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Antibody to galactocerebroside can be produced in rabbits by the injection of the hapten together with a carrier protein (GC.BSA) or by immunization with the myelin fraction of brain homogenates (GC.M). The anti-galactocerebroside activity of both sera is mainly directed towards the galactose moiety of the galactolipid. In a complement fixation system they react with galactocerabroside (galactosylceramide) and monogalactosyldiglyceride, but not with glucocerebroside (glucosylceramide). Passive cutaneous anaphylaxis can be elicited by the carrier, but not the hapten. There is no demonstrable reaction to the glycolipid itself-even when tested in association with auxiliary lipids or the carrier protein; or as a component of solubilized myelin. The specificity of antibody to galactocerebroside appears to be unaffected by the nature of the carrier or associated proteins. This is shown by the fact that antisera to GC.BSA and GC.M react in an identical fashion with the lipid-protein complexes formed when myelin is solubilized with lysolecithin. Complement fixation is increased by solubilization, but increasing the lysolecithin/myelin ratio above the minimum necessary to render myelin completely soluble decreased the specific activity. Following isopycnic centrifugation maximum complement fixing activity was present in a fraction having a density of 1.04 at 10°, corresponding to one of three peaks of galactocerebroside concentration. However, precipitin reactions were obtained over a range of fractions of higher density. These results suggest that galactocerebroside is present in myelin in at least two forms and provides further confirmation of the fact that this hapten is responsible for a large part of the organ-specific complement fixing and precipitating activity of anti-myelin serum. [ABSTRACT FROM AUTHOR]

The primary and secondary immune responses of toads (Bufo marinus) were studied using the bacteriophage f2 as antigen. Antibodies were identified in the serum 2 weeks after primary immunization and peak antibody levels were reached at 6 weeks. Although both IgM and IgG antibodies can be found in toad serum, most of the antibody activity in our animals persisted in the IgM fraction until 8 weeks after immunization. When a second injection of antigen was given 4 weeks after primary immunization, there was a marked increase in the total serum antibody activity and IgG antibodies were found as early as 4 weeks after the second injection of antigen. These observations suggest that the toad, B. marinus, is capable of a true secondary response to the bacteriophage f2. [ABSTRACT FROM AUTHOR]

The antigen stimulation of thymidine uptake by immune rabbit lymph node cells in vitro was studied. The kinetics of the response varied depending on the time between immunization and culture. Cultures set up early after immunization showed a peak response over day 1-2 of culture while those set up late after immunization showed a peak response over day 3-4. Studies using the metabolic inhibitor BUDR suggested that this is due at least in part to a larger recruitment of cells into the response during the third day of culture when lymph node cells taken late after immunization were used. Removal of antigen from cultures after brief exposure of cells at 4° reduced the magnitude but did not eliminate the proliferative response, suggesting that some antigen is specifically bound to the cells in the cold. Readdition of antigen restored normal reactivity. Holding cells at 4° for 4 hours without antigen had no effect on their response to subsequent addition of antigen. However, if cells were held at 4° for 3 hours with antigen present a severe degree of depression of subsequent thymidine incorporation was observed in some but not all experiments. This depression of responsiveness was interpreted as an in vitro phenomenon comparable to immunologic tolerance. [ABSTRACT FROM AUTHOR]

The immunogenic capacity of protein antigens has been compared for the free and peritoneal exudate cell (PEC)-bound forms. The response of OBA mice to BSA provides the reference system, but lysozyme, ovalbumin, HSA and modified BSA were also studied. Uptake is approximately equally efficient in vivo and in vitro. PEG-bound antigen, estimated by radioactivity is far more potent than the free form in inducing primary immunization. The following properties were also found: (i) viable PEG are required; (ii) irradiation of the cell donor 2–7 days before giving antigen inhibits immunization, but irradiation after uptake does not do so; (iii) mice are susceptible to immunization during their phase of recovery from paralysis; (iv) PEG do not retain large amounts of antigen for long; (v) the activity does not depend solely on a minor, phagocytosis- prone fraction of the antigen; (vi) allogeneic transfer of PEG reduces their immunogenic capacity; (vii) paralysed hosts are not susceptible to immunization, but PEG from paralysed donors are effective; and (viii) the enhancement of immunogenic capacity does not apply to the secondary response. The conclusion may be drawn that the retention of small quantities of antigen by macrophages plays an essential role in some, but probably not all types of immune response.

The persistence of immunogenicity of lipopolysaceharide antigens of E. coli and of sheep red blood cells in the mouse was studied by a transfer system. Immunized mice were lethally irradiated and re-populated with non-immune syngeneic lymphoid and bone marrow cells at different times after immunization. Persistence of immunogenicity in the irradiated hosts was revealed as the induction of a primary immune response in the transferred cells. In this system SRBC persisted in an immunogenic form for 14 days whereas the bacterial antigens remained immunogenic for at least 45 days.

