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Micro-scale flow cytometry-based and biochemical analysis of lipid signaling in primary B cell subpopulations

B cell subpopulations in the spleen have been extensively characterized phenotypically; however, biochemical properties of these cell populations following B cell antigen receptor engagement have not been fully determined due to technical difficulties and limiting cell numbers. We therefore employed...

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Detalles Bibliográficos
Autores principales: Antony, Pierre, Hoek, Kristen, Sarmah, Bhaskarjyoti, Khan, Wasif N.
Formato: Texto
Lenguaje:English
Publicado: Biological Procedures Online 2007
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2275047/
https://www.ncbi.nlm.nih.gov/pubmed/18385809
http://dx.doi.org/10.1251/bpo135
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author Antony, Pierre
Hoek, Kristen
Sarmah, Bhaskarjyoti
Khan, Wasif N.
author_facet Antony, Pierre
Hoek, Kristen
Sarmah, Bhaskarjyoti
Khan, Wasif N.
author_sort Antony, Pierre
collection PubMed
description B cell subpopulations in the spleen have been extensively characterized phenotypically; however, biochemical properties of these cell populations following B cell antigen receptor engagement have not been fully determined due to technical difficulties and limiting cell numbers. We therefore employed mini-scale protocols to assess lipid signaling, particularly that of diacylglycerol and inositol trisphosphate, with as few as 0.5x10(6) purified early (T1) and late (T2) transitional B cells. Additionally, utilizing flow cytometric techniques, we determined levels of phosphatidylinositol bisphosphate and calcium mobilization in T1 and T2 cells, as well as mature follicular and marginal zone B cells using less than 1x10(6 )primary B cells. Thus, these biochemical and flow cytometric methodologies can be used to analyse signal-induced changes in phosphatidylinositol bisphosphate levels, diacylglycerol and inositol triphosphate production and calcium in each B cell population.
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spelling pubmed-22750472008-04-02 Micro-scale flow cytometry-based and biochemical analysis of lipid signaling in primary B cell subpopulations Antony, Pierre Hoek, Kristen Sarmah, Bhaskarjyoti Khan, Wasif N. Biol Proced Online Article B cell subpopulations in the spleen have been extensively characterized phenotypically; however, biochemical properties of these cell populations following B cell antigen receptor engagement have not been fully determined due to technical difficulties and limiting cell numbers. We therefore employed mini-scale protocols to assess lipid signaling, particularly that of diacylglycerol and inositol trisphosphate, with as few as 0.5x10(6) purified early (T1) and late (T2) transitional B cells. Additionally, utilizing flow cytometric techniques, we determined levels of phosphatidylinositol bisphosphate and calcium mobilization in T1 and T2 cells, as well as mature follicular and marginal zone B cells using less than 1x10(6 )primary B cells. Thus, these biochemical and flow cytometric methodologies can be used to analyse signal-induced changes in phosphatidylinositol bisphosphate levels, diacylglycerol and inositol triphosphate production and calcium in each B cell population. Biological Procedures Online 2007-12-24 /pmc/articles/PMC2275047/ /pubmed/18385809 http://dx.doi.org/10.1251/bpo135 Text en Article © by the author(s). This paper is Open Access and is published in Biological Procedures Online under license from the author(s). Copying, printing, redistribution and storage permitted. Journal © 1997-2007 Biological Procedures Online.
spellingShingle Article
Antony, Pierre
Hoek, Kristen
Sarmah, Bhaskarjyoti
Khan, Wasif N.
Micro-scale flow cytometry-based and biochemical analysis of lipid signaling in primary B cell subpopulations
title Micro-scale flow cytometry-based and biochemical analysis of lipid signaling in primary B cell subpopulations
title_full Micro-scale flow cytometry-based and biochemical analysis of lipid signaling in primary B cell subpopulations
title_fullStr Micro-scale flow cytometry-based and biochemical analysis of lipid signaling in primary B cell subpopulations
title_full_unstemmed Micro-scale flow cytometry-based and biochemical analysis of lipid signaling in primary B cell subpopulations
title_short Micro-scale flow cytometry-based and biochemical analysis of lipid signaling in primary B cell subpopulations
title_sort micro-scale flow cytometry-based and biochemical analysis of lipid signaling in primary b cell subpopulations
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2275047/
https://www.ncbi.nlm.nih.gov/pubmed/18385809
http://dx.doi.org/10.1251/bpo135
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