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R S Accolla

Publications and source records attributed to R S Accolla.

At least 73 records · Page 4Linked to original sources
PubMed1985

The human Ia-associated invariant chain is synthesized in Ia-negative B cell variants and is not expressed on the cell surface of both Ia-negative and Ia-positive parental cells.

The expression of Ia-associated human Invariant (In) chain glycoproteins was studied in the Raji B cells as well as in their RJ 2.2.5 Ia-negative derived variant cells by using a specific rabbit anti-human In chain antiserum. Two-dimensional gel electrophoresis of immunoprecipitates from either biosynthetically labeled or surface labeled cells were analyzed. In addition, flow microfluorometric analysis of stained cells was performed. The results indicate that the In chain is constitutively produced in the Ia-negative B cell variant. Moreover, it appears that several forms of In chain-related molecules, with different charges and distinct m.w. are equally expressed in Ia-positive and Ia-negative B cells. Finally, no evidence could be obtained that the In molecular family was expressed on the cell surface of Ia-positive Raji and Ia-negative RJ 2.2.5 cells.

Animals↗
PubMed1984

Analysis of the structural heterogeneity and polymorphism of human Ia antigens. Four distinct subsets of molecules are coexpressed in the Ia pool of both DR1,1 homozygous and DR3,W6 heterozygous B cell lines.

Four monoclonal antibodies reacting with distinct human Ia antigenic determinants have been used to demonstrate the coexpression of four distinct subsets, NG1, NG2, H40+-3/4+, and DC1 H40--3/4+, in the Ia pool of DR heterozygous or homozygous B cell lines. By two-dimensional peptide mapping the four subsets within the same Ia pool displayed structurally different beta as well as alpha subunits. The beta chain of the NG1 subset was shown to display considerable structural polymorphism when analyzed in two cell lines with distinct DR but similar DC phenotype, LG2 (DR1,1-DC1) and Raji (DR3, W6-DC1). In contrast, the beta chains of NG2, DC1 H40+-3/4+, and DC1 H40--3/4+ subsets of LG2 cells were shown to be very similar to their homologous Raji cell counterparts, thus indicating a relatively low structural polymorphism. Furthermore, the alpha chains of either one of the four subsets expressed in LG2 cells displayed very high structural similarities to the homologous counterparts in the Raji Ia pool, thus suggesting a relatively low polymorphism for the large Ia subunits described in this study. A striking feature deduced from this study was the selective subunit association of the distinct alpha-beta heterodimers.

Absorption↗
PubMed1983

Human B cell variants immunoselected against a single Ia antigen subset have lost expression of several Ia antigen subsets.

Two monoclonal antibodies, D1-12 and BT 2.2, recognizing two distinct subsets of human Ia molecules, NG1 and NG2, respectively, present in all individuals irrespective of their HLA-DR phenotype, have been used to immunoselect cell variants from the lymphoblastoid cell line Raji. Results showed that, irrespective of the monoclonal antibody used for immunoselection, the cell variants analyzed in this study had lost the expression of both D1-12-and BT 2.2-specific antigenic determinants. Moreover, the expression of antigenic determinants specific for a third family of Ia molecules, the DC-1 subset, were also lost in the cell variants. In contrast, expression of HLA A, B, and C common structures, as recognized by the W6.32 monoclonal antibody, as well as expression of surface immunoglobulins, were not affected. Possible mechanisms inducing such a coordinate loss of expression of several families of human Ia molecules are discussed.

Animals↗
PubMed1983

Distinct HLA-DR epitopes and distinct families of HLA-Dr molecules defined by 15 monoclonal antibodies (mAb) either anti-DR or allo-anti-Iak cross-reacting with human DR molecule. I. Cross-inhibition studies of mAb cell surface fixation and differential binding of mAb to detergent-solubilized HLA molecules immobilized to a solid phase by a first mAb.

