A simple sensitive radioreceptor assay for calcium antagonist drugs

A radioreceptor assay for calcium channel antagonist drugs described here is based on the ability of these drugs to affect ³H-nitrendipine binding to calcium channels. All the known calcium channel antagonists may be assayed in this manner. The assay can detect 10–100 nM (4 – 40 ng/ml) nimodipine, 10–100 nM (3.5 – 35 ng/ml) nifedipine, 3–30 μM (1.2 – 12 μm/ml) prenylamine, 0.1 – 1.0 μM (49 – 490 ng/ml) verapamil and 3–30 μM (1.2 – 12 μg/ml) diltiazem. These values cover the range of concentrations of calcium channel antagonists that are clinically important. As the radioreceptor assay detects active metabolites as well as the parent drugs, it should prove a useful adjunct in cardiovascular therapy. The method is more reproducible, simpler and less expensive than other methods such as high pressure liquid chromatography.     

Preferential association of kappa IIIb light chains with monoclonal human IgM kappa autoantibodies

The predominance of the relatively uncommon V region subgroup isotype kappa III among the light chains of human monoclonal (IgM kappa) anti-IgG antibodies, (i.e., rheumatoid factors), was further documented through sequence analyses of ten such autoantibodies isolated from IgM-anti-IgG cold-insoluble immune complexes (mixed cryoglobulins). The amino-terminal sequence of all ten kappa-chains was characteristic for kappa III proteins and virtually identical to that of a prototype kappa III light chain. Similar sequence identity was found for kappa-chains isolated from three IgM kappa autoantibodies that formed cold-insoluble immune complexes with low-density lipoprotein (LDL). The thirteen light chains were found to be virtually identical in sequence for the first framework region (FR); ten of these proteins sequenced through the first complementarity-determining region (CDR) and into the second FR were markedly similar. The second CDR of five proteins was almost identical in sequence to that of the prototype kappa III-chain. Concordance was also demonstrated between the structural classification of the light chains as kappa III and their immunochemical classification as members of this V region subgroup. Serologic analyses of light chains isolated from seven IgM kappa autoantibodies (six anti-IgG, one anti-LDL) and of one intact IgM kappa anti-LDL antibody showed that each had antigenic determinants common to kappa II proteins. These light chains also expressed the antigenic determinant(s) of a V-region sub-subgroup of kappa III proteins designated kappa IIIb. Our studies confirm the preferential association of kappa III (and kappa IIIb) light chains with IgM kappa anti-IgG antibodies and demonstrate a similar association for IgM kappa anti-LDL antibodies. The finding that these and other types of IgM kappa autoantibodies, e.g., cold agglutinins, have remarkably similar light chains suggests an inherent restriction in the immune response to self-antigens.     

Structural studies of an Inv (1, −2) kappa light chain

A Bence Jones protein with phenotype Inv (1, –2) was isolated from the urine of a patient with multiple myeloma. Inv typing of the patient's relatives established the presence of anInv ¹ allele in the kindred, and that the patient's genotype wasInv ¹/Inv ³. Hence, the absence of Inv (2) in the Bence Jones protein was shown to be genetic and not an artifact caused by the disease. The tryptic peptide-containing residues 191 through 194 were isolated and shown to be composed of Leu, Tyr, Ala, Cys, with Leu at the amino end. Hence, the residue at 191 is the same as that present in Inv (1, 2) Bence Jones proteins. More detailed study of the tryptic peptides established that residue 153 is Val rather than Ala as in all other K chains thus far studied. The primary sequence: Ala¹⁵³, Leu¹⁹¹ determines Inv (1, 2); Ala¹⁵³, Val¹⁹¹ determines Inv (3); and Val¹⁵³, Leu¹⁹¹ determines Inv (1). The Val¹⁵³, Val¹⁹¹ sequence has not been observed. It may correspond to Inv (–). These data are strikingly similar to the data for the Kern and Oz isotypes (changes at 154 and 191, respectively) in the chain. As in the case of theK chain, only three of the four possible combinations have been observed. The implications of this parallelism and of crystallographic findings on chains, reported by others, are discussed.     

Serological Studies of Clostridium botulinum Type E and Related Organisms

Clostridium botulinum type E antigens prepared from washed cells by either Formalin treatment or heating at 100 C were used for immunizing rabbits. Agglutination tests showed that high levels of antibody were produced by both types of preparations. Flagellar antigens were highly strain-specific, whereas the somatic antigens were sufficiently similar to produce complete cross-agglutination. One toxigenic strain produced toxigenic and nontoxigenic progeny which were physiologically and antigenically identical in all other respects. Other nontoxigenic strains whose growth, physiological, and morphological characters were identical to type E and strains which had some physiological differences completely cross-agglutinated with type E strains via the somatic antigen. Neither type of antiserum agglutinated other clostridia against which they were tested except for C. acetobutylicum. This reaction seems to be due to a nonspecific anamnestic response and does not appear to be related to the immunizing strains. The nontoxigenic strains studied seem to have no greater antigenic differences from type E strains than the type E strains have from each other.