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KMID : 0357319960310050599
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Journal of the Korean Society for Microbiology
1996 Volume.31 No. 5 p.599 ~ p.605
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Analysis of the Assembly of CD3 to T Cell Antigen Receptor by Flow Cytometry
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Abstract
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CD3ζ, component of T cell receptor(TCR) complex, is believed to transduce a signal from the T cell receptor into the interior of the cell thus contributing to T cell activation.
Although flow cytometry is a good tool for analysis of the surface molecules expressed on T lymphocyte, it is difficult to analyze CD3ζassociation to TCR by flow cytometry because most of CD3ζresides in intracelluar region rather than
extracellular
surface.
To determine the detectability of CD3ζassociation to TCR by flow cytometry using the partial permeabilization method, we partially permealized T cell lines(CTLL-2: TCR associated with CD3ζTIB-48 not associated with CD3ζ) prior to staining with
rabbit
anti-CD3ζintracellular peptide antibody(N-38) and FITC conjugated anti-rabbit IgG.
We found that the partial permeabilization with lysolecithin(10μg/ml) for 1 min prior to staining with the antibodies enables to detect the intracellular portion of CD3ζassociated with TCR and the results of flow cytometry were comparable with
those
of immunoprecipitation and SDS-PAGE with the same cell lines.
Our result shows that the partial permeabilization of T lymphocyte prior to flow cytometry using antibody specific to intracellular peptide of CD3ζassociated with TCR.
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KEYWORD
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