Ralph E. Schrohenloher

A quantitative assay for IgA rheumatoid factor

We developed a solid-phase radioimmunoassay capable of detecting nanogram quantities of human IgA rheumatoid factor (RF) in biological fluids. Human IgM RF, IgG RF, IgG, IgA, IgM and whole serum did not significantly interfere with the IgA RF assay. Patients with sero-positive rheumatoid arthritis (RA) had significantly higher concentrations of IgA RF than sero-negative RA patients or healthy adult controls. Concentrations of IgA RF in paired sera and synovial fluids from sero-positive RA patients were comparable. Levels of IgA RF demonstrated a moderately good correlation with levels of IgM RF in sero-positive RA sera (r = 0.673). However, the ratio of IgA RF concentration to IgM RF concentration in sero-positive RA sera varied widely.

The degradation of human γ-globulin by trypsin

Abstract The 3.5 S fragments produced from normal human 7 S γ-globulin by the action of trypsin were isolated by DEAE-cellulose chromatography and gel filtration through Sephadex G-75 and were subsequently characterized by ultracentrifugation, immunoelectrophoresis, and double gel diffusion analysis. The degradation of the γ-globulin by trypsin resulted in the production of fragments which closely resembled those produced by cysteine-activated papain. Corresponding fragments split from the same γ-globulin by the two enzymes were indistinguishable by the methods employed with respect to antigenic determinants, sedimentation characteristics, and chromatographic behavior on DEAE-cellulose. Corresponding fragments also appeared identical by immunoelectrophoresis, except for a minor difference in the electrophoretic mobility of one of the fragment types. Two major subfractions of the γ-globulin obtained by DEAE-cellulose chromatography were each similar to the unfractionated γ-globulin with respect to the products obtained by enzymic degradation.

Gamma globulin complexes in rheumatoid pericardial fluid

Cardiac tamponade due to pericarditis occurred in a patient with rheumatoid arthritis. Aspiration afforded us an opportunity to expand the characterization of pericardial fluid. Elevated acid phosphatase levels, decreased whole hemolytic complement and gamma globulin complexes similar to those found in rheumatoid synovial fluid were noted, supporting the concept of a unitary nature of inflammation in rheumatoid disease.

Tryptophan Metabolite Excretion in Connective Tissue Diseases Demonstrating a Difference Between Rheumatoid Spondylitis and Rheumatoid Arthritis.

Summary Urinary excretion of kynurenine was measured in a group of patients with various connective tissue diseases and compared to normal controls. Elevated kynurenine excretion was found in patients with rheumatoid arthritis, systemic lupus erythema-tosus, and in 2 patients with polymyositis; whereas excretion of kynurenine in patients with scleroderma and rheumatoid spondylitis was not significantly different from normal controls. The demonstrated difference in kynurenine excretion between patients with rheumatoid spondylitis and those with rheumatoid arthritis in this study provides additional evidence that the two conditions are distinct clinical entities. The valuable technical assistance of Miss Geneva Blackburn is gratefully (acknowledged. The authors also wish to thank Miss Carol Brewster for supervision of the diet used in this study.

Studies of the component chains of human IgM by citraconylation

Abstract Partial reduction and alkylation of a Waldenstrom IgM followed by reaction with citraconic anhydride resulted in modification of 92–95% of the free amino groups. The citraconylated μ and κ chains were obtained in pure form by chromatography on Sephadex G-200 in a nondenaturing solvent. The molecular weight determined from the sedimentation and diffusion coefficients for the citraconyl μ chain was 83 000 and that for the citraconyl κ chain was 23 000. Substraction of the weight contributed by the citraconyl groups gives estimated residual molecular weight for μ chain of 77 000 and for κ chain of 22 000. Deblocking of citraconyl μ and κ chains was achieved by incubation at pH 4 for 3 h. The deblocked chains showed a multiple banding pattern in acrylamide disc gel electrophoresis. Peptide maps of tryptic digests of deblocked citraconyl μ, maleyl μ and μ chains obtained by conventional methods showed considerable susceptibility to tryptic digestion.

Studies on the mechanism of the degradation of human γ-globulin by trypsin☆

Abstract The presence of 0.05 M N -ethylmaleimide resulted in a marked inhibition of the production of 3.5 S fragments from normal human 7 S γ-globulin by trypsin. The presence of 0.1 M iodoacetamide also resulted in an inhibition of the fragmentation process. However, the component distribution of the resulting digests differed in that the principal component of the N -ethylmaleimide inhibited digests sedimented at the same rate as the unfragmented γ-globulin, while that of the iodoacetamide inhibited digests sedimented at a somewhat reduced rate. Reduction of the N -ethylmaleimide or iodoacetamide inhibited digests by 0.1 M 2-mercaptoethanol resulted in the conversion of faster sedimenting materials to 3.5 S fragments. Inhibition of the production of 3.5 S fragments by tryptic degradation in the absence of added sulfhydryl blocking reagent was also observed following prior treatment of the γ-globulin by N -ethylmaleimide. The findings of this study provide evidence that the free sulfhydryl groups of human γ-globulin are involved in the splitting of disulfide bonds during the fragmentation of the γ-globulin by trypsin. Rabbit 7 S γ-globulin differed from human γ-globulin in that treatment by trypsin did not produce 3.5 S fragments except in the presence of a reducing reagent.

Isolation from plasma of an α1-glycoprotein resembling “orosomucoid”: A simple method for its estimation in blood and tissues☆

Abstract An acid α 1 -glycoprotein has been isolated from human plasma, and closely resembles “orosomucoid” as to its ultracentrifugal and electrophoretic behavior and chemical composition. However, the two preparations differed from each other immunologically. The data obtained suggest that the isolated protein is actually “orosomucoid,” but probably less denatured than the preparations obtained by other procedures. A method is described for the isolation and estimation of this α 1 -glycoprotein from plasma and tissues, which, owing to its simplicity, is suitable for use in routine clinical laboratories. The procedure described is also highly specific, as indicated by over 90% purity of the isolated product.

Serum Immunoglobulins in Gout

Summary Serum levels of IgA, IgM, and IgG were determined in a group of 25 patients with primary and secondary gout. Significant elevations of IgA were found in both groups.