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Biomedical subjects

E Appella

Publications and source records attributed to E Appella.

At least 361 records · Page 20Linked to original sources
PubMed1976

Cell-mediated immunity in mice against papain-solubilized histocompatibility and tumor-specific antigens by a macrophage migration inhibition microassay.

The cell-mediated immune status of B10.D2 (H-2d) mice immunized with spleen cells from a congenic strain, B10.A (H-2a), differing at the H-2 locus and of BALB/c mice immunized with a syngeneic simian virus 40 (SV40)-induced sarcoma (mKSA-TU5) was evaluated by an agarose microassay for migration inhibition factor. The inducing antigens in this experiment were papain-solubilized and partially purified chromatographic preparations of spleen cells from A/J mice (H-2a) and a papain-solubilized antigen extract prepared from a tissue culture-adapted cell line (TU-5), derived from the SV40-induced mKSA tumor. The assay used microliters of normal or immune peritoneal exudate cells (PEC) resuspended in a 2-mul droplet of agarose and cultured in the presence or absence of antigen. Specific migration inhibition of PEC from immunized mice was observed with concentrations of solubilized antigen preparations as low as 2.0 mug/ml (3.67 mug/chamber).

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PubMed1976

The component fragments obtained by acid dissociation of papain-solubilized H-2 molecules.

H-2Kk and H-2Dd molecules were specificially purified from a radioiodinated H-2a preparation obtained by papain digestion of spleen cell membranes of A/J strain mice. The molecules were isolated by binding to H-2 alloantisera of the corresponding private specificity followed by precipitation with rabbit anti-mouse IgG antiserum. The specifically precipitated radioiodinated H-2Kk and H-2Dd molecules were dissociated by acid treatment into large and small components of about 37,000 and 11,000 respectively. These were separated by gel filtration at acid pH or by gel isoelectric focusing in the presence of 6 M urea. Each component separated by gel filtration of the acid-dissociated H-2 molecules showed a high degree of size homogeneity as determined by sodium dodecyl sulphate-acrylamide gel electrophoresis. Upon gel isoelectric focusing, however, the small components showed two peaks of radioactivity closely located together at pH 7-8, both of which had a restricted pH range, while the large components gave one peak of a relatively wide pH range of pH 5-6. The H-2Kk and H-2Dd molecules gave essentially the same pattern in terms of the numbers and the positions of the radioactivity bands. Under the iodination conditions used the large components of H-2Kk molecules contained more radioactivity than the small components, while the reverse was true in case of H-2Dd molecules. Such a difference was also found with H-2Kk and H-2Dd molecules isolated by use of alloantisera of the respective public specificity. The assay of binding of the isolated components with H-2 alloantisera of defined specificity revealed that the large components retain most of the allospecificities of the parental H-2 molecules. No H-2 allospecificities were found on the small components. The small components showed extensive binding with rabbit antiserum against mouse beta2-microglobulin. The same antiserum did not show any binding with the large components. On the other hand, both of the components did bind with rabbit antiserum against papain-solubilized H-2 molecules.

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PubMed1976

Biological and biochemical properties of soluble tumor-specific transplantation antigen of a simian virus 40-induced neoplasm.

We have solubilized by limited papain digestion and partially purified the tumor rejection antigen, tumor-specific transplantation antigens (TSTA), from membranes of a simian virus 40-induced sarcoma. Uniform-sized materials with a molecular weight range of 50,000 have retained their tumor rejection activities through the purification procedures. The simian virus 40 TSTA have been separated from H-2 activity by affinity chromatography on concanavalin A columns and no evidence was found for H-2 antigens in the unbound fraction (I) of concanavalin A containing TSTA activity. A reduced yield from the crude soluble fraction was observed with Fraction I of concanavalin A material and this may indeed represent fragmentation of antigen during papain digestion. These results stand in contrast to purification of histocompatibility antigens (H-2alpha) using the same methods and techniques. Low concentrations of simian virus 40 TSTA crude soluble materials were nervertheless biologically active. A concentration as low as 4 mug protein provided 50% tumor rejection and 0.1 mug protein provided lymphocyte stimulation. Both assays reflected specificity of response.

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PubMed1975

Cellular immune reactivity in vitro and tumor rejection provided by tumor-associated antigens of friend-virus-induced leukemia.

