Rapid, efficient functional characterization and recovery of HIV-specific human CD8+ T cells using microengraving
The nature of certain clinical samples (tissue biopsies, fluids) or the subjects themselves (pediatric subjects, neonates) often constrain the number of cells available to evaluate the breadth of functional T-cell responses to infections or therapeutic interventions. The methods most commonly used t...
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| Format: | Article |
| Language: | en_US |
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National Academy of Sciences
2012
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| Online Access: | http://hdl.handle.net/1721.1/74605 https://orcid.org/0000-0003-0921-3144 |
| _version_ | 1826191660327895040 |
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| author | Varadarajan, Navin Kwon, Douglas Law, Kenneth M. Ogunniyi, Adebola Oluwakayode Anahtar, Melis N. Richter, James M. Walker, Bruce D. Love, John C |
| author2 | Massachusetts Institute of Technology. Department of Chemical Engineering |
| author_facet | Massachusetts Institute of Technology. Department of Chemical Engineering Varadarajan, Navin Kwon, Douglas Law, Kenneth M. Ogunniyi, Adebola Oluwakayode Anahtar, Melis N. Richter, James M. Walker, Bruce D. Love, John C |
| author_sort | Varadarajan, Navin |
| collection | MIT |
| description | The nature of certain clinical samples (tissue biopsies, fluids) or the subjects themselves (pediatric subjects, neonates) often constrain the number of cells available to evaluate the breadth of functional T-cell responses to infections or therapeutic interventions. The methods most commonly used to assess this functional diversity ex vivo and to recover specific cells to expand in vitro usually require more than 10[superscript 6] cells. Here we present a process to identify antigen-specific responses efficiently ex vivo from 10[superscript 4]–10[superscript 5] single cells from blood or mucosal tissues using dense arrays of subnanoliter wells. The approach combines on-chip imaging cytometry with a technique for capturing secreted proteins—called “microengraving”—to enumerate antigen-specific responses by single T cells in a manner comparable to conventional assays such as ELISpot and intracellular cytokine staining. Unlike those assays, however, the individual cells identified can be recovered readily by micromanipulation for further characterization in vitro. Applying this method to assess HIV-specific T-cell responses demonstrates that it is possible to establish clonal CD8[superscript +] T-cell lines that represent the most abundant specificities present in circulation using 100- to 1,000-fold fewer cells than traditional approaches require and without extensive genotypic analysis a priori. This rapid (<24 h), efficient, and inexpensive process should improve the comparative study of human T-cell immunology across ages and anatomic compartments. |
| first_indexed | 2024-09-23T08:59:23Z |
| format | Article |
| id | mit-1721.1/74605 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T08:59:23Z |
| publishDate | 2012 |
| publisher | National Academy of Sciences |
| record_format | dspace |
| spelling | mit-1721.1/746052022-09-30T12:38:23Z Rapid, efficient functional characterization and recovery of HIV-specific human CD8+ T cells using microengraving Varadarajan, Navin Kwon, Douglas Law, Kenneth M. Ogunniyi, Adebola Oluwakayode Anahtar, Melis N. Richter, James M. Walker, Bruce D. Love, John C Massachusetts Institute of Technology. Department of Chemical Engineering Koch Institute for Integrative Cancer Research at MIT Varadarajan, Navin Ogunniyi, Adebola Oluwakayode Love, J. Christopher The nature of certain clinical samples (tissue biopsies, fluids) or the subjects themselves (pediatric subjects, neonates) often constrain the number of cells available to evaluate the breadth of functional T-cell responses to infections or therapeutic interventions. The methods most commonly used to assess this functional diversity ex vivo and to recover specific cells to expand in vitro usually require more than 10[superscript 6] cells. Here we present a process to identify antigen-specific responses efficiently ex vivo from 10[superscript 4]–10[superscript 5] single cells from blood or mucosal tissues using dense arrays of subnanoliter wells. The approach combines on-chip imaging cytometry with a technique for capturing secreted proteins—called “microengraving”—to enumerate antigen-specific responses by single T cells in a manner comparable to conventional assays such as ELISpot and intracellular cytokine staining. Unlike those assays, however, the individual cells identified can be recovered readily by micromanipulation for further characterization in vitro. Applying this method to assess HIV-specific T-cell responses demonstrates that it is possible to establish clonal CD8[superscript +] T-cell lines that represent the most abundant specificities present in circulation using 100- to 1,000-fold fewer cells than traditional approaches require and without extensive genotypic analysis a priori. This rapid (<24 h), efficient, and inexpensive process should improve the comparative study of human T-cell immunology across ages and anatomic compartments. National Institutes of Health (U.S.) (Grant U54-AI057156) National Institute of Allergy and Infectious Diseases (U.S.) 2012-11-08T18:29:23Z 2012-11-08T18:29:23Z 2012-03 2011-07 Article http://purl.org/eprint/type/JournalArticle 0027-8424 1091-6490 http://hdl.handle.net/1721.1/74605 Varadarajan, N. et al. “Rapid, Efficient Functional Characterization and Recovery of HIV-specific Human CD8+ T Cells Using Microengraving.” Proceedings of the National Academy of Sciences 109.10 (2012): 3885–3890. ©2012 by the National Academy of Sciences https://orcid.org/0000-0003-0921-3144 en_US http://dx.doi.org/10.1073/pnas.1111205109 Proceedings of the National Academy of Sciences Article is made available in accordance with the publisher's policy and may be subject to US copyright law. Please refer to the publisher's site for terms of use. application/pdf National Academy of Sciences PNAS |
| spellingShingle | Varadarajan, Navin Kwon, Douglas Law, Kenneth M. Ogunniyi, Adebola Oluwakayode Anahtar, Melis N. Richter, James M. Walker, Bruce D. Love, John C Rapid, efficient functional characterization and recovery of HIV-specific human CD8+ T cells using microengraving |
| title | Rapid, efficient functional characterization and recovery of HIV-specific human CD8+ T cells using microengraving |
| title_full | Rapid, efficient functional characterization and recovery of HIV-specific human CD8+ T cells using microengraving |
| title_fullStr | Rapid, efficient functional characterization and recovery of HIV-specific human CD8+ T cells using microengraving |
| title_full_unstemmed | Rapid, efficient functional characterization and recovery of HIV-specific human CD8+ T cells using microengraving |
| title_short | Rapid, efficient functional characterization and recovery of HIV-specific human CD8+ T cells using microengraving |
| title_sort | rapid efficient functional characterization and recovery of hiv specific human cd8 t cells using microengraving |
| url | http://hdl.handle.net/1721.1/74605 https://orcid.org/0000-0003-0921-3144 |
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