Per Bygren

An ELISA for the detection of anti-neutrophil cytoplasm antibodies (ANCA).

An enzyme-linked immunosorbent assay (ELISA) has been developed for the detection of circulating anti-neutrophil cytoplasm antibodies (ANCA), which are defined by a diffuse, granular staining of the cytoplasm of alcohol-fixed human neutrophils by indirect immunofluorescence (IIF). Detection of antineutrophil cytoplasm antibodies has a high sensitivity and specificity for active Wegeners granulomatosis (WG) and reflects the effect of treatment. In the present enzyme-linked assay, immunoplates were coated with the cytoplasmic alpha fraction of neutrophils obtained from apparently healthy human donors by nitrogen bomb cavitation and subsequent Percoll gradient centrifugation. Alkaline phosphatase-labelled anti-human IgG was used as a secondary antibody. Diluted sera from 70 patients with WG and 16 patients with other diseases with anti-myeloperoxidase antibodies (anti-MPO) were examined. It is concluded that the ELISA accurately detects IIF ANCA positive patients with WG, is helpful in detecting WG patients in remission, is not influenced by the presence of anti-MPO and may help in detecting ANCA in cases with granulocyte-specific anti-nuclear antibodies since this IIF pattern obscures the IIF ANCA patterns. The ELISA with titration can be carried out in 3.5 h whereas a rapid test just to detect ANCA can be performed in 30 min.

Urinary Tract Infections Caused by Staphylococcus Saprophyticus: Recurrences and Complications

AbstractStaphylococcus saprophyticus is a coagulase-negative, novobiocin-resistant staphylococcus known to cause acute urinary tract infections in young women. We believe that the kidney can be involved in such infections. Of 57 randomly selected women with urinary tract infection caused by Staphylococcus saprophyticus clinical symptoms suggestive of renal involvement were reported by 29, 20 of whom had renal tenderness on examination. In 9 of 16 patients studied with the pitressin tannate test the renal concentrating capacity was reduced during the course of infection. All patients had regained the concentrating capacity on followup after antibiotic treatment. In the 57 women studied 38 had a history of recurrent urinary tract infection before entering the study. Recurrent urinary tract infection occurred in 17 patients during followup of 6 to 12 months, with Staphylococcus saprophyticus in 10 and gram-negative rods in 7. Twelve other women who had received treatment with nalidixic acid had persistent ur...

Circulating autoantibodies as serological markers in the differential diagnosis of pulmonary renal syndrome

Abstract. Objectives. Pulmonary renal syndrome (lung haemorrhage and glomerulonephritis) is a fulminant condition that warrants a rapid diagnosis and treatment to prevent mortality and preserve renal functions. However, the patients frequently present with non‐specific pulmonary symptoms in the early phase of the syndrome and the diagnosis is often missed. Recently, several autoantibodies have been described in association with various forms of glomerulonephritis. We evaluated the association as well as the diagnostic and the prognostic significance of these antibodies in pulmonary renal syndrome.

Anti-basement membrane antibody: immunoenzymatic assay and specificity of antibodies

A sensitive enzyme linked immunosorbent assay has been developed for circulating anti-glomerular basement membrane antibodies and optimal conditions for each steps have been determined. Structurally different preparations of antigens from glomerular basement membrane are coated to the walls of polystyrene tubes. Sera containing either specific antibody or negative control are added and allowed to react. Immunoglobulins bound to the antigens on the tube are then detected using anti-immunoglobulin antibodies coupled to alkaline phosphatase. The enzymic activity is measured using the fluorogenic substrate methylumbellipheryl phosphate. The fluorescence recorded is proportional to the concentration of specific antibodies in the serum tested. The specificity of the assay has been determined. The two different structural components of glomerular basement membrane were used as antigens. Type IV collagen was prepared after pepsin digestion, while glycoprotein components were solubilized by digestion of glomerular...

A rapid assay for circulating anti-glomerular basement membrane antibodies in Goodpasture syndrome.

A rapid ELISA for the detection of circulating anti-glomerular basement membrane antibodies in Goodpasture syndrome is described. The specificity of the test was shown to be highly dependent on the antigens used. Using the purified Goodpasture antigen it was possible to shorten the incubation times to 10 min in a routine assay using alkaline phosphatase-labeled second antibodies and the total assay was complete in 30 min. 200 reference sera, 500 sera from patients with various types of glomerulonephritis and 32 sera from patients with Goodpasture syndrome were analyzed by this rapid assay. The assay was able to discriminate between Goodpasture syndrome and other forms of glomerulonephritis. Using enzyme amplification it was possible to further shorten the incubation times to 1 min and the total time of the assay to 6 min.

