Faculty Bibliography

A discriminating direct immunofluorescent test has been used to identify immunoglobulin inclusions in polymorphonuclear leucocytes (PMNs) isolated from the blood of patients with rheumatoid arthritis. These inclusions are thought to represent phagocytosed immune complexes, since normal PMNs incubated in RA sera known to contain raised levels of immune complexes developed similar immunoglobulin inclusions. Inclusions did not develop in normal PMNs incubated in normal serum. No correlation was found between the percentage of either RA blood PMNs with immunoglobulin inclusions or normal PMNs developing inclusions after incubation in RA sera, and levels of immune complexes in the corresponding sera. Using heat-aggregated IgG as a laboratory model of immune complexes, a simple relationship has been demonstrated between the uptake of IgG aggregates by normal PMNs and the concentrations of IgG aggregates in the test solutions over a concentration range of 12.5-200 micrograms . ml-1. These results indicate that the C1q- PEG test gives no measure of the actual amounts of immune complexes available in serum for phagocytosis

Tests for antiperinuclear factor (APF) demonstrable by indirect immunofluorescence (IF) on smears of human buccal mucosal cells and for antibodies to keratin (AKA) detected on cryostat sections of rat oesophagus were performed on serum from 102 cases of rheumatoid arthritis (RA) and 117 controls. APF was detected in 92% of the cases of RA; positive tests obtained with non-RA sera were generally weaker than those given by the RA group, and the antibody in both RA and non-RA serum was predominantly IgG class. The difficulty in obtaining suitable substrate material previously reported was confirmed, and only 2 satisfactory donors were identified among 27 individuals tested. The incidence of keratin antibodies detected was found to be related to the site from which the tissue was taken; low oesophagus provided the best discrimination between RA and controls (51% and 5% positive respectively), and cardia of the stomach gave the highest incidence of staining in all groups. A laminar staining pattern was seen with most positive sera, but occasionally the keratinised layer was diffusely stained. The presence of AKA showed a marked correlation with both IgM rheumatoid factor and increased Clq binding in RA, but APF did not

Measurement of the absorbance due to light scattering at 40 nm proved to be a simple and reliable way of assessing the extent of aggregation in heat-treated IgG solutions. Using this technique the rate of aggregation was demonstrated to be markedly temperature dependent with a sharp inflexion in the curve close to 63 degrees C. During heating at 63 degrees C the concentration of unaggregated IgG fell in a manner consistent with a first-order process and the mean size of the IgG aggregates increased with time. IgG aggregates could be selectively removed from heat-treated IgG solutions and concentrated by precipitation with 3.5% polyethylene glycol without altering their size distribution. Furthermore, and in contrast with a previous report, the IgG aggregates examined in this study were remarkably stable in the absence of any other protein such as serum albumin. In consequence, several important practical recommendations concerning the production of heat-aggregated IgG for use in immune complex assays are made

The reactivity of heat-aggregated IgG of known size, in the Raji cell assay, the C1q binding assay and the C1q solid phase radioimmunoassay as a function of concentration, has been investigated. Marked differences were found in the way that the three assays behave when the IgG concentration and aggregate size are varied. These findings indicate the pitfalls in attempting to express the results of immune complex assays performed on biological fluids in terms of equivalent concentrations of aggregated IgG