A series of HLA-DR-reactive mouse anti-human B cell or anti-Ia monoclonal antibodies (mAb) have been used to explore the serological complexity of human class II antigens at the determinant level, using two techniques: (a) cross antibody-binding competitor assays using 125I-labeled and-unlabeled mAb were performed to study the topological organization of the corresponding determinants to determine epitopic clusters recognized by this collection of mAb and (b) differential reactivity of mAb to detergent-solobilized solid-phase-immobilized HLA-DR molecules to determine epitopes expressed on identical DR isotypes. The fifteen mAb could be classified according to the first technique as falling into three different epitopic clusters. Using the second technique, we were able to define at least two independent molecular subsets, one co-expressing two of the three epitopic clusters and the second expressing only the third one. We could not formally identify molecular subsets expressing only one of the first two clusters, using the second technique. The precise serological mapping of the determinants recognized by various anti-class II mAb should prove very useful if such mAb were to be introduced in anti-class II-specific T cell clone blocking experiments. We anticipate that some of them should facilitate the correlation at the clonal level between the T cell repertoire and the epitopes or molecular subsets defined by these mAb. However, within mAb belonging apparently to a same cluster, some could mediate different biological effects.

Animals↗
PubMed1983

The importance of cross-reactions between species: mouse allo-anti-Ia monoclonal antibodies as a powerful tool to define human Ia subsets.

Three mouse allo-anti-Ia monoclonal antibodies recognizing epitopes specific for either I-E gene products (H81.98.21 and H40.315.7 Mabs) or shared between I-E and I-A gene products (H82.246.3 Mab) were used to analyze the human Ia molecules against which they cross-react. The results indicated that in the Ia pool of the human B cell line Raji the three Mabs were able to recognize at least three distinct subsets differing each other both in their alpha as well as in their beta subunits. The comparison with the previously defined human Ia subsets NG1, NGe, and DC1 suggested the following similarities: H81.98.21 specific Ia molecules similar to NG1; H82.246.3 specific Ia molecules similar to NG2, and including probably NG1; H40.315.7 specific molecules similar to DC1 in their alpha subunits and probably heterogeneous in their beta subunits with characteristics of the DC1, NG1, and NG2 subunits altogether. The heterogeneity of the human Ia molecules and the possible similarities with the mouse Ia system were discussed.

Animals↗
PubMed1983

Isolation of cDNA clones for the p33 invariant chain associated with HLA-DR antigens.

HLA-DR antigens are polymorphic cell surface glycoproteins involved in the control of the immune response in man. They consist of two subunits, the alpha and the beta chains. In addition, an invariant glycoprotein of Mr 33,000 (DRp33) is associated intracellularly with HLA-DR antigens. A cDNA clone for DRp33, called 33-10, was isolated. Because no amino acid sequence has yet been determined for DRp33 the identification of cDNA clone 33-10 was based on selection of mRNA by hybridization, subsequent translation in a rabbit reticulocyte lysate supplemented with microsomes, and translation in microinjected Xenopus oocytes followed by immunoprecipitation with an anti-DR antiserum. The translation products assembled with DR alpha and beta chains in oocytes coinjected with all three mRNAs. Assembly of DR alpha and beta chains was also observed in the absence of DRp33 mRNA. Furthermore, when compared with DRp33 immunoprecipitated from a human B-cell line, translation products of the hybrid-selected mRNA showed (i) identical migration in two-dimensional gel electrophoresis, (ii) identical apparent molecular weight in the absence of N-linked glycosylation, and (iii) a very similar two-dimensional peptide map. Transcription of the DRp33 gene into a mRNA 1,400 nucleotides long was observed in B cells but was undetectable in T-cell lines and was very low in liver. Thus, DRp33 appears to be coordinately expressed with DR alpha and beta chains. Hybridization to DNA of mouse-human somatic cell hybrids showed that DRp33 is encoded by a gene that is located outside the major histocompatibility complex.

Animals↗
PubMed1983

Structural heterogeneity of the human Ia molecular pool as detected by cross-reacting mouse monoclonal antibodies.

Three monoclonal antibodies (Mab), H39.49.5, H40.242.3, and H40.164.3, produced in an A.TH anti-A.TL system and reacting with determinants expressed either on I-Ak, on I-Ek, or on I-Ak and I-Ek molecules, respectively, have been used to analyze, at the structural level, the molecules of the human Ia pool against which they cross-react. The results indicate that the human Ia pool can be dissected out by the use of the A.TH anti-A.TL Mab in distinct families, which display structural variations not only among each other but also with respect to the previously defined NG1 and NG2 Ia subsets. In two distinct Ia pools analyzed (from Raji cells: HLA-DR 3,W6, and from LG-2 cells: HLA DR 1,1), the three A.TH anti-A.TL cross-reacting antibodies recognized molecules in which the allelic polymorphism was confined to the beta subunits. Moreover, within the same Ia pool, the alpha subunits as well as the beta subunits were shown to be different from each other. These results are discussed in the context of the present knowledge of the heterogeneity of the human Ia pool and of the possible similarity between gene products of the mouse H-2-I region and the human HLA-D counterpart.

