan="4"><sup>Low</sup>: low expression levels, <sup>Hi</sup>: high expression levels, <sup>+</sup>: expressed marker is present in cell subtype, <sup>-</sup>: expressed marker is not present in cell subtype.</td></tr></tbody></table></div></div><div class="heading section"><h2 id="disease"><br>Regulatory T cell in disease </h2></div><div class="text parbase section"><p>There are many different immunosuppressive mechanisms reportedly used by Treg. Cell contact–dependent suppression can be mediated by TIGIT, CD39, CD73, and CD152 (CTLA-4), as well as by cytolysis of target cells via granzymes A and B. While Treg do not secrete IL-2 upon activation, they can secrete immunomodulatory cytokines, such as IL-10, TGFβ, and IL-35. Depletion of IL-2 from the microenvironment and other metabolic perturbations have been reported as mechanisms used by Treg to suppress immune responses.</p><p>Several therapeutic drugs target Treg in order to enhance Treg function in autoimmune diseases and graft-versus-host disease (GVHD) or to block Treg function in some cancers. For example, anti-TNF antibodies were shown to expand the Treg population in patients with rheumatoid arthritis and stabilize their function, whereas low-dose IL-2 increased expression of CD25 on Foxp3<sup>+</sup> cells and expanded the Treg population with promising effects in graft-versus-host disease, Hepatitis C virus–induced vasculitis, type 1 diabetes, alopecia areata, and SLE. Additional studies have looked at using anti-CD3 antibodies, anti-CD25 antibodies, IL-2/anti-IL-2 complexes, anti–IL-6 receptor antibodies, and CTLA-4:Ig to target Treg in autoimmune disease. Methods to expand and stabilize Treg <i>ex vivo</i> for therapeutic use are being explored.</p></div><div class="heading section"><h2 id="isolation"><br>Isolation and sorting of viable Treg </h2></div><div class="text parbase section"><p>Treg were originally identified as a CD4<sup>+</sup> CD25<sup>+</sup> T cell population with the capacity to suppress an immune response. Magnetic cell separation approaches leverage the high expression of CD25 on Treg to enrich Foxp3<sup>+</sup> cells from both humans and mice. The identification of Foxp3 as the “master-regulator” of Treg was a critical step in defining Treg as a distinct T cell lineage. However, CD25 and Foxp3 expression may also be induced in T cells that lack suppressive function. Moreover, the localization of Foxp3 to the nucleus prevents its use as a marker for the isolation of viable Treg.</p><p>The identification of additional antigenic markers on the surface of Treg has enabled identification and flow cytometry–based sorting of viable Treg, resulting in a more highly enriched and suppressive Treg population. In addition to CD4 and CD25, both mouse and human Treg can express high levels of CD357 (GITR/AITR) and CD152 (CTLA-4) but express low levels of CD127 (IL-7Ra). In humans, the addition of markers such as CD45RO, CD39, and CD73 can improve the enrichment of Treg, and in fact several studies have shown that using CD4, CD25, and CD127 only to isolate Treg in humans can result in significant enrichment of activated conventional T cells. CD45RA has also proven to be a useful marker for the distinction of resting and effector Treg in humans. Similarly, in the mouse model, the addition of CD39, CD45RB, CD101, FR4, and CD73 to a panel of CD4, CD25, and CD127 can enhance enrichment of Treg.</p><p>Treg can exist in different states (resting or activated) and in specific tissues (gut, adipose, skin, or muscle), which can be further identified based on their expression of additional homing and migration markers. The identification and isolation of live Treg using a panel of antibodies for a variety of surface markers enables enrichment of Treg subpopulations and downstream analysis of their suppressive activity, as well as ex vivo expansion and transfer back into mice or humans for the regulation of immune responses.