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Biomedical subjects

S Avrameas

Publications and source records attributed to S Avrameas.

At least 271 records · Page 15Linked to original sources

Solid-phase immunoassays using chimera antibodies prepared with monoclonal or polyclonal anti-enzyme and anti-erythrocyte antibodies.

The possibility of using covalently prepared chimera antibodies in solid phase immunoassays was tested. The antibody specific for the antigen to be quantified was conjugated with a polyclonal or monoclonal antibody specific for an enzyme or for erythrocytes. Using these conjugates, labelling was done by enzyme/anti-enzyme or erythrocyte/anti-erythrocyte interactions. The chimera antibodies were used for quantitation of human IgE and for titration of human anti-Echinococcus granulosus antibody by solid-phase immunoassays, and were found to possess equal or higher amplification than conventionally prepared enzyme-antibody conjugates. A washing device for erythroadsorption test is also described. It allows such a test to be expressed in quantitative terms, as is the case in enzyme-immunoassays.

Animals↗

Cross-reactive idiotypic reactions between hybridoma proteins with or without anti-peroxidase antibody function.

Hybridomas from lymph node cells of a mouse C57BL/6 immunized with peroxidase were prepared. Two hybridoma proteins, one with and one without reactivity towards peroxidase, were selected and used to prepare antiidiotypic antisera in rabbits. The idiotypic cross-reactions between these two hybridoma proteins were studied using these antiidiotypic antisera. The results obtained indicate that at least two main idiotopes are present on the non-peroxidase binding hybridoma protein, only one of which is shared by the peroxidase-binding hybridoma protein.

Animals↗

Sandwich enzyme immunoassay of human chorionic gonadotropin using polystyrene beads as solid support.

A solid-phase sandwich enzyme immunoassay for human chorionic gonadotropin (hCG) employing monoclonal antibodies directed against beta- and alpha-subunits is described. Polystyrene beads used as solid support were coated with monoclonal anti-beta-hCG antibody. Monoclonal anti-alpha-hCG antibody was labelled with the enzyme, horseradish peroxidase. The assay could be performed by a "one-step" or "two-step" procedure. The sensitivity of the two-step procedure was 7.8 ng hCG/ml with a linear range up to 500 ng hCG/ml. The one-step procedure had a measurement range between 15.6 and 125 ng hCG/ml. Within the limits of measuring ranges, the two procedures had similar accuracy, the correlation coefficient being 0.987. Twenty-one urine samples from pregnant women were analysed by the two-step procedure and also by radioimmunoassay. The results were comparable, with r = 0.956.

Antibodies, Monoclonal↗

Both natural and induced murine anti-TNP antibodies possess anti-DNA activity.

Sera of BALB/c mice immunized with either trinitrophenyl (TNP) coupled to bovine serum albumin (TNP/BSA) and Freund's adjuvant or TNP/BSA and TNP coupled to mouse serum albumin (TNP/MSA) without adjuvant were tested for antibody activity against a panel of antigens: TNP/BSA, DNA, actin, myosin, tubulin and transferrin. Anti-DNA antibody activity in the serum increased significantly in immunized mice; this activity followed the increase in the anti-TNP titres. Sera from MRL/lpr/lpr autoimmune mice were also tested for antibody activity against a panel of antigens. Again, it was found that the level of the anti-TNP antibodies followed the level of anti-DNA antibodies. Purified antibodies isolated from both immunized BALB/c and MRL/lpr/lpr mice on a dinitrophenol(DNP)-lysine immunoadsorbent shared antibody activity against both TNP and DNA. The results obtained suggest that a single region in the antibody binding site is responsible for both anti-DNA and anti-TNP activities, that TNP-like groups may represent a "public epitope" carried by various antigens and that antibodies obtained after experimental immunization and antibodies appearing in autoimmunopathological situations apparently possess similar specificities.

Animals↗

Detection of natural autoantibodies in the serum of anti-HIV-positive individuals.

