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Techniques have been developed to measure the reactivity of free thiols in the HLA class I antigen-binding cleft. HLA-B27, which sequencing predicts has a free cysteine at position 67, reacts rapidly with the positively charged thiol reagent monobromotrimethyl-ammoniobimane bromide (qBBr) to give products which are identifiable by isoelectric focusing. HLA-B38, B39, B64 and B65, all of which have a similar Cys 67, react less strongly. Several other class I molecules, notably HLA-C antigens, are reactive in this system, and it may be capable of recognizing subtypes such as A*0207 which also carry free cysteine. The accessibility of thiol to qBBr depends both on the chemistry of the class I molecule and other factors in the cell. Two human cell lines which are known to carry identical B27 genes but do not present the same peptides, differ considerably in the accessibility of their B27 thiol. Evidence from mouse cells transfected with mutant B27 genes suggests that a unique lysine at position 70 in the wild-type molecule increases reactivity to thiol-reactive metabolites. The failure of B27 to give a complete reaction with qBBr in our model systems suggests that it can exist in more than one chemical form. This may leave the molecule susceptible to oxidation, causing errors in T cell recognition and an exaggerated inflammatory response.

SUMMARY A mouse alloantiserum, raised by immunizing B10.D2/n mice with B10.M spleen, has been found to retain cytotoxic activity against a proportion of lymph node cells even after exhaustive absorption with B10.M red blood cells. Titration and absorption with a number of strains showed that the absorbed serum retained antibodies against two new specificities. The absorbed serum gave significant enhancement of B10.M skin grafts on B10.D2 and reacted mainly with B cells.

Immune responses to conserved, immunogenic homologues of the mycobacterial 65 kDa stress protein (SP65) have been implicated in inflammatory arthritis. Serum anti-SP65 was measured in AS, RA and healthy controls using an indirect enzyme immunoassay with recombinant SP65. IgA anti-SP65 was elevated in 19 of 59 AS patients, but the elevation in median level was not statistically significant. Anti-SP65 of all isotypes was increased in RA, but achieved significance (P less than 0.01) for IgA only. Adjusting specific antibody results for elevations in total serum Ig levels reduced AS and RA anti-SP65 to near normal levels, suggesting that a major component of the increased anti-SP65 may be secondary to polyclonal activation.

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