Ki-67 staining for determination of rhesus macaque T cell proliferative responses ex vivo
(c) 2010 International Society for Advancement of Cytometry..
The capacity for robust proliferation upon re-infection is a hallmark of adaptive immunity and the basis of vaccination. A widely used animal model for the study of human disease is the rhesus macaque (RM), where capacity for proliferation can be assessed ex vivo using carboxyfluorescein succinimidyl ester (CFSE)-based dilution assays. However, we show over the course of the standard ex vivo proliferation assay that CFSE-labeling at commonly used dye concentrations induces significant cell death, but that this phenomenon is dose-dependent. Here, we describe an alternative semiquantitative method for estimating T cell proliferative responses that avoids the putative biases associated with chemical modification. RM peripheral blood mononuclear cells were stimulated ex vivo with cognate peptides for 5 days, immunostained for intracellular Ki-67, and then analyzed by flow cytometry. We describe a gating strategy using Ki-67 and side light scatter, also a marker of blastogenesis, which correlates strongly with data from CFSE dilution. We show that this method is a valid tool for measuring RM antigen-specific cellular proliferation ex vivo and can be used as an alternative to CFSE dilution assays.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2010 |
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| Erschienen: |
2010 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:77 |
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| Enthalten in: |
Cytometry. Part A : the journal of the International Society for Analytical Cytology - 77(2010), 3 vom: 01. März, Seite 275-84 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Shedlock, Devon J [VerfasserIn] |
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| Links: |
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| Anmerkungen: |
Date Completed 04.06.2010 Date Revised 20.10.2021 published: Print Citation Status MEDLINE |
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| doi: |
10.1002/cyto.a.20857 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM194641988 |
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| 245 | 1 | 0 | |a Ki-67 staining for determination of rhesus macaque T cell proliferative responses ex vivo |
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| 500 | |a Date Revised 20.10.2021 | ||
| 500 | |a published: Print | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a (c) 2010 International Society for Advancement of Cytometry. | ||
| 520 | |a The capacity for robust proliferation upon re-infection is a hallmark of adaptive immunity and the basis of vaccination. A widely used animal model for the study of human disease is the rhesus macaque (RM), where capacity for proliferation can be assessed ex vivo using carboxyfluorescein succinimidyl ester (CFSE)-based dilution assays. However, we show over the course of the standard ex vivo proliferation assay that CFSE-labeling at commonly used dye concentrations induces significant cell death, but that this phenomenon is dose-dependent. Here, we describe an alternative semiquantitative method for estimating T cell proliferative responses that avoids the putative biases associated with chemical modification. RM peripheral blood mononuclear cells were stimulated ex vivo with cognate peptides for 5 days, immunostained for intracellular Ki-67, and then analyzed by flow cytometry. We describe a gating strategy using Ki-67 and side light scatter, also a marker of blastogenesis, which correlates strongly with data from CFSE dilution. We show that this method is a valid tool for measuring RM antigen-specific cellular proliferation ex vivo and can be used as an alternative to CFSE dilution assays | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, N.I.H., Extramural | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 7 | |a 5-(6)-carboxyfluorescein diacetate succinimidyl ester |2 NLM | |
| 650 | 7 | |a Antigens |2 NLM | |
| 650 | 7 | |a Coloring Agents |2 NLM | |
| 650 | 7 | |a Fluoresceins |2 NLM | |
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| 700 | 1 | |a Talbott, Kendra T |e verfasserin |4 aut | |
| 700 | 1 | |a Morrow, Matthew P |e verfasserin |4 aut | |
| 700 | 1 | |a Ferraro, Bernadette |e verfasserin |4 aut | |
| 700 | 1 | |a Hokey, David A |e verfasserin |4 aut | |
| 700 | 1 | |a Muthumani, Karuppiah |e verfasserin |4 aut | |
| 700 | 1 | |a Weiner, David B |e verfasserin |4 aut | |
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