Antibodies, Monoclonal↗
PubMed1982

Monoclonal antibodies against carcinoembryonic antigen (CEA) used in a solid-phase enzyme immunoassay. First clinical results.

A solid-phase enzyme immunoassay using both mouse monoclonal and goat polyclonal antibodies against carcinoembryonic antigen (CEA) was developed. The assay detects 0.6 to 1.2 ng of CEA per ml of serum and has 3 incubation steps which can be performed in 1 day. Polystyrene balls coated with polyclonal goat anti-CEA antibodies are first incubated with heat-extracted serum samples. Bound CEA is then detected by addition of mouse monoclonal antibodies, followed by goat IgG anti-mouse IgG1 coupled to alkaline phosphatase. Results with this enzyme immunoassay using monoclonal antibodies (M-EIA) have been compared with those obtained by the conventional inhibition radioimmunoassay (RIA) using goat antiserum. Three hundred and eighty serum samples from 167 patients with malignant or non-malignant diseases and from 134 normal individuals with or without heavy smoking habits were analyzed by the 2 assays. Excellent correlation between the results of the 2 assays was obtained, but the M-EIA, using monoclonal antibodies from a single hybridoma, did not discriminate better than the conventional RIA between CEA produced by different types of carcinoma and between CEA associated with malignant or non-malignant diseases. Follow-up studies of several patients by sequential CEA determinations with the 2 assays showed that the M-EIA was as accurate as the RIA for the detection of tumor recurrences.

Animals↗
PubMed1982

IL-2-mediated T cell proliferation in humans is blocked by a monoclonal antibody directed against monomorphic determinants of HLA-DR antigens.

We have studied the effect on the interleukin (IL-) 2-dependent human T cell growth of two distinct monoclonal antibodies (Mab), D1-12 and 4F2, with specificity for common determinant of human Ia antigens and for a differentiation antigen expressed on all activated T cells, respectively. Strong inhibition of cell growth was found in cultures supplemented with the anti-Ia D1-12 Mab but not in cultures supplemented with 4F2 Mab. These results were obtained when either total mixed leukocyte culture (MLC) T cells or an MLC-derived T cell clone were used as indicator cell systems for IL-2 activity. The inhibition of cell growth appears to be mediated by a direct interaction of D1-12 Mab with the cells and not by a direct inactivation of the growth factor, as addition of the antibody to murine MLC T cells, which do not express the determinant defined by D1-12 Mab, resulted in no inhibition of their proliferation induced by the same source of human IL-2.

Animals↗
PubMed1982

Demonstration at the single-cell level of the existence of distinct clusters of epitopes in two predefined human Ia molecular subsets.

The expression at the cell surface level of two previously defined human Ia antigen subsets, NG1 and NG2, was studied. Two monoclonal hybridoma antibodies, D1-12 (anti-NG1) and D4-22 (anti-NG2), labeled with distinct fluorochromes, were used as probes. The results indicate that NG1 and NG2 molecules are simultaneously expressed on the same cell, and moreover, that they are identifiable as spatially separate structures. In addition, when a series of 6 other anti-Ia monoclonal antibodies previously described were analyzed in a checkerboard test of binding inhibition at the cell surface level, they were found to closely mimic the reactivity pattern of either D1-12 or D4-22 antibody, suggesting that NG1 and NG2 Ia subsets express independent clusters of highly immunogenic epitopes.

Antibodies, Monoclonal↗
PubMed1982

Sandwich enzyme immunoassay using three monoclonal antibodies against different epitopes of carcinoembryonic antigen (CEA).

