Heterologous Serum Proteins Research Articles

Modified rosette inhibition test with mouse lymphocytes for detection of early pregnancy factor in human pregnancy serum

The rosette inhibition test has been modified so that early pregnancy factor (EPF) in human pregnancy serum can be detected with mouse lymphocytes. Interference with the assay, which would result from incubation of lymphocytes with heterologous serum proteins, is prevented by the introduction of an ion exchange chromatography step. This provides a simple and rapid method for separating EPF from interfering serum proteins. Human pregnancy and non-pregnancy sera were assayed for EPF with human lymphocytes and results compared with those obtained with DEAE-Sephacel fractions of the same sera tested with mouse lymphocytes. The 2 systems showed good agreement but the mouse assay gave greater differentiation between positive and negative results. When monoclonal anti-T cell antibodies, Hu Ly-m1 and anti-Ly-1.1, were substituted for anti-lymphocyte sera in the human and mouse assay systems respectively, similar results were obtained. The mouse assay system has several advantages over the human assay, including stability of the anti-mouse lymphocyte serum and the ready availability of mouse lymphocytes. Moreover the modified method may be applied not only to the assay of human EPF, but also to the assay of EPF from other species. This would be of value if several species were being studied in 1 laboratory, or if sufficient quantities of lymphocytes from a particular species were not available.

Augmentation of delayed-type hypersensitivity to serum proteins by vesicular stomatitis virus infection in mice: virus-suppressor cell interactions.

The effects of infection with vesicular stomatitis virus (VSV) on delayed-type hypersensitivity (DTH) to heterologous serum proteins were investigated in mice. DTH was induced by a subcutaneous injection of antigen in complete Freund's adjuvant. Infection with VSV at the time of immunization did not affect the level of DTH elicited 3 wk later. Marked augmentation of DTH was observed only when previously immunized mice were infected with VSV simultaneously with restimulation by soluble antigen; either soluble antigen or the virus infection alone was ineffective. The augmentation was specific to the antigen used for the restimulation; in the mouse previously immunized with both bovine serum albumin (BSA) and human alpha-globulin (HGG), DTH to BSA but not to HGG was augmented by injecting soluble BSA and VSV, and vice versa. These results strongly suggest that cells involved in the suppression of DTH manifestation became susceptible to the virus after specific antigenic restimulation and were then eliminated.

Increase in immunogenicity of a pulmonary squamous-cell carcinoma, propagated in vitro.

The chemically induced, non-immunogenic lung squamous-cell carcinoma (MSC-10) propagated in vitro gradually loses tumorigenicity in immunocompetent hosts with increasing in vitro passage. This was found to be related to an increase in antigenicity, since immunosuppressed hosts (thymectomy plus 600 rads whole body X-irradiation) supported the growth of tumor cells, whereas immunocompetent recipients did not. The antigens involved in rejection are not heterologous serum proteins present in culture media since the cell line grown in isologous serum is also rejected. Immunization with the in vitro tumor line partially protected against the parental in vivo line, therefore the antigens involved must be present on both tumor lines. Inoculation of the cultured cell line into normal or immunosuppressed hosts produced tumors with the same histological characteristics as those of the in vivo tumor line. We concluded that by in vitro culture the weakly antigenic carcinoma becomes more immunogenic and thereby capable of inducing transplantation resistance. The cultured tumor cells retain their antigenic specificity and histologic characteristics while increasing their antigenic potency.

SERUM REACTIONS AN ANALYSIS OF COMMERCIAL ANTIVENOMS AND THE POSSIBLE ROLE OF ANTICOMPLEMENTARY ACTIVITY IN DE‐NOVO REACTIONS TO ANTIVENOMS AND ANTITOXINS

Infusion of concentrated heterologous serum proteins may precipitate severe reactions in humans. Some of these reactions are not due to a preexisting hypersensitivity to the foreign protein, but rather to the strongly anticomplementary activity (ACA) of the infused protein. The latter mechanism could account for the occasional anaphylactoid reaction seen in a patient who has had no known previous exposure to the foreign protein. Similarly, it could account for the patient who fails to react to a preliminary subcutaneous trial dose, but then collapses when he is infused later with the same preparation. Nine commercial antivenoms from nine countries were examined and most were found to have high levels of anticomplementary activity. Considerable variation was found in both the immunochemical properties and the anticomplementary activity of these antivenoms. Antitoxins of equine origin to the toxins of diphtheria, tetanus and gas gangrene were also found to have high anticomplementary activity. Because of this activity these preparations should always be diluted and infused slowly. Lack of reaction to a test dose will never guarantee an uneventful infusion, and thus all infusions of heterologous immunoglobulin should be undertaken with the greatest caution and vigilance.

