ANALYSIS OF T-CELL RECEPTOR-BETA CHAIN EXPRESSION BY ISOELECTRIC-FOCUSING FOLLOWING GENE AMPLIFICATION AND IN-VITRO TRANSLATION

We describe a new approach to analysis of T cell receptor diversity ba sed on isoelectric focusing of in vitro translation products of amplif ied V region genes. The method is illustrated by analysis of V beta 2 profiles in peripheral blood lymphocytes from normal donors. The prime rs used for V beta 2 analysis spanned the V-(D-)J junction and include d the segment from amino acid residue position 53 in the variable regi on to residue 132 of the constant region. The isoelectric focusing pat terns display approximately 13-14 bands of varying intensity. Differen ces in expression of V beta 2-derived peptides were detected in compar isons of the isoelectric focusing profiles from different individuals, suggesting that the method may be useful for detecting genetically de termined, immune response related or disease associated differences in Tcr V region expression. The major isoelectric focusing bands have be en interpreted as representing groups of V beta 2 sequences sharing J beta region and NDN region charge similarity. Quantitative differences were detected in V beta 2 profiles of CD4 and CD8 T cell subpopulatio ns indicating there may be selection for different charge characterist ics in NDNJ sequences in the two T cell subsets. The method provides a new dimension for the detection of perturbations in the T cell repert oire.