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Biomedical subjects

S Avrameas

Publications and source records attributed to S Avrameas.

At least 163 records · Page 9Linked to original sources
PubMed1976

Cooperative binding of concanavalin A to thymocytes at 4 degrees C and micro-redistribution of concanavalin A receptors.

The mode of binding of 125I-labelled concanavalin A and succinyl-concanavalin A to rat thymocytes at 4 degrees C was investigated. Simultaneously, the free binding sites of the cell-bound lectin molecules were quantified by horseradish peroxidase binding. Concanavalin A showed cooperative binding while succinyl-concanavalin A did not. The number of molecules of concanavalin A bound to the cell surface when it was saturated was twice the number of molecules of succinyl-concanavalin A. We interpret these results as showing that the binding of native concanavalin A to thymocytes at 4 degrees C brings about a cooperative modification of the membrane which leads to appearance of new receptors. Divalent succinyl-concanavalin A has no such effect. Horseradish peroxidase binding to cell-bound lectin was shown to be related to the immobilization of membrane receptors; the more they are immobilized, the more receptor-associated lectin can bind horseradish peroxidase. This allowed us to establish that post-binding events, which we called micro-redistribution, occurred at 4 degrees C when either concanavalin A or succinyl-concanavalin A binds to cells. A cooperative restriction of the micromobility of cell receptors is produced by increasing concentrations of concanavalin A. Succinyl-concanavalin A does not restrict cell receptor mobility at any concentration tested. The results are discussed in terms of cell stimulation and cell agglutination.

Animals↗
PubMed1976

The relationship between storage and secretion of specific antibody by immune lymphoid cells: ultrastructural localization of anti-peroxidase antibodies in plaque-forming cells of the rabbit popliteal lymph node.

The ultrastructure of antibody-forming cells (AFC) has been studied in the lymph node cell population from rabbits locally immunized with horseradish peroxydase (PO) incorporated in complete Freund's adjuvant, and the kinetics of AFC development followed from day 7 to day 18 after one injection of PO. Identification of the AFC was done by local hemolysis assay, using carboxymethyl cellulose solidifying medium and PO-coated sheep erythrocytes. AFC were thereafter transferred by micromanipulation into a Beem capsule, fixed, treated by PO for fine ultrastructure detection of anti-PO antibodies, included, sectioned and studied by electron microscopy. It was found that the AFC were essentially of three categories: lymphocytes, proplasmacytes and plasmacytes, with (+), or without (-), intracellular antibody. The proportion of these categories varies with the time elapsed since the injection of antigen and with the plaque-forming activity of the population: lymphocytes (-) are relatively more numerous (over 30%) at the early stages of immunization (day 7). The number of plasmacytes increases with immunization. If most of them contain intracellular antibody at the early stages (up to day 9), the proportion of plasmacytes (+) decreases markedly afterwards. The cell type distribution is compatible with the idea that the lymphocytes are the precursors of plasmacytes, proplasmacytes being transitional forms, but no direct filiation scheme can actually be deduced from these experiments.

Animals↗
PubMed1976

Detection of anti-TNP antibody-forming cells (AFC) with TNP-enzyme and TNP-Fab anti-enzyme conjugates.

Peroxidase (PO), alkaline phosphatase, and glucose oxidase, as well as Fab anti-PO, were coupled with varying molar ratios of trinitrophenyl (TNP) hapten. These reagents were evaluated for their ability to detect anti-TNP antibodies in the lymph node cells of Balb/c mice immunized with heavily-substituted TNP45 alkaline phosphate, which gave rise only to anti-hapten antibody-forming cells (AFC). The best results were obtained with lightly-substituted TNP-Fab anti-PO plus PO, TNP-alkaline phosphatase, and TNP-glucose oxidase. These reagents gave strong, specific staining of AFC, and negative background staining. Anti-hapten and anti-carrier AFC could be stained in contrasting colors on the same slide, when immunization was performed with lightly-substituted TNP5.9 alkaline phosphatase. Anti-hapten AFC were detected with TNP-Fab anti-PO or TNP-glucose oxidase, and unsubstituted alkaline phosphatase was used to reveal anti-carrier AFC. The number of AFC detected with these reagents was compared with the number of direct and indirect anti-TNP plaque-forming cells (PFC). At three and five weeks after primary immunization, 40 and 70% more AFC than PFC were detected. These methods can be employed alone, to enumerate anti-TNP AFC and, if desired, anti-carrier AFC; they can also be used in parallel with anti-TNP PFC assays, to determine the fractions of AFC that are not actively involved in antibody secretion.