Immunologic and fine structure evidence of avidly bound host serum proteins in the surface coat of a bloodstream trypanosome.

Intact, washed Trypanosoma lewisi bloodstream forms, isolated from rats, were agglutinated specifically by antisera against rat whole serum, albumin, alpha2-macroglobulin, and IgG. However, trypsinized bloodstream and intact culture forms lacking surface coat were not agglutinated by these antisera. Trypsinized bloodstream forms, incubated in dilute rat or heterologous host serum proteins, were agglutinated with specific antisera. The characteristic surface coat of intact bloodstream forms was absent from trypsinized cells; however, trypsinized and serum-incubated bloodstream forms reacquired a surface coat similar to that of intact cells. Gel-diffusion and immunoelectrophoretic results showed that rat albumin, alpha2-macroglobulin, and IgG were present in the surface coat of bloodstream forms. Results of quantitative rocket immunoelectrophoresis demonstrated that the adsorbed rat serum proteins constituted by weight about 5% of the trypanosome total surface coat protein.

The Role of T Cells in IgG Production; Thymus-Dependent Antigens Induce B Cell Memory in the Absence of T Cells

B cell memory was shown to develop in congenitally athymic (nu/nu) mice after injection with small amounts of thymus-dependent antigens, in particular heterologous serum proteins, such as fown gamma-globulin (FGG) or DNP-bovine-serum albumin (DNP-BSA). Large doses of proteins (10 mg) tended to produce a specific B cell unresponsiveness, although there was still some evidence of B cell priming. The antigen did not have to be in a multivalent form to interact with B cell so as to induce immunologic memory or tolerance. In contrast to the induction of B cell memory, the production of IgG antibody in this system was found to be strongly T cell dependent. Thymus-independent antigens like LPS or POL with pronounced adjuvant effects on IgG production in normal or surgically thymectomized mice, could not replace T cells in allowing an IgG response against thymus-dependent antigens in congenitally athymic mice. However, the action of T cells once activated is likely to be non-antigen-specific, since it was shown that supernatants of antigen-activated-syngeneic T cells stimulated IgG production in cultures of primed B cell populations non-antigen-specifically.

Immunological studies in patients receiving repeated allogeneic anc xenogeneic liver hemoperfusions.

Serological and immunopathological studies were carried out in five patients during and after repeated allogeneic and xenogeneic liver perfusions for hepatic coma. Patients received from 1 to 16 perfusions, each lasting 5–52 hr, using 24 livers obtained from four animal species and two human cadavers. All patients recovered consciousness; three survived for 1, 4, and 11 weeks, and two are alive and well 3 years after treatment. Before, and at intervals after treatment, patients' sera were examined for changes in the titer of preformed species-specific antibodies, cross reacting heterologous antibodies, the presence of heterologous serum proteins and antibody to these proteins, changes in the level of serum immunoglobulins and complement, and the development of antibodies to human transplantation antigens. Sections of the perfused livers and biopsies of patients' own livers and kidneys were examined histologically and by immunofluorescence. All perfusions were accompanied by rapid absorption of preformed heterologous antibody. After porcine liver perfusions, there followed a marked rise in the titer of species-specific antibodies, heterologous cross reacting antibodies, human anti-A and anti-B isoantibodies, and porcine serum protein antibody. Repeated baboon perfusion gave rise to only small elevations in the titer of heteroantibody but was not accompanied by clinical reactions and did not give rise to antibaboon serum protein antibody. Rising titer of cytotoxic antibody to human lymphocytes was observed in one patient who had also received blood and platelet transfusions. Examination of the perfused organs showed acute humoral xenograft rejection in all pig livers, but only mild or absent cell-mediated rejection in the baboon livers and in the human cadaver livers. Immunoglobulin deposition was not seen in patients' own livers or kidneys. These findings should have important implications in human heterotransplantation because they suggest that an accelerated liver homograft rejection may not occur when patients supported by hepatic perfusion subsequently receive orthotopie liver allografts.