</p></div><div class="heading section"><h2 id="ex-vivo"><br>Treg <i>ex vivo</i>: expansion, differentiation, and function </h2></div><div class="text parbase section"><p>Treg are potent inhibitors of immune responses; however, they are not a very abundant population of cells. This lack of abundance has necessitated several approaches for <i>ex vivo</i> manipulation, including expansion of existing Treg or the differentiation of Treg from conventional CD4<sup>+</sup> T cells, in order to obtain sufficient numbers of cells to study their function, both <i>in vivo</i> and <i>in vitro</i>.</p><p>After isolating live Treg from humans or mice, either by magnetic cell separation or multiparameter flow cytometric approaches, this cell population may be expanded <i>in vitro</i> through stimulation of the TCR and addition of IL-2 with or without rapamycin. Stimulation of the TCR may be achieved using plate-bound anti-CD3 antibody, anti-CD3/anti-CD28 antibodies bound to beads, or anti-CD3 antibody–coated antigen presenting cells. Co-stimulation is routinely used for expansion of human Treg but may be dispensable for the <i>in vitro</i> expansion of mouse Treg. Adding rapamycin can be beneficial when using magnetic separation approaches, as it is known to inhibit the expansion of conventional T cells, thereby allowing Treg to preferentially expand and survive.</p><p>In mice, Treg may be generated <i>in vitro</i> from naive CD4<sup>+</sup> Foxp3<sup>–</sup> T cells when stimulated through the TCR using anti-CD3 antibody in the presence of IL-2 and TGFβ. Co-stimulation with anti-CD28 antibody may be dispensable, as its ability to promote Foxp3 expression is due primarily to the enhancement of endogenous IL-2 production. Interestingly, although naive CD4<sup>+</sup> T cells from humans can be induced to express Foxp3 under similar culture conditions, these cells do not have suppressive function and proliferate and produce inflammatory cytokines in response to re-stimulation. Some studies suggest that the addition of rapamycin or retinoic acid may help promote suppressive function in human cells, but the results have not been reproduced by other laboratories.</p><p>The suppressive function of Treg can be studied <i>in vitro</i> using a co-culture system of Treg and effector T cells. The source of Treg may be freshly isolated, expanded <i>ex vivo</i>, or iTreg. Typically, the Treg and effector T cells are cultured at varying ratios with TCR stimulation for up to 4 days. The most common readout is the proliferation of the effector T cells, either by dilution of CFSE (carboxyfluorescein succinimidyl ester), incorporation of BrdU (5-bromo-2'-deoxyuridine) or 3H-thymidine. Cytokine production, upregulation of activation markers, and cell counts may also be assessed. Interestingly, a two-day protocol using expression of CD25 and CD134 (OX-40) on effector T cells has been shown to correlate with proliferation and can serve as a shorter, surrogate measurement of suppressive function of Treg.</p></div><div class="text parbase section"><h4>Table 2. Reagents for <i>in vitro</i> expansion of tTreg.</h4><div class="table-component-wrapper"><table class="rte-table" id="table-1772143455"><tbody><tr><th style="width: 55%;">Mediators</th><th>Function</th></tr><tr><td><a href="/antibody/product/CD3-Antibody-clone-OKT3-Monoclonal/16-0037-81">Anti-CD3</a></td><td>TCR crosslinking</td></tr><tr><td><a href="/antibody/product/CD28-Antibody-clone-CD28-2-Monoclonal/16-0289-81">Anti-CD28</a></td><td>Co-stimulation</td></tr><tr><td><a href="/order/catalog/product/40203D#/40203D">Gibco CTS (Cell Therapy Systems) Dynabeads CD3/CD28</a></td><td>TCR crosslinking and co-stimulation</td></tr><tr><td><a href="/order/catalog/product/PHC0021#/PHC0021">IL-2</a></td><td>Growth factor</td></tr><tr><td><a href="/order/catalog/product/PHZ1235">Rapamycin</a></td><td>Suppress expansion of conventional T cells</td></tr></tbody></table></div></div><div class="text parbase section"><h4>Table 3. Reagents for <i>in vitro</i> generation of iTreg.