Alkaline Phosphatase↗
PubMed1976

Quantitative ultrastructural autoradiographic studies of iodinated plasma membranes of lymphocytes during segregation and internalization of surface immunoglobulins.

Rat spleen lymphocytes were iodinated (125 I) with lactoperoxidase. Quantitative autoradiographic studies on cells fixed immediately after iodination showed 19-24% of intracytoplasmic grains at 3HD and over from the plasma membrane. Normalization of grain density distribution and comparison of resulting curves with the universal curve of grain scatter of 125 I showed that a significant percentage of intracytoplasmic grains (36%) originates from intracytoplasmic labeled sources rather than from scattering from the heavily labeled plasma membrane. Damaged cells had a threefold grain density than intact cells. Radioactivity counts in sliced polyacrylamide gels of iodinated cells revealed 65-72% of total radioactivity in five peaks of apparent mol wt of 44, 50, 57, 90 and 195 thousand daltons. Segregation and internalization of anti-immunoglobulin-Ig-horseradish peroxidase (HRP) complexes from the iodinated plasma membrane proteins of lymphocytes was studied with quantitative autoradiography (125 I) and peroxidase cytochemistry; 64% of grains at 1.5HD (1,500 A) from the plasma membrane were within the cap zone, and 36% of grains remained outside the capped immunoglobulins; 45-57% of grains internalized together with Fab-anti-Ig-Ig-HRP, and 68% of grains internalized together with anti-Ig-Ig-HRP. These studies indicate that (a) iodination of rat spleen lymphocytes results in a significant internal labeling and that (b) immunoglobulins segregate into caps and internalize together with other iodinated plasma membrane proteins while a significant percentage of iodinated proteins (36%) are excluded from the immunoglobulin caps or internalization sites (32-55%).

Animals↗
PubMed1976

Receptor mobility and its cooperative restriction by ligands.

The functional significance of membrane fluidity and receptor mobility in lymphoid cells has been studied in the recent literature. Although far from clarified, the role of membrane fluidity in achieving control over cell activity is probably important; it allows cooperative interactions over long distances. Here, the emphasis is put on the phenomenon of restriction of receptor mobility by ligands such as Concanavalin A, a phenomenon discovered in recent years using morphological techniques. We discuss in some detail our own approach for studying this phenomenon. This consists of quantitatively measuring the active sites on cell-bound lectin molecules by subsequent fixation of horse-radish peroxidase. This study has shown a cooperative binding of Concanavalin A to cells which corresponds to a modification of the membrane, leading to the recruitment of new receptors. The existence of a post-binding event, that we have called micro-redistribution, has been shown at 4 degrees C, through the use of peroxidase binding to cell-bound lectin. A cooperative restriction of receptor microredistribution is observed when the cooperative recruitment of receptors induced by increasing concentrations of Concanavalin A occurs. Both phenomena were shown to be modulated by drugs such as colchicine and cytochalasin B. The characteristics of this modulation suggest that density and distribution of receptors are dependent upon the state of a multimeric submembrane structure which is still functional at 4 degrees C.

Lectins↗
PubMed1976

Development of immunoglobulin and antibody-synthesizing cells after immunization with different doses of antigen.