Route of antigen administration for tolerance production

Tolerance production is readily induced by inoculation of heterologous serum proteins in mice. The intravenous and intraperitoneal route are generally preferred because they allow rapid antigen equilibrium in the intra and extravascular spaces of the host. This report shows that the administration of particle-free antigen in the foot pad produces a greater number of tolerant animals at all concentrations used. The active participation of regional lymph nodes in tolerance production is postulated.

The enhancement of muscle trichinosis in rabbits after immunization with heterologous serum proteins

The enhancement of muscle trichinosis in rabbits after immunization with heterologous serum proteins. International Journal for Parasitology 4, 193–196. Rabbits were immunized with electrophoretically-purified rat serum albumin, β-globulin or γ-globulin in Freund's Complete Adjuvant and then inoculated with Trichinella spiralis larvae. Muscle parasitism was greatly enhanced in all the treated animals; this, plus the high levels of anti-rat and low levels of antiparasite antibody, indicates that antigenic competition can increase parasite numbers in trichinosis. Antigenic competition may play a role in the establishment of natural infections by T. spiralis by deflecting a protective immunological response by the host.

Radiolabeled Toxin for Studying Binding of Cholera Toxin and Toxoids to Intestinal Mucosal Receptor Sites

SummaryRecent evidence suggests that the mode of action of the cholera toxin involves at least two steps, which include binding to a membrane receptor and stimulation of the membrane-bound enzyme adenyl cyclase. A sensitive in vitro technique for studying toxin binding to tissue receptor sites is described and appears to have considerable potential usefulness in the study of toxin interaction with mucosal tissue. Highly purified 125I-labeled cholera toxin was found to bind specifically to insoluble mucosal homogenates from the small intestines of normal adult guinea pigs. Binding of labeled toxin could be inhibited by nanogram levels of both cholera toxin and choleragenoid (a spontaneously formed toxoid), but not by heterologous rabbit serum proteins. Artificial cholera toxoids prepared by treatment of purified toxin with formaldehyde or glutaraldehyde failed to inhibit radiolabeled toxin binding to mucosal homogenates. The data indicate that cholera toxin binding is highly specific for the toxin and is n...

Isolation of human reagins differing in their affinity for ragweed immunosorbent.

Reaginic antibodies to ragweed pollen which had been adsorbed to cellulose-allergen immunosorbents were eluted in the presence of homologous or heterologous serum protein(s) at a concentration of 3–7 mg/ml with gly-HCl at pH 2.5, or 6 m urea at pH 7.4. These extraneous serum proteins exerted a stabilizing effect on the eluted reagins. Two groups of reagins differing in their affinity for the allergen(s) were sequentially eluted from the immunosorbent with gly-HCl and Nal regardless of the order of addition of these eluting agents. Moreover, specific elution of some reagins was achieved with a low molecular weight, hapten-like preparation of ragweed pollen.

Experimental Trypanosomal Glomerulonephritis: Evidence for Selective Renal Deposits Containing Reactants of the Alternate C Pathway

Abstract Three rhesus monkeys infected with Trypanosoma rhodesiense developed a sustained parasitemia, hypocomplementemia, and glomerulonephritis. In each of the three cases glomerulonephritis was characterized by mild proliferative and exudative changes. Immunofluorescent studies on renal glomeruli were done with antibody to human γG, γM, γA, C3, C4, and properdin. For each antibody a strong cross-reaction developed with the heterologous (monkey) serum protein. Two nephritic animals showed no deposits of Ig in glomeruli while a third had faint deposits of γM. All three animals showed heavy deposits of properdin and C3, in a granular pattern, but no evidence of C4. Glomerular deposits appeared during the period of hypocomplementemia which developed after approximately 8 days of infection. Non-infected animals (two) failed to show evidence for glomerular deposits of any of the proteins. These findings suggest that the glomerulonephritis of experimental trypanosomiasis is related to selective deposits of reactant proteins of the alternate C pathway (properdin system).

Immune Complex Disease in Experimental Animals and Man

Publisher Summary No experimental model has provided greater insight into the mechanism of immune complex disease than the experimental serum sickness. The morphological, immunohistological, and serological features of the laboratory models have provided a basis for understanding the pathogenic mechanisms responsible for human glomerulonephritis, vasculitis, and a variety of systemic connective tissue diseases. The subject of experimental acute and chronic immune complex disease produced by nonliving antigens has received extensive review in this series. This chapter summarizes the data presented in these two reviews and discusses in detail more recent studies. Experiments to study acute immune complex disease (serum sickness) have been performed in rabbits almost exclusively. Experimental chronic immune complex disease has proved to be a most useful model in understanding human glomerulonephritis. When injected daily with heterologous serum protein antigens, rabbits with strong antibody responses develop chronic membranous glomerulonephritis in about 5 weeks.

