Biomedical subjects
Publications and source records attributed to A Malley.
A method for the consistent production of rabbit anti-mouse IgE sera is described. Rabbits were immunized intravenously with IgE-anti-normal mouse serum mixtures over 8 to 12 weeks. Ten microliters of the anti-mouse IgE sera completely neutralized all the mouse IgE in the serum (50 microliter) possessing a passive cutaneous anaphylaxis titer of 20,000.
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Rhesus monkeys were immunized with normal human lymphoid cells, cultured lymphoid cells, and chronic leukemic lymphocytes. Antisera were analyzed by cytotoxicity and immunofluorescence techniques to study the antigenic characteristics of human lymphocytes. In an attempt to obtain a reagent specifically reactive with T (thymus-derived) lymphocytes, an antispleen antiserum was absorbed with cellf from five B- (bone marrow-derived) cell lines. After absorption, the antiserum killed 60-75% of peripheral blood lymphocytes and 40-50% of tonsil cells, so that there was a relationship between the percentage of killed cells and the proportion of T lymphocytes. However, when cells after cytotoxic treatment were assayed for rosette formation with sheep erythrocytes (a T-cell marker) 5-20% of viable rosette-forming lymphocytes were found. Therefore, this antiserum was cytotoxic for only 75-90% of T cells. From studies performed with antisera prepared against spleen and B-cell lines, we conclude that lymphoblastoid cells are antigenically different and deficient in comparison to normal B lymphocytes. In addition, cultured B-cell lines appear to be antigenically heterogenous, as shown by the cytotoxic activity remaining in antispleen and anti-B-cell lines sera after absorption with various numbers and types of lymphoid cell lines. After absorption with normal lymphocytes, an antiserum produced against chronic lymphatic leukemia cells had specific activity associated with 12 chronic lymphatic leukemia cells tested. Absorption of the same antiserum with leukemic cells from two patients showed that a certain degree of antigenic heterogeneity also exists among chronic leukemic lymphocytes.
Purified populations of splenic B and T lymphocytes from LAF mice immunized with a crude extract of timothy pollen (WST) responded specifically to pollen antigens in an in vitro lymphocyte transformation system. The peak lymphocyte transformation response occurred 5 days after a secondary immunization and was the result of T-B cell cooperation in vitro. Wtih two purified pollen antigens as in vitro stimulants we were able to define at least two antigen-specific population of B cells and one population of T cells. These results were confirmed by inhibition studies with a monovalent hapten from WST, Antigen D.
An unusually heavy infestation of the tussock moth resulted in a high incidence of symptoms affecting the skin and mucous membranes of those exposed to high concentrations of particulate matter of this insect. Extensive epidemiological studies proved these manifestations to be both toxic and allergic. The authors have given this condition the name, Tussockosis. Clinical and immunoligical studies including in vitro procedures and direct patient testing confirmed the allergenicity, as well as the non-specific irritation of material from this moth.
A low molecular weight allergen fragment present in the pea dialysate fraction was purified by ion-exchange chromatography and gel filtration. The highly purified allergen fragment inhibits both antigen-indiced passive cutaneous anaphlaxix reactions in guinea-pigs sensitized with rabbit anti-pea extract sera and Prausnitz-Küstner reactions in non-allergic volunteers sensitized with the sera of patients sensitive to green peas. Preliminary analysis of the purified allergen fragment indicates that it is a glycoprotein with a molecular weight of 1800 +/- 250.
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The aqueous extract of green peas was separated into 3 fractions (albumin, legumin, and vicilin) by dialysis against distilled water and isoelectric precipitation. The major antigenic and all of the allergenic activity of the pea extract was associated with the albumin fraction. The albumin fraction retains its allergenicity upon heating at 60 degrees C for 30 min or boiling at 100 degrees C for 5 min, but becomes partially inactivated by autoclaving at 120 degrees C for 15 min. The allergenic determinant expressed by the albumin fraction appears to be common to several other members of the legume family. In addition, the pea dialysate fraction was shown to specifically inhibit precipitin and passive cutaneous anaphylaxis (PCA) reactions involving rabbit antipea serum and the pea albumin fraction, and histamine release from passively sensitized monkey lung tissue using the serum of pea-sensitive patients.
Spleen cells from LAF1 mice were stimulated in a microculture system with T and B cell mitogens or antigens of timothy pollen. Only cells from mice immunized with crude timothy pollen extract (WST) or a major antigen of timothy pollen conjugated to Ascaris (antigen B-Ascaris) responded to timothy antigens in vitro. Optimum responses were obtained at 120 to 144 hr of culture with 5 to 10 mug WST per culture and ranged from three to 10 times greater than cell background. No correlations could be found between the optimum antigen concentration or the maximum response and the immune status of the spleen cell donor. Response could be inhibited by a dialyzable fraction of timothy pollen, antigen D, which is a monovalent form of a major antigen of timothy pollen.
Previous studies with timothy pollen extracts demonstrated that a low molecular weight dialyzable fraction, antigen D, possessed the allergenic determinant of the major allergen of timothy pollen (allergen B). Bio-gel P-2 gel-filtration of the dialysate fraction resulted in the isolation of a fragment, antigen D3 with a molecular weight near 1000. The relationship of antigen D3 to the antigenic and allergenic determinants of allergen B was evaluated by passive cutaneous anaphylaxis (PCA) in guinea pigs sensitized with rabbit antitimothy sera and by allergen-induced histamine release from monkey lung tissue passively sensitized with the serum of timothy-sensitive patients. These studies demonstrated that 10 units of antigen D3 gave 1) a 70% inhibition of PCA reactions in guinea pigs challenged with allergen B, and 2) a 73% inhibition of allergen-induced histamine release from monkey lung tissue sensitized with timothy reagin and challenged with the crude pollen extract (WST). Although chemical characterization studies of the antigen D3 fragment are still being carried out it appears that the major components fo this fragment are 1) a flavonoid pigment - quercitin, 2) a disaccharide moiety - cellobiose, and 3) the amino acid threonine linked together in an )O-glycosidic type linkage.
The IgG1 and IgE homocytotropic antibody responses of LAF and C3H mice to timothy pollen antigens are defined. Both mouse strains responded to low doses of crude timothy pollen extract (WST) or a major antigen of timothy pollen coupled to a purified fraction of Ascaris suum (Antigen B-Ascaris). Titers in LAF mice were greater than those in C3H mice. Regardless of the immunogen, antigen B was the major determinant recognized by the homocytotropic antibodies; PCA titers with WST or antigen B for challenge were equivalent and PCA activity could be inhibited by antigen D, a dialyzable fraction of timothy pollen possessing the antigen B determinant in monovalent form. The possible usefulness of antigen D for in vivo and in vitro studies of specific immune suppression of cellular activity is discussed.
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