Twenty-six sera containing anti-HIV antibodies from individuals with AIDS, AIDS-related complex, lymphadenopathy syndrome, or from seropositive individuals without clinical symptoms, were tested for the presence of natural autoantibodies (NAb) to actin, DNA, tubulin, thyroglobulin, albumin, myosin and TNP-BSA. Sera from 22 seronegative individuals at high risk for HIV infection were also examined. NAb titres to all these antigens were found to be elevated in both groups as compared to the titres in pooled normal human sera. The most prevalent NAb were those of the IgG class directed against TNP found in high titre in seropositive individuals. On the basis of the above results, sera from 50 anti-HIV-positive and 67 anti-HIV-negative individuals were tested for IgG antibody activity against a panel of protein antigens with different degrees of TNP substitution. The greater the TNP substitution, the higher the titres of IgG antibodies detected in anti-HIV-positive sera. Antibody titres in seronegative individuals at high risk for HIV infection were independent of the degree of TNP substitution. In almost all cases, the highest titres of IgG anti-TNP antibodies were found in anti-HIV-positive sera from individuals with clinical manifestations.

AIDS-Related Complex↗

[Anti-actin and anti-tubulin antibodies in the serum of non-immunized animals].

The preparation and the purification of anti-action and anti-tubulin antibodies is reported. The occurrence of low amount of IgG anti-tubulin was also found in the sera of non-immunized animals from various species. The staining of the cellular network of microtubules as well as of tubulin paracrystals was observed with both induced and natural antibodies.

Actins↗

Analysis of autoantibody reactivities in hybridoma collections derived from normal adult BALB/c mice.

Hybridoma collections were produced from lipopolysaccharide-stimulated adult BALB/c spleen cells, small resting spleen B cells and large activated spleen lymphocytes. The hybridomas were examined for production of immunoglobulins and of antibodies directed against a panel of self (actin, myosin, tubulin, DNA) and non-self antigens (myoglobin, spectrin, trinitrobenzene). From the 345 hybridomas secreting immunoglobulin, 68% did not react with any antigen of the panel, 17% reacted with only one, 5.5% with 2 and 7.9% with 3 or more. Apparently, each of the monoclonal multispecific antibodies exhibited a pattern of reactivity which was quite unique. There were no apparent differences in antibody reactivities between self and non-self antigens; moreover, with the exception of DNA, no differences were noted among the 3 different hybridoma collections.

Animals↗

[Relationship between dose of injected serum albumin and appearance of cells synthesizing immunoglobulins or antibodies].

Mice were given increasing doses of sheep serum albumin emulsified with Freund's complete adjuvant in hind footpads. The kinetics of development of cells which synthesize antibody or immunoglobulins without antibody function was studied in the draining lymph nodes. Results show the relationship between the two cell populations: as the number of antibody synthesizing cells increased, the number of immunoglobulin synthesizing cell increased, and conversely. Between 0.050 and 2.5 mg of antigen, the number of cells of each population increased with the dose injected whereas with 5 and 10 mg, this number is smaller than that obtained with smaller doses and decreased with an increasing dose.

Animals↗

[Antibody producing cells and immunoglobulin synthesizing cells lacking antibody function].

After primary immunization the first immunoglobulin-synthesizing cells which appeared, contained immunoglobulins devoided of antibody function. These cells were progressively replaced by antibody synthesizing cells. The results we reported here demonstrated that except the repartition of Ig between IgM and IgG, there are no differences in the molecular composition and in the rate of biosynthesis between these two populations of cells.

Animals↗

Biology of isolated immunocytes. II. Simultaneous detection of cell surface Ig and theta-antigen by immunoperoxidase staining at the ultrastructural level.

Surface Ig and theta-antigen of lymphocytes collected on the bottom of Beem capsules were stained by an indirect immunoperoxidase technique to compare B and T cells at the ultrastructural level. To distinguish the two markers, the surface Ig was allowed to cap during the incubation with anti-Ig antiserum while the incubation with anti-theta alloantiserum was performed at 4 degrees to obtain a staining in a ring pattern. This method was applied to the study of lymphocytes from spleen, lymph nodes and thymus of normal mice. Lymphocytes forming rosettes with SRBC were also studied at different days after immunization with SRBC.