Cell-mediated immunity (CMI) and tumor rejection were studied in the Friend virus leukemia system of C57Bl/6 mice. Mice were immunized with Friend leukemia virus (FLV) or X-irradiated FBL-3 leukemic cells and studied temporally for the development of CMI reactivity by assays of 51Cr release lymphocyte cytotoxicity, lymphocyte transformation, migration inhibition, Winn tumor cell neutralization and transplantation rejection. High levels of specific lymphocyte cytotoxicity were observed by day 7 f0llowing FLV infection; this reactivity reached a peak between 17 and 21 days, and returned to background levels by day 36. Further, positive Winn assays were obtained with spleen cells from mice immunized with FLV at times when the mice resisted live FBL-3 tumor challenge. Positive lymphocyte transformation was obtained with spleen cells from mice immunized with FLV or FBL-3, but not with cells from normal mice or mice immune to a syngeneic methycholanthrene-induced tumor, when cultured with papain-soluble FBL-3 or RBL-5 tumor-cell extracts or mitomycin-C (MMC)-treated FBL-3 or RBL-5 cells. Positive reactivity in the lymphocyte transformation assay occurred after reactivity had peaked in the lymphocyte cytotoxicity test. Similar positive macrophage migration inhibition patterns were also obtained with peritoneal exudate cells (PEC) from FLV-immunized mice using papain-solubilized tumor-associated antigen (TAA) from FBL-3 cells. These data suggest that sequential development and modulation of CMI reactivity occurs as observed in different assays following immunization in this system.

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PubMed1975

Studies of lymphocyte stimulation by intact tumor-cell and solubilized tumor antigen.

BALB/c mice were rendered immune to syngeneic SV40-induced sarcoma by subcutaneous injection of mKSA-TU5 tissue-culture adapted cells. Spleen cells from immune mice were examined for tumor-cell neutralization in the Winn assay as well as in in vitro lymphocyte stimulation assays. A microculture (200 mul) lymphocyte stimulation (LS) assay utilizing immune spleen cells was employed with mixed lymphocyte/tumor-cell cultures (MLTC) and the papain crude soluble (CS) extracts from mKSA-TU5 cells. Specificity in the LS assay was determined using spleen cells from mice immune to other syngeneic tumors and by soluble antigenic preparation of normal BALB/c spleen cells. The Winn assay studies demonstrated that spleen cells from mKSA-sensitized mice neutralized mKSA tumor cells and this was corroborated by their resistance to direct tumor challenge. Positive lymphocyte transformation responses in MLTC were observed when mKSA-TU5-sensitized spleen cells were mixed with mitocycin-C-treated intact tumor cells or when papain-solubilized antigens of mKSA cells were employed, but not with non-immune spleen cells or with a soluble antigen from normal cells. Papain-solubilized antigen preparations employed in in vitro assays also immunized against challenge with mKSA tumor cells. Specificity of these lymphocyte transformation reactions was demonstrated with non-sensitized lymphoid cells or lymphoid cells from mice sensitized with a syngeneic Kirsten virus-induced respond. Thus, mKSA tumor surface antigens were recognized on intact tumor cells or with microgram quantities of papain-solubilized extracts from these tumor cells. We believe the lymphocyte stimulation assay affords a method for demonstrating the presence of tumor-specific antigen and for monitoring further purification procedures.

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PubMed1975

Distinction between tumor-specific transplantation antigen and virion antigens in solubilized products from membranes of virus-induced leukemic cells.

A membrane antigen from RBL-5 leukemic cells that was solubilized and partially purified is further characterized in this study. This soluble antigen is capable of immunizating syngeneic hosts to reject neoplastic cells and thus resembles TSTA. It also induces cytotoxic antibody in syngeneic hosts capable of specifically lysing RBL-5 cells in vitro. RBL-5, however, releases infectious virus (RLV); it was necessary therefore to rule out virus or structural virion antigens as the effective immunogen. Infectious virus was not detectable in our initial crude membrane (CM) material, nor in the papain-solubilized CS or the G-150 Sephadex-chromatographed fraction. Virus-neutralizing antibody was not detected, under stringent assay conditions, in the syngeneic anti-CM sera. Antigen preparations CM, CS and the chromatographeal fractions F1, F2 and F3 were assayed in a complement-fixation test against brood-reacting antisera capable of detecting virus envelope antigen and gs antigen and against syngeneic antisera. Although our antigen preparations were positive for virion antigens, CS and F2 contained an antigen that reacted only with syngeneic antiserum. These same fractions were those reactive as immunogens. On the basis of these data, it is postulated that a cellular membrane component, other than viral, functions as TSTA.

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