</h4><div class="table-component-wrapper"><table class="rte-table" id="table-2128186451"><tbody><tr><th style="width: 55%;">Mediators</th><th>Function</th></tr><tr><td><a href="/antibody/product/CD3-Antibody-clone-OKT3-Monoclonal/16-0037-81">Anti-CD3</a></td><td>TCR crosslinking</td></tr><tr><td><a href="/antibody/product/CD28-Antibody-clone-CD28-2-Monoclonal/16-0289-81">Anti-CD28</a></td><td>Co-stimulation</td></tr><tr><td><a href="/order/catalog/product/PHC0021#/PHC0021">IL-2</a></td><td>Growth factor</td></tr><tr><td><a href="/order/catalog/product/PHG9214">TGFβ</a></td><td>Growth factor</td></tr><tr><td>Retinoic acid</td><td>Stabilization of Foxp3 and suppressive function?</td></tr><tr><td><a href="/order/catalog/product/PHZ1235">Rapamycin</a></td><td>Stabilization of Foxp3 and suppressive function?</td></tr></tbody></table></div></div><div class="text parbase section"><h4>Table 4. Reagents for studying the suppressive function of Treg <i>in vitro</i>.</h4><div class="table-component-wrapper"><table class="rte-table" id="table-604400381"><tbody><tr><th style="width: 55%;">Mediators</th><th>Function</th></tr><tr><td><a href="/antibody/product/CD3-Antibody-clone-OKT3-Monoclonal/16-0037-81">Anti-CD3</a></td><td>TCR crosslinking</td></tr><tr><td><a href="/antibody/product/CD28-Antibody-clone-CD28-2-Monoclonal/16-0289-81">Anti-CD28</a></td><td>Co-stimulation</td></tr><tr><td><a href="/order/catalog/product/40203D#/40203D">Gibco CTS (Cell Therapy Systems) Dynabeads CD3/CD28</a></td><td>TCR crosslinking and co-stimulation</td></tr><tr><td><a href="/order/catalog/product/PHC0021#/PHC0021">IL-2</a></td><td>Growth factor</td></tr><tr><td><a href="/sg/en/home/life-science/cell-analysis/flow-cytometry/flow-cytometry-assays-reagents/cell-proliferation-flow-cytometry.html">CFSE</a></td><td>Cell division</td></tr><tr><td><a href="/sg/en/home/life-science/cell-analysis/flow-cytometry/flow-cytometry-assays-reagents/cell-proliferation-flow-cytometry.html">CellTrace dyes</a></td><td>Cell division</td></tr><tr><td><a href="/antibody/primary/query/CD25">Anti-CD25</a></td><td>Activation marker</td></tr><tr><td><a href="/antibody/primary/query/CD134">Anti-CD134</a></td><td>Activation marker</td></tr><tr><td><a href="/sg/en/home/life-science/cell-analysis/flow-cytometry/flow-cytometry-assays-reagents/cell-proliferation-flow-cytometry.html">Brdu or EdU</a></td><td>DNA synthesis/proliferation</td></tr><tr><td><a href="/sg/en/home/life-science/cell-analysis/flow-cytometry/flow-cytometry-assays-reagents/cell-proliferation-flow-cytometry.html">3H-Thymidine</a></td><td>DNA synthesis/proliferation</td></tr></tbody></table></div></div><div class="heading section"><h2 id="CytokineP"><br>Cytokine profiling </h2></div><div class="text parbase section"><p>Treg cells have been shown to restrict T cell function through diverse methods including contact-dependent and cytokine-mediated mechanisms. These include the secretion of TGF-b which has been shown to be a potent regulator of effector T cell function, IL-10 which can function as a T cell inhibitory cytokine in a context-dependent manner, and IL-35 which some studies have shown to have an inhibitory effect on T cell proliferation. In addition, studies have demonstrated that the chemokines CCL3 and CCL4 can serve as chemoattractants that can affect cytotoxic T cell suppression by bringing them into close proximity to Tregs. Experiments have also suggested that Treg cells indirectly influence T cell proliferation and function due to surface expression of high-affinity IL-2 receptors when compared to stimulated naïve T cells, thus competing for IL-2.</p><h4>Table 5: Key cytokines involved in Treg differentiation and secretion.