The phenomenon of antigenic competition has been investigated by the use of synthetic compounds and guinea-pigs of inbred strain 2 as experimental animals. The effect of addition to the immunizing mixture of immunogenic and non-immunogenic synthetic compounds has been studied. Antigenic competition has been demonstrated between the synthetic antigen 252,p(Tyr,Glu,Lys) and DNP-p(Lys) or 28,pDLAla-p(Tyr, Glu)--pLys. The non-immunogenic compounds 251,p(DTyr,DGlu,DLys), 33,pTyr-pDLAla--pLys, and p(Tyr) have been found respectively to inhibit, enhance, or not affect the immune response to 252,p(Tyr,Glu,Lys). Two weak antigens, namely 509,p(Tyr, Glu,)-pDLAla--pLys and p(Lys) also enhanced the response to 252,p(Tyr, Glu, Lys). Possible explanations for these findings are discussed.

Immunization of HSA-tolerant animals with sulphanil-HSA induces the formation of antibodies that react with native HSA. To test whether the anti-HSA response does actually represent complete restoration of the original reactivity, the antibodies produced following immunization of tolerant animals with sulphanil-HSAS were compared to those produced by normal rabbits immunized with the native antigen. Gel-diffusion analysis of the precipitins disclosed that the patterns of the two types of antibody were not completely identical. The antibodies of the 'restored' tolerant rabbits were directed mainly towards an antigenic determinant of the native HSA, to which normal rabbits immunized with HSA did not respond. When the 'restored' rabbits were challenged with native HSA after a prolonged rest period, the antibodies formed thereafter were completely identical with anti-HSA produced by normal HSA-immunized rabbits. Thus, complete restoration of the original immunological reactivity to HSA was achieved following an intermediary stage of atypical anti-HSA elicited by the conjugated protein. Attempts to break down natural tolerance to RSA by treatment with sulphanil-RSA failed to give any evidence of an autoimmune elimination of RSA. To test whether this inability to break tolerance to RSA was due to the presence of an excess of native protein, animals that had been made tolerant to HSA were immunized with a mixture of sulphanil-HSA and native HSA. Under such conditions, the termination of acquired tolerance was inhibited. The possible relevance of these observations to the cellular basis of immunological tolerance is discussed. [ABSTRACT FROM AUTHOR]

Mice were immunized with BSA or HSA in Freund's adjuvant, and their lymph node and spleen cells transplanted into syngeneic hosts, which in most experiments had been irradiated. After transplantation the cells do not synthesize much antibody if left without stimulation, but can be stimulated to do so by injection of BSA or HSA in solution. The response has been studied over a dose range of 10-3–105 μg. antigen. Stimulation can be detected down to 10-3 μg. antigen, and reaches a maximum at middling doses. Middling doses stimulate proliferation of the primed cells to an extent which can be measured by 131IUdR uptake. At high doses both antibody production and IUdR uptake are inhibited. The conclusion is drawn that high concentrations of antigen can paralyze the immunological reaction of primed cells. [ABSTRACT FROM AUTHOR]

Rats, injected with crude bovine heart extracts, showed dissociation of their immunological response. Complement-fixing and capillary precipitating anti-bodies were produced, active against particulate antigens, while tanned-cell haemagglutinating and gel-diffusing antibodies were active against soluble antigens. Particulate antigens were centrifuged down from tissue extracts as 60,000 g but not at 12,000 g. The kinetics of production and specificity differed in the two groups of antibody. When rats were injected with bovine tissues from birth, to produce immuno-tolerance, they did not produce complement-fixing or capillary precipitating antibodies on later challenge with bovine heart but did produce haemagglutinating and gel-diffusing antibodies. Even before challenge, pretrated rats showed haemagglutinating antibodies, while the response by this test to the further immunization was partially suppressed. [ABSTRACT FROM AUTHOR]

Studies were continued on the mode of action of a methanol-insoluble moiety (methanol-extraction residue) of attenuated tubercle bacilli which enhances the immune response. We analysed the effects of treatment with this material on the primary and secondary immune responses of mice to two antigens, sheep red blood cell and T2 phage. Pretreatment with methanol-extraction residue enhanced the primary response to sheep red blood cells, accelerating the formation of both 19S and 7S plaque-forming cells and of circulating haemagglutinins, and prolonging high levels of 19S and 7S plaque-forming cells. Methanol-extraction residue delayed the secondary response to sheep red blood cells when administered between primary and secondary immunization. Methanol- extraction residue administration subsequent to immunization did not affect an ongoing primary response to this antigen. Administration of methanol-extraction residue before primary immunization with T2 phage, or between primary and secondary immunizations, enhanced both the primary and secondary circulating antibody responses to this antigen. The findings are discussed in relation to possible sites of action of methanol-extraction residue. [ABSTRACT FROM AUTHOR]