Plasma membranes of pig liver were isolated by means of an aqueous two-phase polymer system and sucrose gradient. The plasma membranes were demonstrated to have high purity when measured by 5'-nucleotidase activity. To our knowledge, this is the first demonstration of radioreceptor assay for insulin using pig liver plasma membranes. The technical performance of the assay is easy and rapid, and the assay system has a detection limit of 15μU/m/ of porcine insulin. A radioreceptor assay for insulin should provide a useful tool for evaluation of biological activity of circulating human insulin and its derivatives.
Conventional insulin preparations (Novo), 5 times recrystallized, were contaminated with approximately 1 to 2 % proinsulin-like substances. Anti-insulin antibodies and even proinsulin-specific antibodies were found in many cases of diabetics treated with these insulin preparations. Monocomponent insulin, purified in order to eliminate its antigenicity for insulin, appeared to be less immunogenic than conventional insulin. In this study, the contaminating proinsulin-like substances in Monocomponent Lente insulin (Batch 522*5), Monocomponent Actrapid insulin (Batch 500*5) and conventional Lente insulin (Lot. No.297961) were estimated directly and after gel filtration by porcine proinsulin-speufic specific radioimmunoassay. Although gel filtration of conventional insulin (500 ng) revealed proinsulin-like components (PLC), these PLC were not detectable after gel filtration of 500 ng and 5000 ng of Monocomponent insulin by specific radioimmunoassay. Monocomponent insulin preparations contained less than O.0001% of porcine proinsulin-like immunoreactivity (PLI) by weight as estimated directly utilizing specific radioimmunoassay for porcine PLI.
An association has been demonstrated between the HLA antigens and Bw 15 and juvenile dibetes mellitus in Caucasians. However HLA-B 8 is almost absent in the Japanese. For this reason, a racial point, HLA typing was performed by a lymphocytotoxicity micromethod on 60 insulin-dependent diabetes mellitus patients who had developed diabetes mellitus by the age of 15 years, and on 355 healthy controls. 27 HLA antigens were tested with 100 selected antisera. In these patients, we observed an increased incidence of HLA-BW 22.1: 25.0 per cent in patients as against 5.6 per cent in controls, HLA-BW 22.2: 23.3 per cent in patients 4.5 per cent in controls. This difference is highly significant.