Screening anti-human IgG for colloidal gold conjugate with dot immunogold filtration assay / 中国免疫学杂志

Objective:To screen the appropriate anti-human IgG (secondary antibody) for conjugating with colloidal gold by dot immunogold filtration assay,and put forward the method of using multiple evaluation indicators for secondary antibody comparing and analyzing. Methods:Three different secondary antibodies A,B and C from three different companies were screened. Firstly the titration was used to determine the optimal reaction conditions. Then three secondary antibodies were conjugated to the colloidal gold,tested with a dot immunogold filtration assay for hydatidosis specific antibody detection. The optimal conjugated secondary antibody were compared with the standard indirect enzyme-linked immunosorbent assay. Results:The optimal pH of three secondary antibodies were 8. 5,the binding capacity were 38. 4,24 and 19. 2μg /ml colloidal gold. According to the comprehensive evaluation,the diagnostic effects of sec-ondary antibody B was better than others. Its diagnostic effects in dot immunogold filtration assay was compared with enzyme-linked im-munosorbent assay. The Kappa was 0. 895(P<0. 05) and showed the two methods were in good agreement. Conclusion:The appropriate secondary antibody for conjugating with colloidal gold could be screened by dot immunogold filtration assay and the evaluation indicators which put forward by this study,the screening method would have an important reference value for all kinds of colloidal gold test kit to screen the suitable secondary antibody.

Partial characterization of a 29 kDa cysteine protease purified from Taenia solium metacestodes

A 29 kDa cysteine protease of Taenia solium metacestodes was purified by Mono Q anion-exchanger and Superose 6 HR gel filtration chromatography. The enzyme was effectively inhibited by cysteine protease inhibitors, such as iodoacetic acid (IAA) and trans-epoxy-succinyl-L-leucyl-amido (4-guanidino) butane (E-64) while inhibitors acting on serine- or metallo-proteases did not affect the enzyme activity. The purified enzyme degraded human immunoglobulin G (IgG), collagen and bovine serum albumin (BSA), but human IgG was more susceptible for proteolysis by the enzyme. To define the precise biological roles of the enzyme, more detailed biochemical and functional studies would be required.

A Pemphigus Vulgaris with IgG1 and IgG4 Subclass Autoantibodies

The distribution of pemphigus subclass autoantibodies in a patient with pemphigus vulgaris (PV) has been investigated by semiquantitative indirect immunofluorescence (IIF), using the HP series monoclonal antibodies specific for four human IgG subclasses on human foreskins. IgG1 and IgG4 intercellular substance-specific autoantibodies were detected in the serum of the patient, whereas IgG2 and IgG3 autoantibodies were absent. In addition to foreskins, human tonsillar epithelia were used as substrates of IIF for detecting the PV autoantibodies and it was one of satisfactory substitutes for monkey esophagus.

Assessment of three simple techniques for the screening of circulating immune complexes: Correlation with a parameter of complement consumption.

The sera of 36 normal controls, 45 patients with various diseases and 11 pregnant women were screened for circulating immune complexes using three relatively simple and inexpensive techniques. These included inhibition of agglutination of IgG coated latex particles with a serum having rheumatoid factor activity, polyethylene glycol precipitation and anti-complementary activity test. The circulating immune complexes were detected in a significantly higher proportion of patients as compared to normal controls. In the patients, the presence of circulating immune complexes did not always correlate with clinically detectable immunoinflammatory tissue damage indicating that pathogenic as well as nonpathogenic immune complexes were being detected by the above mentioned techniques. The alpha-1- antitrypsin/C3 ratio, however, correlated well with clinically apparent immunoinflammation.