PLAZ~ZS 92 2"o 96 (Received for publication, July 24, 1959) The rheumatoid factor, a group of unusual macroglobulins, is demonstrable in the serum of the majority of patients with rheumatoid arthritis and in a relatively small proportion of patients with other diseases (1). The presence of this factor can be demonstrated by its reaction with human v-globulin (Cohn fraction II) in such serological procedures as the tanned sheep cell agglutination (2) and latex fixation tests (3), as well as by precipitin tests (4). The reactive component of "y-globulin seems to be an aggregate (5) with high sedimentation coefficients, which forms an insoluble complex with the rheumatoid factor. The origin and site of production of the rheumatoid factor has been unknown. I n the present study, fluorescein-labelled aggregated human 7-globulin was used as a specific reagent for its detection in situ. The results show that the factor is present in certain cells of the synovial tissues, lymph nodes, and subcutaneous nodules in active rheumatoid arthritis. Materials and MethodsFluorescein-Labdled Aggregated Human T-Globutin.--Human T-globulin (Cohn fraction II, E. R. Squibb Lot 1788), obtained as a gift from the American National Red Cross, was conjugated at a concentration of 20 rag. protein per ml. with fluorescein isothiocyanate (Sylvana Chemical Co., Orange, New Jersey), in accordance with the procedure of Riggs and associates (6), and dialyzed against buffered saline (0.15 ~ sodium chloride containing 0.01 M phosphate, pH 7.0) until free of unconjugated fluorescein. The fluorescein-labelled human "y-globuiin preparation was heated in a water bath to 62°C. for l0 minutes, and 40 per cent by volume of 2.18 M Na2SO, was then added, with stirring, in a manner used by Christian (5). A precipitate was collected after overnight storage in the cold, dissolved in saline, and dialyzed against phosphate buffer (0.15 x¢, pH 7.0). The dialyzed preparation was centrifuged at 18,400 a~.v.~, for 1 hour prior to use. This preparation will be referred to in this report as fluorescein-labelled aggregated human ~,-globulin. This reagent was six times more reactive in precipitation reactions with rheumatoid serums than a similarly prepared but unheated q,-globulin preparation, and furthermore was live times more reactive than a heated preparation in which the Na2SO4 treatment was omitted. The various preparations were also analyzed by ultracentrifugation. Finorescein-labelled aggregated human T-globulin contained approximately 34 per cent of the 7S component and 66 per cent of an aggregate having sedimentation coefficients varying from approximately 10S to 70S. The analysis was performed in a Spinco,

No abstract

Sera of individuals with Burkitt's lymphoma and nasopharyngeal carcinoma, tested by consumption of hemolytic complement, were found by comparison with healthy individuals to have significantly increased levels of circulating immune complexes. The identity of the immune complexes was established by sucrose density gradient ultracentrifugation, which showed them to sediment between 10 and 19S. As they were adsorbable by rheumatoid factor--Sepharose 4B conjugates, it appeared that these complexes were composed of IgG. The complexes were retained by Concanavalin A--Sepharose columns and eluted by alpha methyl-D-mannoside, suggesting by analogy with model complexes that the antigens might be glycoproteins.

13 The interpretation of an experiment of this type given in our previous publication (op. cit.) is in error. The results of that experiment are in qualitative agreement with those of similar experiments reported in the present paper.

Radioimmunoassays for anti-F(ab')z antibodies, which feature the use of goat anti-human Fc antibody for correcting potentiation of IgM anti-F(ab')z antibody titers by endogenous IgM anti-Fc antibodies (rheumatoid factors), are described. Individuals with classic rheumatoid arthritis had significantly more IgM anti-F(ab')2 antibody (P < 0.001) and IgG anti-F(ab')2 antibody (P = 0.05) than did individuals with systemic lupus erythematosus or normal volunteers. There is some similarity in patterns of isotype distribution of anti-F(ab')z antibodies and rheumatoid factors.