RESUMO
The two mouse MHC class I alleles, L(d) and L(q), share complete amino acid sequence identity except in the alpha2 domain, where they differ at six positions. Despite their similarity, L(q) has a stronger association with beta2-microglobulin (beta2m), is expressed at higher levels on the cell surface, demonstrates an increased cell surface half-life, and has fewer open forms on the cell surface than L(d). To determine the basis for their phenotypic differences, L(d) molecules containing chimeric L(d)-L(q) alpha2 domains were characterized, and these analyses implicated residue 97 (L(d)Trp and L(q)Arg) as the polymorphic site responsible for the disparity in beta2m association between the two alleles. Single substitution analysis at this site (L(d)W97R and L(q)R97W) confirmed this. Furthermore, the L(d)W97R mutant molecule has a longer cell surface half-life than either L(q) or L(d), and fewer open forms of L(d)W97R are observed on the cell surface. In addition, both L(d)W97R and L(q) possess decreased binding affinity for the L(d)-restricted tum(-) P91A(14-22) peptide compared with L(d). Collectively, these results and the known location of Trp(97) in the peptide binding cleft of L(d) strongly suggest that the substitution of Arg for Trp(97) in L(d) alters the peptide binding cleft, increasing its affinity for endogenous peptides, which results in greater cell surface stability and better retention of beta2m. Furthermore, these results imply that Trp(97) plays an important role in the ability of L(d) to efficiently participate in alternative MHC class I Ag presentation pathways.
Assuntos
Substituição de Aminoácidos/imunologia , Epitopos/genética , Epitopos/imunologia , Antígenos H-2/genética , Peptídeos/genética , Peptídeos/imunologia , Polimorfismo Genético/imunologia , Microglobulina beta-2/metabolismo , Alelos , Substituição de Aminoácidos/genética , Animais , Arginina/genética , Linhagem Celular , Membrana Celular/genética , Membrana Celular/imunologia , Membrana Celular/metabolismo , Epitopos/metabolismo , Antígenos H-2/metabolismo , Meia-Vida , Antígeno de Histocompatibilidade H-2D , Isoantígenos/genética , Isoantígenos/metabolismo , Camundongos , Peptídeos/metabolismo , Ligação Proteica/genética , Ligação Proteica/imunologia , Transfecção , Triptofano/genéticaRESUMO
A number of classes of endogenous antibodies, including heterophile, rheumatoid factor, and autoantibodies, can interfere with immunoassay measurements of many different analytes. Heterophile and rheumatoid factor antibody interferences have been described previously for the AxSYM cardiac troponin I assay. Several commercial products have been developed to neutralize heterophile antibody interferences. We describe a patient with multiple apparently falsely elevated cardiac troponin I results that were unique to the AxSYM analyzer. These cardiac troponin I results diluted linearly. When treated with 2 different heterophile-blocking reagents, the magnitudes of the falsely elevated results increased 17- and 26-fold, and these results also demonstrated dilution linearity. This interfering substance could be removed by passage through an immobilized protein A column and by polyethylene glycol precipitation. It does not appear to be a classic heterophile antibody, nor is it a paraprotein. Laboratorians must remain constantly vigilant for immunoassay interferences that lead to clinically significant inaccurate results and must recognize that accepted methods for detecting and neutralizing the interference may be ineffective.