</h4><div class="table-component-wrapper"><table class="rte-table" id="table--1620845553"><tbody><tr><th> </th><th width="33%">Differentiation</th><th width="33%">Secreted</th></tr><tr><td>Cytokines and chemokines</td><td>IL-2, TGF-b</td><td>IL-10, TGF-b, IL-35, IL-9, CCL3, CCL4</td></tr></tbody></table></div><p>Tregs also express several immune checkpoint molecules including CTLA‐4, PD‐1, Tim‐3, LAG‐3, and TIGIT which deliver positive or negative immune modulatory signals on engagement with their cognate ligands expressed on antigen presenting cells. Several of these immune checkpoint molecules have been found in their soluble form, either formed due to the proteolytic cleavage of their membrane associated forms or expressed as soluble forms. These soluble forms could have functional activities that can activate or inhibit anti-tumor activity and can be measured in serum or plasma using multiplex immunoassays.</p></div><div class="text parbase section"><div class="table-component-wrapper"><table class="rte-table" id="table--852978349"><tbody><tr><th>Species</th><th width="26%">Description</th><th>Analytes</th><th width="20%">Catalog number</th></tr><tr><td rowspan="3">Human</td><td><a href="/order/catalog/product/EPX180-12165-901#/EPX180-12165-901">Th1/Th2/Th9/Th17 Cytokine 18-Plex Human ProcartaPlex Panel</a></td><td>GM-CSF, IFN gamma, IL-1 beta, IL-2, IL-4, IL-5, IL-6, IL-9, IL-10, IL-12p70, IL-13, IL-17A (CTLA-8), IL-18, IL-22, IL-23, IL-27, TNF alpha</td><td><a href="/order/catalog/product/EPX180-12165-901#/EPX180-12165-901">EPX180-12165-901</a></td></tr><tr><td><a href="/order/catalog/product/EPX14A-15803-901#/EPX14A-15803-901">Immuno-Oncology Checkpoint 14-Plex Human ProcartaPlex Panel 1</a></td><td>D27, CD28, CD137 (4-1BB), GITR, HVEM, BTLA, CD80, CD152 (CTLA4), IDO, LAG-3, PD-1, PD-L1, PD-L2, TIM-3</td><td><a href="/order/catalog/product/EPX14A-15803-901#/EPX14A-15803-901">EPX14A-15803-901</a></td></tr><tr><td><a href="/order/catalog/product/EPX140-15815-901#/EPX140-15815-901">Immuno-Oncology Checkpoint 14-Plex Human ProcartaPlex Panel 2</a></td><td>MICA, MICB, Perforin, ULBP-1, ULBP-3, ULBP-4, Arginase-1, CD73 (NT5E), CD96 (Tactile), E-Cadherin, Nectin-2, PVR, Siglec-7, Siglec-9</td><td><a href="/order/catalog/product/EPX140-15815-901#/EPX140-15815-901">EPX140-15815-901</a></td></tr><tr><td rowspan="3">Mouse</td><td><a href="/order/catalog/product/EPX170-26087-901#/EPX170-26087-901">Th1/Th2/Th9/Th17/Th22/Treg Cytokine 17-Plex Mouse ProcartaPlex Panel</a></td><td>GM-CSF, IFN gamma, IL-1 beta, IL-2, IL-4, IL-5, IL-6, IL-9, IL-10, IL-12p70, IL-13, IL-17A (CTLA-8), IL-18, IL-22, IL-23, IL-27, TNF alpha</td><td><a href="/order/catalog/product/EPX170-26087-901#/EPX170-26087-901">EPX170-26087-901</a></td></tr><tr><td><a href="/order/catalog/product/EPX070-20835-901#/EPX070-20835-901">Immuno-Oncology Checkpoint 7-Plex Mouse ProcartaPlex Panel 2</a></td><td>CD137L (4-1BBL), CD152 (CTLA4), CD276 (B7-H3), CD80, PD-1, PD-L1, PD-L2</td><td><a href="/order/catalog/product/EPX070-20835-901#/EPX070-20835-901">EPX070-20835-901</a></td></tr><tr><td><a href="/order/catalog/product/EPX040-20830-901#/EPX040-20830-901">Immuno-Oncology Checkpoint 4-Plex Mouse ProcartaPlex Panel 1</a></td><td>BTLA, CD27, LAG-3, TIM-3</td><td><a href="/order/catalog/product/EPX040-20830-901#/EPX040-20830-901">EPX040-20830-901</a></td></tr></tbody></table></div><p><a name="references" id="references"></a></p></div><div class="accordion section"><div class="accordion" id="49787e8de0ef490d874a0ea6b1bf556f" style='margin-bottom: 0;'><div class="parsys"><div class="accordionitem section"><div class='accordion-group' ><div class='accordion-heading '><a class="accordion-toggle" data-toggle="collapse" data-parent="#49787e8de0ef490d874a0ea6b1bf556f" href="#referencesaccordion"><span class="caret"></span> References </a></div><div class="accordion-body collapse " id="referencesaccordion"><div class="accordion-inner"><div class="accordion-content"><div class="itemParsys parsys"><div class="text parbase section"><ol><li><a href="https://www.ncbi.nlm.nih.gov/pubmed/32305296" target="_blank">Zhang X, Olsen N, Zheng SG (2020) The progress and prospect of regulatory T cells in autoimmune diseases. 