Studies on the induction of antibody synthesis against sulfanilic acid in rabbits. III. Effect of antibodies directed against new antigenic determinants in homologous protein. The effect in primed or unresponsive rabbits.

The following three results emerged from the present experiments concerning the induction of antibody synthesis against sulfanilic acid: 1) the anti‐Sulf antibody response in rabbits seems to be much more carrier‐dependent when Sulf is coupled to heterologous serum protein compared to homologous (autologous) serum protein; 2) anti‐NAD antibodies (antibodies directed against the new antigenic determinants introduced into the carrier through the coupling with hapten) were able to induce a secondary response to Sulf11RSA in rabbits primarily stimulated with Sulf10BGG; and 3) anti‐NAD antibodies were able to induce an immune response to Sulf11RSA in rabbits made unresponsive to Sulf11RSA by i.v. injections of soluble Sulf11RSA.

Further studies on guinea pig and rabbit cytophilic antibodies directed against human serum albumin. II. In vitro binding properties and the role of cytophilic antibody in plague-formation as measured using the indirect technique of localized hemolysis in gel.

The fixation of guinea pig and rabbit cytophilic antibodies to normal cells of homologous origin was investigated. For both systems, PE cells were found to fix antibody most efficiently. Spleen cells from other animal species were also shown to fix both guinea pig and rabbit cytophilic antibodies; human spleen cells showed the highest levels of fixation. Interference in the fixation of cytophilic antibodies to homologous spleen cells of guinea pig and rabbit origin was effected when absorptions were carried out in the presence of homologous and heterologous normal serum protein; with the guinea pig system, the presence of one ‘cytophilic’ antibody did not lead to levels of interference in antibody fixation of a second cytophilic antibody which were higher than those induced with normal serum proteins. Destruction of antibody receptor site activity by trypsin was not demonstrable for PE cells whereas the receptor site responsible for fixation of rabbit IgG but not IgM cytophilic antibody was shown to be highly susceptible to proteolytic enzyme treatment. Localized hemolysis in gel was produced using normal guinea pig and rabbit cells passively coated with homologous cytophilic antibody. Whereas the cytophilic antibody activity of guinea pig serum was previously shown to involve only the IgG class of immunoglobulins, the present report provides evidence for both IgM and IgG cytophilic antibody production in the rabbit.

Cell-to-cell interaction in the immune response. VI. Contribution of thymus-derived cells and antibody-forming cell precursors to immunological memory.

Collaboration between thymus-derived lymphocytes and nonthymus-derived antibody-forming cell precursors occurs in the primary antibody response of mice to heterologous erythrocytes and serum proteins. The purpose of the experiments reported here was to determine whether collaboration took place in an adoptive secondary antibody response. A chimeric population of lymphocytes was produced by reconstituting neonatally thymectomized CBA mice soon after birth with (CBA x C57BL)F(1) thymus lymphocytes. These mice could be effectively primed to fowl immunoglobulin G (FgammaG) and their thoracic duct lymphocytes adoptively transferred memory responses to irradiated mice. The activity of these cells was impaired markedly by preincubation with CBA anti-C57BL serum and to a lesser extent by anti-theta-serum. Reversal of this deficiency was obtained by adding T cells in the form of thoracic duct cells from normal CBA mice. Cells from FgammaG-primed mice were at least 10 times as effective as cells from normal mice or from CBA mice primed to horse erythrocytes. These results were considered to support the concept that memory resides in the T cell population and that collaboration between T and B cells is necessary for an optimal secondary antibody response. Poor antibody responses were obtained in irradiated mice given mixtures of thoracic duct cells from primed mice and of B cells from unprimed mice (in the form of spleen or thoracic duct cells from thymectomized donors). In contrast to the situation with T cells, the deficiency in the B cell population could not be reversed by adding B cells from unprimed mice. It was considered that memory resides in B cells as well as in T cells and that priming probably entails a change in the B cell population which is fundamentally different from that produced in the T cell population.

