The relationship between fluorescent, agglutinating, and precipitating antibodies to Candida albicans and their immunoglobulin classes

A parallel study of fluorescent, agglutinating, and precipitating antibodies to Candida albicans revealed that precipitating antibodies belong to the IgG class, whereas agglutinating antibodies reside in the IgG, IgM, and IgA classes. The three types as well as the three classes of antibodies were found in Candida endocarditis and mucocutaneous candidiasis. Immuno-absorption studies suggest that the three serological tests estimate antibodies to mannan determinants of Candida albicans.     

Demonstration by immunoelectro-osmophoresis of precipitating antibodies to a purified rubella virus antigen.

The nonionic detergent Triton X-100 was used to extract antigens of rubella virus from infected tissue culture cells. Three virus-specific antigens were demonstrated by crossed immunoelectrophoresis by using a pool of human gamma globulin as antiserum. The most dominant of these antigens were purified by ion-exchange chromatography on diethylaminoethyl-cellulose. This antigen was of glucoprotein nature and had slow electrophoretic motility and low binding capacity to diethylaminoethyl-cellulose. Thus, it seems likely that the antigen is identical with the precipitating antigen of rubella virus designated b-antigen or tro-osmophoresis with precipiting antibody in sera obtained from patients recovering from acute postnatal rubella. The precipitin reaction that could be correlated to the hemaglutination-inhibition titers of the same sera appeared 12 days after onset of the disease and remained positive for several years.     

Production of agglutinating monoclonal antibody against antigen 8 specific for Cryptococcus neoformans serotype D.

A hybridoma (clone CRND-8) that produced agglutinating monoclonal antibody (MAb) against Cryptococcus neoformans serotype D was established by using a soluble capsular polysaccharide-keyhole limpet hemocyanin conjugate for immunization. The isotype was immunoglobulin M(kappa). Specificity was determined by cell slide agglutination and enzyme-linked immunosorbent assay (ELISA). In both tests, the MAb reacted to serotypes D and A-D but not to serotypes A, B, and C. Furthermore, the specificity of the MAb determined by ELISA was the same as that of polyclonal antibody factor serum (PAb factor) 8, which showed high-level reactivity with serotypes D and A-D. These results supported the deduced specificity of the PAb-based antigenic factor 8. A total of 15 isolates of serotypes D and A-D but no serotype A isolates reacted with the MAb in cell slide agglutination tests. CRND-8 MAb can be used in place of PAb factor 8 for serotyping C. neoformans isolates and for the analysis of the antigen 8 epitope.     

Characterization of precipitating antibodies to E. coli O antigen in infants and children with acute pyelonephritis

Precipitating serum antibodies to Escherichia coli O antigens from some infants and children with acute pyelonephritis were studied.     

The occurrence of antigen reacting with antibody to porcine neurophysin

1. Antiserum was raised in rabbits to neurophysin prepared from posterior pituitaries of pigs. The presence of antibody reacting with porcine neurophysin was demonstrated in precipitation, gel diffusion and immunoelectrophoretic tests.

2. The antibody was species specific and did not react with protein from rat, bovine or guinea-pig neurohypophyses.

3. The occurrence of antigen reacting with anti-neurophysin serum was demonstrated in acetic acid extracts of porcine kidney but there was no evidence for the presence of antigen in extracts of liver or spleen prepared in the same way.

4. Protein fractions (`N-fractions') from various organs and tissues of pigs were obtained by the same methods used to prepare neurophysin from posterior pituitaries and were tested for antigenicity in reaction with neurophysin antibody. N-fractions from kidney, uterus, mammary gland and serum contained antigen while fractions from liver, spleen, brain and skeletal muscle did not react with the anti-neurophysin serum.

     

A pomona serogroup-specific, agglutinating antigen in Leptospira, identified by monoclonal antibodies

Monoclonal antibodies were produced by hybridoma cell lines derived by fusion of mouse NS-1 myeloma cells with splenocytes from mice immunized with Leptospira interrogans serovar pomona. One hybridoma (A3) produced an IgG2a antibody which agglutinated all leptospires of the Pomona serogroup but not leptospires representative of serovars of any other serogroup. The antibody precipitated in immunodiffusions with either alkali- or phenol-extracted lipopolysaccharide and also with the TM antigen of Yanagawa et al., indicating a common determinant on both antigens. A3 antibody opsonized viable leptospires for phagocytosis by mouse macrophages in vitro.     

Purification of TR-b, a Reiter treponeme protein antigen precipitating with antibodies in human syphilitic sera.

TR-b is a Reiter treponeme antigen, cross-reacting with an antigen in Treponema pallidum (Nichols pathogenic strain). Sera from patients with secondary syphilis contain precipitating antibodies against TR-b. The isolation of TR-b from a bacterial sonic extract is described here. It involved five fractionation steps: anion-exchange chromatography (DE-52 Whatman), gel filtration (Ac-A-22 Ultrogel), and affinity chromatography on phenyl-Sepharose CL 4B, iminodiacetic acid-Sepharose CL 4B, and lysine-Sepharose 4B, respectively. The purified TR-b was enriched 199 times compared with the starting material, and the recovery was 12%. TR-b was shown to be a protein; it did not bind to a series of lectins, and by gel filtration and polyacrylamide gel electrophoresis, the molecular weight was determined to be 610,000 to 630,000. It was found by sodium dodecyl sulfate-polyacrylamide gel electrophoresis to be composed of identical 70,000-dalton subunits. The isolated TR-b was immunologically pure when tested in crossed immunoelectrophoresis against polyspecific anti-Reiter immunoglobulin. The purified TR-b antigen was used for the production of a monospecific rabbit antiserum, giving strong fluorescence with both the Reiter treponeme and T. pallidum in an indirect immunofluorescence test.     