Purified monoclonal antibodies (Mab) produced by 3 hybridomas and reacting with 3 different epitopes of carcinoembryonic antigen (CEA) were used in a solid phase enzyme immunoassay. Two Mabs were physically adsorbed to polystyrene balls and the third Mab was coupled to alkaline phosphatase using the bifunctional reagent N-succinimidyl-3-(2-pyridyldithio)-propionate. During a first incubation, CEA from heat-extracted serum samples was immunoadsorbed to the antibody coated balls. After washing of the balls, bound CEA was detected by a second incubation with the enzyme coupled Mab. The sensitivity of the assay was 0.6 ng per ml of serum. A total of 196 serum samples from patients with various types of carcinoma, with liver cirrhosis, or from healthy blood donors with or without smoking habits, were tested. The results obtained with the monoclonal enzyme immunoassay (M-EIA) were compared with those obtained with perchloric acid extracts of the same serum samples tested by an inhibition radioimmunoassay using conventional goat anti-CEA antiserum. There was an excellent correlation between the two assays. In particular, the new M-EIA gave good results for the detection of tumor recurrences in the follow-up of colon carcinoma patients. However, despite the use of exclusively monoclonal antibodies the new assay detected a similar percentage of slightly elevated CEA values as the conventional assay in patients with non-malignant disease, suggesting that the CEA associated with non-malignant diseases is immunologically identical to the CEA released by colon carcinoma.

Antibodies, Monoclonal↗
PubMed1982

Isolation of distinct cDNA clones encoding HLA-DR beta chains by use of an expression assay.

cDNA clones encoding different human Ia antigen beta chains were isolated by use of a complementation-expression assay in Xenopus oocytes. The assay was based on two previous findings. First, oocytes injected with mRNA from a human B-cell line express HLA-DR antigen. The three intracellular DR chains are assembled in oocytes and can be immunoprecipitated with anti-DR monoclonal antibodies. Second, we have isolated cDNA clones encoding DR alpha and intermediate chains. In order to identify beta-chain cDNA clones, mRNA was hybrid-selected with pools of cDNA clones, mixed with mRNA for the alpha and intermediate chains, and injected into oocytes. We isolated two distinct clones that could select DR beta-chain mRNA as demonstrated by assembly of the translation product with DR alpha chains and immunoprecipitation with DR-specific monoclonal antibodies. One clone is specific for a beta chain of the DR locus. The other clone, much weaker in its ability to select DR mRNA, encodes another Ia-like beta chain. Full-length cDNA clones corresponding to the DR and Ia-like beta chains were isolated and compared. Cross-hybridization was detectable in the coding regions but not in the 3' untranslated regions. Distinct RNAs homologous to the DR and the Ia-like beta-chain clones were present in B cells but were undetectable in three T-cell lines.

Base Sequence↗
PubMed1981

Antibody-mediated activation of genetically defective Escherichia coli beta-galactosidases by monoclonal antibodies produced by somatic cell hybrids.

Six hybridomas producing monoclonal antibodies against Escherichia coli beta-galactosidase (beta-D-galactoside galoctohydrolase, EC 3.2.1.23) have been derived from two separate somatic cell fusions. Three of these antibodies can activate defective enzymes produced by strains of E. coli carrying Z-gene point mutations. In antigen excess, one monoclonal antibody shows similar enzyme binding and mutant-activating capacity. Characteristically, the former reaction has a 200-fold higher equilibrium constant. These data provide direct evidence that the enzyme-activation reaction is a single-hit event in which one antibody site favors the correct conformation of one active center of the enzyme. Because each "activating" hybridoma is able to activate several but not all point mutant enzymes tested, it appears that the correction of the genetic defect is produced by binding key sites of the protein three-dimensional structure rather than the sites affected by the mutation.

Animals↗
PubMed1981

Distinct forms of both alpha and beta subunits are present in the human Ia molecular pool.

Two distinct subsets of human Ia molecules, called NG1 and NG2, present in all individuals irrespective of their HLA-DR phenotype, which were previously defined by their reactivity with two monoclonal hybridoma antibodies, D1--12 and D4--22, were analyzed by two-dimensional peptide mapping techniques. Results show that, in the Ia molecular pool from a single individual, small beta subunits of the NG1 and NG2 subsets display significant differences from each other. In addition, beta subunits of the same subset from two different allotype Ia molecular pools are also different from each other, thus indicating that NG1 and NG2 subsets carry polymorphic specificities. Moreover, large alpha chains of NG1 and NG2 subsets are different from each other; however, no significant differences are observed in alpha chains of the same subset when different allotype Ia pools are analyzed. The possible genetic implications of these findings are discussed.

Antibodies, Monoclonal↗