Fibronectin-immunoglobulin complexes in the early course of IgA and Henoch-Schönlein nephritis

We have previously reported the presence of circulating IgA-fibronectin complexes in adult patients with primary IgA nephropathy. In the present study five children were serially investigated during the early course of IgA nephropathy and Henoch-Schönlein glomerulonephritis. Using affinity chromatography procedures and enzyme-linked immuno-sorbent assay, IgA, IgG and IgM in complex with fibronectin were repeatedly demonstrated during the follow-up period in both groups of patients. Most patients had, at the same time, IgA, IgG, as well as IgM deposits in the glomerular mesangium. The simultaneous presence of IgA and IgG in complexes purified from serum was furthermore demonstrated. The results are thus in contrast to the findings in adults with IgA nephropathy, in whom the immunoglobulin-fibronectin complexes only contained IgA. Whether this reflects different subgroups of patients or a different pathophysiology in children and adults remains to be elucidated.

Persistent High Prevalence of Thyroid Antibodies after Immunosuppressive Therapy in Subjects with Glomerulonephritis

The prevalence of thyroid antibodies, indicating an autoimmune thyroiditis, has been shown to be significantly increased in patients with autoimmune diseases. A 3-year prospective follow-up study of 42 patients with biopsy-confirmed glomerulonephritis is presented. Although the majority of patients had been treated with immunosuppressants, the prevalence of thyroid peroxidase antibodies was unchanged in both females and males, 47 and 15% respectively, at follow-up. Likewise, the prevalence of thyroglobulin antibodies was unaffected as was that of antinuclear antibodies (ANA) when analysing males and females together. However, for males there was a trend to higher prevalence for ANA at follow-up. On the other hand, the prevalence of antineutrophil cytoplasmic antibodies declined. Furthermore, thyroid antibodies were not restricted to membranous nephropathy, and notably found in 4 out of the 8 patients with vasculitis.

Human glomerular basement membrane. Heterogeneity of antigenic determinants

Human glomerular basement membrane was solubilized by digestion with proteolytic enzymes and immunoreactive components were quantitated and characterized by using rabbit antibodies raised against the particulate membrane. A number of antigens were demonstrated but they did not separate on gel filtration. However, two antigenic components in a collagenase digest of the membrane could be separated and isolated by Sepharose 6B chromatography. Chemical characterization suggests that both fragments are noncollagenous glycopeptides (molecular weights approx. 1,000,000 and 60,000--200,000, respectively).

Human glomerular basement membrane. Antigenic determinants in human urine

Antisera against particulate human glomerular basement membrane prepared from cadaver kidneys were raised in rabbits. It was shown that both normal individuals and patients with glomerular and tubular diseases excrete in their urine several antigens reactive with these antibodies. One antigen crossreacted immunologically with an antigen from human glomerular basement membrane while several others did not. One of the urinary antigens and the antigen crossreacting with the basement membrane were separated from the others by ion exchange chromatography and gel filtration, respectively. The pattern of anttigen excretion differed depending on the underlying renal disease but the multitude of different antigens detected complicates the interpretation of the patterns of excretion in different diseases.

Autoimmunity and glomerulonephritis.

Autoimmunity is now unequivocally regarded as the predominant pathogenic process underlying most forms of primary and secondary glomerulonephritis in humans. Most of the investigations so far have been focused upon humoral mechanisms. Consequently, the role of cell-mediated immunity in nephritis is still incompletely understood. Nonetheless, as a result of contemporary studies, a number of previously unidentified auto-antibodies in association with glomerulonephritis have been discovered. However, apart from anti-NC1 antibodies in the classical Goodpasture syndrome, the exact role of these auto-antibodies in the pathogenesis of glomerulonephritis yet remains undefined. This fact, however, does not undermine the relevance of exploring these auto-antibodies. They have been of immense help in sub-classifying glomerulonephritis previously thought homogeneous (Figure 3). Besides, analysis of auto-antibodies has assisted tremendously in the early diagnosis of rapidly progressive glomerulonephritis. This, in turn, has aided in early commencement of therapy thus contributing to regression in morbidity and mortality resulting from these disorders. Moreover, investigation of these auto-antibodies is of enormous value for future studies aimed at understanding the pathogenic mechanisms involved in glomerulonephritis.