Animals↗

Development of immuneeoglobulin and antibody-forming cells in different stages of the immun response.

After primary immunization of mice, rats and rabbits with antigens (horse radish peroxidase, bovine serum albumin and muchroom tyrosinase) emulsified in complete or incomplete Freund's adjuvant, both cells synthesizing immunoglobulin without detectable antibody function and antibody-producing cells were detected. The first cells which appeared were synthesizing and secreting IgG and IgM immunoglobulins without antibody function. These cells were progressively replaced by cells synthesizing and secreting antibodies. In some plasma cells of mice, rats and rabbits immunized with peroxidase, antibody activity was detected only in restricted areas of the cytoplasm ; the remainder contained antigenic determinants of immunoglobulins. After secondary immunization the results were the following: in mice, both cells containing immunoglobulins without antibody function and antibody-containing cells appeared simultaneously and they were present in equal amount; in rats, only the antibody-containing cells were present in high number. Immunizations performed using different protein antigens (horse radish peroxidase, human and bovine serum albumin, aggregated and desaggregated human IgG, and ovalbumin) injected as a solution in saline have shown that after antigenic stimulation both populations of cells appeared, their number depending on the dose of the antigen injected. Further experiments carried out with tolerant mice, with germ-free animals and with "B" mice have shown that the appearance or not of antibody-producing cells was always related with respectively the presence or absence of cells synthesizing immunoglobulins without detectable antibody function. Finally experiments performed on rabbits have shown that some cells containing immunoglobulins without antibody function share idiotypic determinants in common with cells synthesizing antibodies.

Animals↗

[An immunoenzyme technic for demonstrating the molecular hybridization of nucleic acids].

An immunoenzymatic procedure has been developed based on the use of monoclonal antibodies specific for 5-bromodeoxyuridine (BdUr). It allows the detection of BdUr-labelled DNA immobilized on a nitrocellulose filter. Using this procedure, it was possible to detect up to 0.5 pg of mammalian DNA labelled in vivo with BdUr, 5 pg of nick-translated BdUr-labelled PBR-322 and, using this latter probe and dot-blot hybridization, 50 pg of native unlabelled PBR-322.

Animals↗

A new method using p-benzoquinone for coupling antigens and antibodies to marker substances.

A method using p-benzoquinone for coupling antigens and antibodies to enzymes and erythrocytes is described. The method involves the treatment of proteins (or polysaccharides) at pH 6 or 7 with an excess of p-benzoquinone. After removal of the unreacted reagent by gel filtration, the "activated" proteins were coupled at pH 8-9 with enzymes or erythrocytes. Biological activities of the proteins were not substantially modified by this treatment since 80-100% of the antigen binding capacity was found to be preserved in p-benzoquinone treated antibodies or Fab fragments. Anti-Ig antibodies (or Fab) were coupled by this procedure to peroxidase, alkaline phosphatase, lactoperoxidase, glucose oxidase and beta-galactosidase, and the conjugates obtained were found to be highly effective in detecting intracellular Ig by immunohistochemical techniques. Erythrocytes coated with sheep anti-mouse Ig antibody or Fab were used to titrate by passive hemagglutination serum Ig. The same erythrocytes were employed to detect by plaque assay mouse Ig secreting cells. Erythrocytes coated with peroxidase, alkaline phosphatase, bovine serum albumin, ribonuclease, Salmonella polysaccharide (B 27 +) and pneumoccocal polysaccharide SIII were employed to titrate serum antibody by passive hemagglutination and hemolysis and to detect mouse antibody secreting cells by plaque assay. All the antigens and antibodies coated erythrocytes prepared gave highly satisfactory and reproducible results.

Alkaline Phosphatase↗