HLA Class I Biochemistry: Definition and Frequency Determination of Subtypes by One‐Dimensional Isoelectric Focusing and Immunoblotting

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Abstract
Previous biochemical studies of HLA class I gene products were based on the use of radiolabelled lymphoblastoid cell lines and/or mitogen-stimulated peripheral blood lymphocytes (PBL). We present here a nonradioactive methodology for defining HLA class I polymorphism on a small number of cells, such as 5times106lymphocytes, the vitality of which is not a requirement in contrast to radiolabelling techniques, where 1–2times106PBL are needed. We analyzed 54 serologically typed families with a total of 261 individuals and 214 haplotypes by one- dimensional isoelectric focusing and subsequent immunoblotting, using a polyclonal anti-class I heavy chain serum. The banding patterns segregated within families and correlated with the serological results. A substantial number of HLA-A and HLA-B products appeared biochemically heterogeneous allowing the definition of subtypes and the calculation of their frequencies.