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Curr Protoc Immunol Chapter 7:Unit 7.4B.</a><span class="icon icon-outbound"></span></li></ol></div></div></div></div></div></div></div></div></div><script> $j(document).ready(function(){ $j('#49787e8de0ef490d874a0ea6b1bf556f').on('hide', function(e) { $(e.target).find(".accordion-content").hide(); }); $j('#49787e8de0ef490d874a0ea6b1bf556f').on('show', function(e) { $(e.target).find(".accordion-content").show(); }); }); </script></div><div class="well-cmp section"><div class="well "><div class="well-container" style=""><div class="well-content"><div class="parsys"><div class="heading section"><h2 id="resources"> Related articles and resources </h2></div><div comp-order="bycol" class="parsys_column cq-colctrl-lt0"><div class="parsys_column cq-colctrl-lt0-c0"><div container-id="cq-colctrl-lt0" comp-id="A1"><div class="well-cmp section"><div class="well well-gray-lighter well-mini"><div class="well-container" style=""><div class="well-content"><div class="parsys"><div comp-order="bycol" class="parsys_column cq-colctrl-lt8"><div class="parsys_column cq-colctrl-lt8-c0"><div container-id="cq-colctrl-lt8" comp-id="A1"><div class="image parbase section"><div id="cq-image-jsp-cb55b0fa-3021-4418-b36c-884782fec6a0MainParsyswell_9410uiparwell_ed63uiparimage_9f7f"><a href="http://assets.thermofisher.com/TFS-Assets/BID/posters/t-helper-cell-paradigm-poster.pdf" target='_blank'><picture><source srcset='https://www.thermofisher.com/sg/en/home/life-science/cell-analysis/cell-analysis-learning-center/immunology-at-work/regulatory-t-cell-overview/jcr:content/MainParsys/well_9410/uipar/well_ed63/uipar/image_9f7f/foregroundimg.img.full.high.jpg/1680611313562.jpg' media="(min-width: 1px)"><img srcset="https://www.thermofisher.com/sg/en/home/life-science/cell-analysis/cell-analysis-learning-center/immunology-at-work/regulatory-t-cell-overview/jcr:content/MainParsys/well_9410/uipar/well_ed63/uipar/image_9f7f/foregroundimg.img.full.high.jpg/1680611313562.jpg,https://www.thermofisher.com/sg/en/home/life-science/cell-analysis/cell-analysis-learning-center/immunology-at-work/regulatory-t-cell-overview/jcr:content/MainParsys/well_9410/uipar/well_ed63/uipar/image_9f7f/foregroundimg.img.320.high.jpg/1680611313562.jpg,https://www.thermofisher.com/sg/en/home/life-science/cell-analysis/cell-analysis-learning-center/immunology-at-work/regulatory-t-cell-overview/jcr:content/MainParsys/well_9410/uipar/well_ed63/uipar/image_9f7f/foregroundimg.img.480.high.jpg/1680611313562.jpg,https://www.thermofisher.com/sg/en/home/life-science/cell-analysis/cell-analysis-learning-center/immunology-at-work/regulatory-t-cell-overview/jcr:content/MainParsys/well_9410/uipar/well_ed63/uipar/image_9f7f/foregroundimg.img.320.medium.jpg/1680611313562.jpg,https://www.thermofisher.com/sg/en/home/life-science/cell-analysis/cell-analysis-learning-center/immunology-at-work/regulatory-t-cell-overview/jcr:content/MainParsys/well_9410/uipar/well_ed63/uipar/image_9f7f/foregroundimg.img.620.high.jpg/1680611313562.jpg,https://www.thermofisher.com/sg/en/home/life-science/cell-analysis/cell-analysis-learning-center/immunology-at-work/regulatory-t-cell-overview/jcr:content/MainParsys/well_9410/uipar/well_ed63/uipar/image_9f7f/foregroundimg.img.320.low.jpg/1680611313562.jpg,https://www.thermofisher.com/sg/en/home/life-science/cell-analysis/cell-analysis-learning-center/immunology-at-work/regulatory-t-cell-overview/jcr:content/MainParsys/well_9410/uipar/well_ed63/uipar/image_9f7f/foregroundimg.img.480.medium.jpg/1680611313562.jpg" alt='T-helper cell paradigm poster cover' title='T-helper cell paradigm poster'></picture><style> #cq-image-jsp-cb55b0fa-3021-4418-b36c-884782fec6a0MainParsyswell_9410uiparwell_ed63uiparimage_9f7f img{ width:100%; 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