An Optimized and Validated Method for Isolation and Characterization of Lymphocytes from HIV+ Human Gut Biopsies
The gastrointestinal (GI) tract harbors most of the body's immune cells and is also a major HIV reservoir in ART-treated patients. To achieve a cure, most HIV-infected cells must be identified and eliminated. While obtaining gut biopsies is a relatively noninvasive method of sampling relevant t...
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| Veröffentlicht in: | AIDS research and human retroviruses 2017-11, Vol.33 (S1), p.S31-S-39 |
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| creator | Trapecar, Martin Khan, Shahzada Roan, Nadia R Chen, Tsui-Hua Telwatte, Sushama Deswal, Monika Pao, Montha Somsouk, Ma Deeks, Steven G Hunt, Peter W Yukl, Steven Sanjabi, Shomyseh |
| description | The gastrointestinal (GI) tract harbors most of the body's immune cells and is also a major HIV reservoir in ART-treated patients. To achieve a cure, most HIV-infected cells must be identified and eliminated. While obtaining gut biopsies is a relatively noninvasive method of sampling relevant tissue for monitoring HIV activity, immune cell isolation from these limited tissue samples has proven to be challenging. Enzymatic tissue digestion is required for maximal immune cell isolation from gut biopsies. However, these enzymatic digestions can also be detrimental for preservation of cellular surface markers that are required for accurate identification of various subsets of leukocytes. In this study, we describe an optimized protocol for isolation of lymphocytes from human gut biopsies. We also discuss our validation results, which show that compared with several other collagenase preparations, the use of CSLPA maintains high lymphocyte recovery while preserving the integrity of most cellular surface antigens that we tested. Importantly, chemokine receptors that are used to characterize various subsets of T cells, which are notorious for being digested during a typical enzymatic tissue digestion, are highly preserved using this protocol. |
| doi_str_mv | 10.1089/AID.2017.0208 |
| format | Article |
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To achieve a cure, most HIV-infected cells must be identified and eliminated. While obtaining gut biopsies is a relatively noninvasive method of sampling relevant tissue for monitoring HIV activity, immune cell isolation from these limited tissue samples has proven to be challenging. Enzymatic tissue digestion is required for maximal immune cell isolation from gut biopsies. However, these enzymatic digestions can also be detrimental for preservation of cellular surface markers that are required for accurate identification of various subsets of leukocytes. In this study, we describe an optimized protocol for isolation of lymphocytes from human gut biopsies. We also discuss our validation results, which show that compared with several other collagenase preparations, the use of CSLPA maintains high lymphocyte recovery while preserving the integrity of most cellular surface antigens that we tested. 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To achieve a cure, most HIV-infected cells must be identified and eliminated. While obtaining gut biopsies is a relatively noninvasive method of sampling relevant tissue for monitoring HIV activity, immune cell isolation from these limited tissue samples has proven to be challenging. Enzymatic tissue digestion is required for maximal immune cell isolation from gut biopsies. However, these enzymatic digestions can also be detrimental for preservation of cellular surface markers that are required for accurate identification of various subsets of leukocytes. In this study, we describe an optimized protocol for isolation of lymphocytes from human gut biopsies. We also discuss our validation results, which show that compared with several other collagenase preparations, the use of CSLPA maintains high lymphocyte recovery while preserving the integrity of most cellular surface antigens that we tested. Importantly, chemokine receptors that are used to characterize various subsets of T cells, which are notorious for being digested during a typical enzymatic tissue digestion, are highly preserved using this protocol.</abstract><cop>United States</cop><pub>Mary Ann Liebert, Inc</pub><pmid>28882052</pmid><doi>10.1089/AID.2017.0208</doi><oa>free_for_read</oa></addata></record> |
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| ispartof | AIDS research and human retroviruses, 2017-11, Vol.33 (S1), p.S31-S-39 |
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| source | MEDLINE; Alma/SFX Local Collection |
| subjects | AIDS/HIV Antigens Biopsy Biopsy - methods CD4 Lymphocyte Count CD4-Positive T-Lymphocytes - immunology Cell Separation - methods Chemokine receptors Chemokines - analysis Collagen Collagenase Digestion Digestive system Gastrointestinal tract Gastrointestinal Tract - immunology Gastrointestinal Tract - virology HIV HIV Infections - immunology HIV-1 - immunology Human immunodeficiency virus Humans Immune system Immunology Leukocytes Lymphocytes Lymphocytes T Preservation Receptors Surface antigens Surface markers T cell receptors |
| title | An Optimized and Validated Method for Isolation and Characterization of Lymphocytes from HIV+ Human Gut Biopsies |
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