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J. Cell Biol.,
Volume 140, Number 1, January 12, 1998 71-79
Department of Biological Sciences, University of Essex, Wivenhoe Park, Colchester CO4 3SQ, United Kingdom
The technique of single-particle fluorescence
imaging was used to investigate the oligomeric state of
MHC class II molecules on the surface of living cells.
Cells transfected with human leukocyte antigen
(HLA)-DR A and B genes were labeled at saturation
with a univalent probe consisting of Fab coupled to
R-phycoerythrin. Analysis of the intensities of fluorescent spots on the cell surface revealed the presence of
single and double particles consistent with the simultaneous presence of HLA-DR heterodimers and dimers of dimers. The proportion of double particles was lower
at 37°C than at 22°C, suggesting that the heterodimers
and dimers of dimers exist in a temperature-dependent
equilibrium. These results are discussed in the context
of a possible role for HLA-DR dimers of dimers in T
cell receptor-MHC interactions. The technique is validated by demonstrating that fluorescence imaging can
distinguish between dimers and tetramers of human
erythrocyte spectrin deposited from solution onto a
solid substrate. The methodology will have broad applicability to investigation of the oligomeric state of immunological and other membrane-bound receptors in
living cells.
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