The kinetics of development of antibody-synthesizing cells and of cells synthesizing immunoglobulins without detectable antibody function were studied in rats immunized with different doses (0-1, 1, 10, 100 mg) of horse radish peroxidase, bovine serum albumin, human serum albumin, hen ovalbumin, or human IgG, which had been deaggregated or heat-aggregated. Each antigen was injected once or twice as a solution in saline. Antibody and immunoglobulin-producing cells were detected in draining lymph nodes by immunohistochemical staining. In the primary response a few antibody-synthesizing cells were found whatever the dose injected. No increase or some increase was found with the amount of antigen injected, according to the protein used, but with all doses of antigen injected, the population of cells remained small, except with human IgG where a relatively high number of positive cells was detected even after injection of 1 mg of antigen. In the secondary response a few antibody-forming cells were also detected with the lower doses of antigen, but this population increased after boosting with 100 mg of antigen. With human IgG a greater number of positive cells was induced withall the doses tested. A correlation between the number of cells synthesizing immunoglobulins without antibody function and the amount of antigen injected was observed in the primary and secondary responses. The relative size of these two populations varied with the stage of immunity of the animals. In the primary response, the population of cells synthesizing immunoglobulins without antibody function was larger than the population of antibody-forming cells. The same was true in the secondary response, but if after a booster injection the level of antibody-synthesizing cells exceeded that reached in the primary response, the increase of cells synthesizing Ig without antibody function was smaller than the increase in antibody-forming cells. In general the more immunogenic an antigen was, the smaller was the ratio between antibody-forming cells and cells producing immunoglobulin without antibody function.

Animals↗
PubMed1976

A simple immunoenzymatic method for measuring IgE in human sera.

A reliable and relatively simple method for the estimation of serum IgE by single radial immunodiffusion is described. The method requires glucose oxidase-labelled antibodies. The method permits the measurement of IgE concentrations ranging from 20 to 700 i.u./ml. The values obtained in unknown samples were in good agreement with those obtained by radioimmunoassay (correlation coefficient r=0-9557).

Glucose Oxidase↗
PubMed1976

Correlations between immunoglobulin- and antibody-synthesizing cells during primary and secondary immune responses of rats immunized with peroxidase.

The development of cells synthesizing immunoglobulins without detectable antibody activity and of antibody-synthesizing cells was studied during primary and secondary immune responses of rats immunized with horseradish peroxidase. After primary immunization with peroxidase emulsified in Freund's complete or incomplete adjuvant, the first antibody-producing cells appeared 4 days after injection. They were preceded by cells synthesizing IgG and IgM without antibody function, appearing 3 days after giving antigen. The ratio between the latter and the former population of cells regularly decreased during the primary response. Seventy to 100 per cent of cells synthesizing immunoglobulins without antibody activity were induced by the antigen, the remainder being induced by the adjuvant. In both populations, the positive cells were always immature or mature plasmocytes. At various times after primary injection, animals received a booster inoculation of soluble peroxidase or of peroxidase emulsified in Freund's adjuvant. Antibody-producing cells, in early stages of differentiation, appeared between 2 and 3 days after challenge and were not preceded by cells synthesizing immunoglobulins without antibody function. These latter cells were reduced or absent after secondary challenge. Increasing the sensitivity of detection of active sites of antibodies, by using direct methods of staining with fixed or unfixed cells gave no increase of antibody-producing cells.

Animals↗
PubMed1975

Lymphocyte differentiation and antibody synthesis in the secondary immune response of peroxidase stimulated lymph nodes of rat.

Lymphocyte differentiation and specific antibody synthesis were studied in the course of the secondary immune response of horseradish-peroxidase immunized rat popliteal lymph nodes by electron microscopic immunocytochemistry. From primary immunization, mature plasma cells containing specific antibody remained as long as 10 months. Antihorseradish peroxidase antibodies were localized in lymphocytic blast cells on Day 3 after restimulation with antigen. In the subsequent days blast cells were still present, and shifting to mature stages occured. During immunocyte differentiation and maturation vesicles and tubules of the Golgi complex became filled with specific antibody suggesting occasional secretion of antibody via the Golgi apparatus. At very mature cell stages, antibody was accumulated in distended cisternae.

Animals↗
PubMed1975

Presence of "receptors" for growth hormone on membranes of rat thymocytes.

Growth hormone conjugated to Sepharose-4 B was found to mimic the action of the free hormone in the stimulation of the uptake and incorporation of radioactive uridine into RNA in thymocytes in vitro. The integrity of the cellular structure was essential for the stimulating effect of the hormone. The bonding of the immunologically reactive determinants of the hormone on membranes of thymocytes was demonstrated by electron micrographs of preparations incubated sequentially with rabbit antigrowth hormone globulins and monospecific sheep anti-rabbit globulins tagged with horse-radish peroxidase.

Animals↗