The detection of precipitating antibodies in equine infectious anemia and partial purification of the antigen

Summary Spleens from 12 ponies experimentally infected with the virus of equine infectious anemia (EIA) were evaluated as antigen sources against sera from known infected and normal horses using the Ouchterlony double diffusion technique. Spleens from 4 normal horses served as controls. Two of the 12 infected spleens could be used as antigen for detecting precipitating antibodies when the spleens were frozen and thawed, minced and placed in the antigen wells. When 9 of the infected spleens were treated to partially purify and concentrate the antigen, all 9 could be used as antigen. Using partially purified antigen, precipitating antibodies could not be detected in any of 107 non-EIA infected sera while antibodies could be demonstrated in 71 of 83 sera from infected horses. The 12 non-reacting sera were from 10 horses infected 14 days or less, from one horse infected 15 days, and from one horse infected 25 days. Eighty-two of 83 horses infected from 18 days to 5 1/2 years had precipitating antibodies detectable by this technique. A precipitinogen can be partially purified from EIA infected spleens and used to detect precipitating antibodies in EIA infected horses.     

Characterization of a precipitating antigen detected in the serum of patients with viral hepatitis

During a search for the aetiological agent of non-A non-B hepatitis, a precipitating antigen was detected in the sera of some patients during the acute phase of their illness. The antigen was detected by agar gel diffusion using antibody from convalescent sera obtained from patients with non-A non-B hepatitis, and from haemophiliac sera. The antigen was usually detected early in the patient's illness, disappearing as liver function tests returned to normal. In some patients specific antibody appeared during the convalescent phase of the disease. The antigen does not appear to be specific for non-A non-B hepatitis, as it could be detected with similar frequency in patients with hepatitis A or hepatitis B and some patients with other liver disorders. Biochemical and biophysical studies suggest that the antigen is probably an abnormal lipoprotein produced as a result of acute liver damage.     

Influence of agglutinating antibody in experimental cryptococcal meningitis.

A model for chronic Cryptococcus neoformans meningitis in corticosteroid-treated rabbits was used to determine the influence of pre-formed agglutinating antibody to cryptococcal polysaccharide on the progress of this infection. Immunized rabbits developed serum agglutinating antibody with a geometric mean titre of 1:32, but none was detected in cerebrospinal fluid. Prior immunization did not enhance immunity to infection, had no effect on the number of viable cryptococci in cerebrospinal fluid, and did not prevent dissemination outside the central nervous system. Future investigations in this field should focus on cellular rather than humoral defence mechanisms.     

An immunochemical analysis of precipitating and non-precipitating idiotype-anti-idiotype reactions.

Two monoclonal IgG3 syngeneic anti-idiotypes are described which form soluble and insoluble complexes with anti-alpha(1----6)dextran hybridoma and myeloma proteins. Specific precipitation was seen when purified anti-alpha(1----6)dextrans were added to ascitic fluid containing IgG3 kappa anti-idiotype. Analysis of the supernatants of the idiotype-anti-idiotype precipitates demonstrated the presence of soluble complexes whose mobilities in polyacrylamide gels could, in some cases, be distinguished from that of free anti-idiotype. An IgG1 kappa anti-idiotype is described which did not form precipitates with anti-alpha(1----6)dextrans unless 3% PEG 6000 was added.     

Studies on the nature of the cell surface antigen reacting with cytolytic T lymphocytes in murine oncornavirus-induced tumors.

The nature of the target antigen, expressed on murine sarcoma virus (MSV) and related murine tumors which reacts with T killer lymphocytes, remains ill-defined. The experiments reported here show that: (a) the previously described H-2 restriction phenomenon is found under all experimental conditions including 3–4 and 16–20-h chromium release tests. With 16–20 h tests and highly efficient T lymphocytes however, quantitative methods are necessary to demonstrate the H-2 restriction. These results support the hypothesis that H-2 molecules may be determinant in the structure and/or in the function of the cytolytic T lymphocyte (CTL) reactive antigen. (b) Under syngeneic conditions (i.e. using H-2-identical immune lymphocytes, stimulators and target cells), the pattern of specificities recognized by anti-MSV or anti-Friend T killer cells on 20 different lymphomas suggests that the main reactive antigen is an “FMR-like” substance. Identical conclusions were drawn from competition experiments. (c) Blockings were obtained by pre-incubation of the target cells with a goat anti-gp70, suggesting a possible role of the viral gp70 in the antigen recognized. However, this could be due to nonspecific reactions as two other anti-gp70 sera as well as with antisera directed against the viral components gp45, pr60, p30, p15, p12 and p10 did not block. The CTL/tumor cell interaction was not inhibited by virion-associated antigens added to the medium. Lysostrip and co-capping experiments have failed to reveal an association between H-2 and gp70. The nature of the viral protein bearing the “FMR-like” substance therefore remains to be established.     

Improved methods of producing precipitating aspergillus antigens.

Improved methods for producing Aspergillus fumigatus antigens capable of giving sensitive and reproducible results by counterimmunoelectrophoresis (CIEP) are described. Among the factors investigated were the source of casein hydrolysate used for N source, method of incubating the CIEP slide following a run, effect of freeze pressing and lyophilization, age of culture and ratio of C to N source in the medium.