The Effect of Carrier Tolerance on the Antibody Response of Guinea Pigs to the Azobenzenearsonate Hapten

Recently several investigators have shown that the immune response of an animal immunized with hapten-carrier conjugates is significantly influenced by the nature of the carrier molecule. Siskind et at. (1) demonstrated in guinea pigs that the carrier to which DNP is coupled influences the amount, type, and functional activity (i.e., affinity) of anti-DNP antibodies, and Fronstin, Sage, and Vazquez (2) have shown that the degree of anti-DNP responsiveness elicited in mice is directly related to the antigenicity of the carrier.The present study examines the antihapten antibody response of guinea pigs immunized with arsanilic acid conjugates of homologous and heterologous serum protein carriers, and compares the effect of natural and acquired tolerance to the carrier on these responses.Materials and Methods. Random bred, adult male albino guinea pigs (300-360 g) were used in these experiments.Reagents. Azobenzenearsonate (ABA) conjugates were made as before (3) by diazoti-zation and coupling in the ratio of...

Immunological Unresponsiveness to Heterologous Serum Albumins in Adult Mice: Effect of Antigen Quality on Tolerogenicity

STUDIES of experimental tissue transplantation have revealed that tolerance to transplantation antigens is more easily achieved across weak histocompatibility barriers than across strong ones1–4. Also, the degree of foreignness between donor and recipient determines the results obtained in clinical transplantations5–7. Taxonomic origin and thus the immunogenicity of red blood cells are important in producing drug-induced immunological unresponsiveness to heterologous erythrocytes in adult mice8,9. In experimental work on tolerance to heterologous serum proteins the degree of dissimilarity between antigens and “self”-constituents has only rarely been considered. A study by Tempelis revealed that in chicks comparatively smaller amounts of an avian than of a mammalian serum protein were needed to induce a state of immunological tolerance. Furthermore, turkey globulin was more effective in producing unresponsiveness than globulin obtained from the goose10. The following hypothesis might thus be put forward: the smaller the phylogenetic disparity between the species from which the antigen is obtained and the experimental animal, the greater will be the tolerogenicity of the antigen in question. I have tested this hypothesis by assessing in adult mice tolerogenic properties of serum albumins obtained from species either closely related to the mouse or from species taxonomically more distant.

The Mechanism of Immunological Paralysis

Publisher Summary This chapter presents the mechanism of immunological paralysis. The paralysis is the result of a central failure of responsiveness brought about by exposure to antigen that prevents potentially competent cells from initiating a specific antibody response. In cellular terms, paralysis results from an adverse effect of antigen directly on lymphocytes, and the recovery from paralysis appears to involve the recruitment of new competent cells, not a recovery of responsiveness by once paralyzed cells. The quantitative factors involved in this antigenic exposure are determined, and these data not only shed light on the process of immunological paralysis but also on those conditions essential to the initiation of an antibody response. The barrier that the homograft reaction imposes to tissue transplantation can be overcome by paralyzing the host with donor antigens. This form of treatment that is already a routine procedure in animals provides the best hope of suppressing a homograft reaction without interfering with other immunological defenses. The chapter focuses on protein antigens, particularly heterologous serum proteins. These are of special interest because of their application in exploring the induction of paralysis in adult animals by low doses. The competent cell that is affected appears to be the lymphocyte. Macrophages in or from paralyzed animals function normally, but lymphocytes display altered reactivity. There is some evidence to suggest that antigen acts directly upon lymphocytes in the induction of paralysis.

In Vivo Behavior of Homologous and Heterologous Thyroglobulin and Induction of Immunologic Unresponsiveness to Heterologous Thyroglobulin

Summary Homologous and heterologous thyroglobulins were observed to have an in vivo behavior, following injection, very similar to that of homologous and heterologous serum proteins. Following injection, thyroglobulin equilibrated between the intra- and extravascular fluid spaces, and persisted in both the blood and lymph with elimination at an exponential rate. A consistent immune response was made to the injected heterologous thyroglobulin but not to the injected homologous thyroglobulin. As with serum proteins, the half-life of heterologous thyroglobulin was usually shorter than the homologous thyroglobulin and varied with the host animal. An acquired immunologic unresponsiveness was induced in rabbits by periodic injections of a heterologous bovine thyroglobulin starting on the day of birth. Thyroglobulin was found to be an excellent antigen for the study of various immunological mechanisms. In addition to the properties listed above, in small doses heterologous thyroglobulins induce a much greater antibody response than comparable heterologous serum proteins.