PMID: 8450239Mar 15, 1993Paper

A sensitive method to detect defined peptide among those eluted from murine MHC class II molecules

Journal of Immunological Methods
T D BrumeanuH Zaghouani

Abstract

We developed a sensitive competitive inhibition radioimmunoassay able to trace pmoles of a defined peptide eluted from major histocompatibility complex (MHC) class II molecules that were subsequently fractionated by RP-HPLC. In this assay we used a model synthetic peptide corresponding to amino acid residues 110-120 from the hemagglutinin (HA) of PR8 influenza virus, and affinity purified rabbit antibodies specific for this peptide. The HA110-120 peptide binds to I-Ed class II molecules on the surface of APCs and is recognized by specific CD4+ T helper cells. 2PK3 B lymphoma cells (H-2d) were pulsed with HA110-120 peptide or PR8 virus, lysed, the MHC class II molecules extracted, and bound peptides eluted. After separation by RP-HPLC, the fractions were tested for inhibition of the binding of rabbit anti-HA110-120 antibodies to peptide coated microtiter plates. A significant inhibitory activity was observed with one peak when the cells were pulsed with HA110-120 peptide and two peaks when pulsed with PR8 virus. The inhibitory activity was correlated with the presence of HA110-120 peptide as demonstrated by peptide sequencing. The assay is reproducible and sensitive to 1 pmol of antigenic peptide. This assay can be useful to ide...Continue Reading

References

Aug 15, 1992·Proceedings of the National Academy of Sciences of the United States of America·C A NelsonE R Unanue
Aug 1, 1987·Immunological Reviews·P M AllenE R Unanue
Aug 1, 1987·Immunological Reviews·T J BracialeV L Braciale
Jan 1, 1989·Annual Review of Immunology·A Townsend, H Bodmer

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Citations

Jun 28, 1995·Journal of Immunological Methods·T D BrumeanuC A Bona
Jun 1, 1996·Immunotechnology : an International Journal of Immunological Engineering·T D BrumeanuH Zaghouani

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