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PubMed1983

In vitro antibody synthesis in 20 microliters hanging drops: initiation of secondary responses and a simple method of cloning hybridomas.

An in vitro culture system is described in which antibody synthesis is initiated by 10(5) hapten-primed spleen cells in 20 microliters hanging drops. Plaque-forming cell (PFC) responses to thymus dependent and thymus independent antigens were obtained in drop cultures comparable to the micro Mishell--Dutton system. Antigens could be administered in soluble form or on antigen-pulsed peritoneal cells. The hanging drop method also provides a reliable yet simple procedure for cloning hybridoma cells while permitting direct visualization of the number of cells in each drop.

Animals↗

PubMed1994

Serum antibody responses of Texel sheep experimentally infected with Haemonchus contortus.

The primary and secondary serum antibody responses of Texel sheep to infective larvae (L3) and adult worms of Haemonchus contortus were studied. Ten-month-old sheep were infected with 20,000 H contortus L3, treated with ivermectin seven weeks later and, after four weeks, reinfected with 10,000 L3 once a week for six weeks. Faecal egg counts were significantly lower during the secondary infection than during the primary infection, but both infections induced antibody responses, as demonstrated by an enzyme-linked immunosorbent assay (ELISA). The primary antibody response developed rather slowly, but the secondary response developed more rapidly and the IgA responses against L3 antigens and the IgG1 and IgG2 responses against adult antigens were twice those observed during the primary infection. These accelerated and enhanced responses after the reinfection suggest an immunological memory for H contortus antigens.

Animals↗

PubMed1984

Factors influencing susceptibility of LEW rats to Heymann nephritis.

Although most LEW rats develop the proteinuria of Heymann nephritis (HN) within 2 months after immunization with Fx1A, protein excretion of some animals remains normal. We have compared nonproteinuric rats with those that developed HN in order to identify factors that influence susceptibility to immunologically medicated kidney disease. In the primary response to Fx1A, immunofluorescence tests showed that antibrush border titers in serum and immunoglobulin deposition in vivo were similar in all rats. However, complement was detected only in rats with proteinuria. Reimmunization with Fx1A at 30 weeks stimulated anamnestic antibody responses in all rats. Following reimmunization, 60% of nonproteinuric rats developed severe HN with an unusually rapid (1 week) onset. Once again, complement was present only in glomeruli of rats with proteinuria. It appears that titers of antibodies to brush border, measured by immunofluorescence tests, are not an index of the pathogenicity of the immune response to Fx1A. Immunological memory, leading to rapid expression of autoimmune disease upon reexposure to antigen, can be established by a primary immunization that does not produce clinical symptoms. Abnormal urine protein composition may provide a clue to subclinical immunopathology of the kidney.

Animals↗

PubMed1990

Pseudoautoimmunity in normal mice: anti-histone antibodies elicited by immunization versus induction during graft-versus-host reaction.

Native preparations of evolutionarily conserved intracellular macromolecules are generally nonimmunogenic when injected in soluble form. However, vigorous immune responses were observed when common autoantigens such as histones, DNA or Sm antigen, or homologous liver homogenate were noncovalently coupled to latex beads prior to injection into mice. Antibody response to histone beads displayed immunologic memory and required a functional thymus, suggesting that T-helper cells were involved. However, bead-elicited autoantibodies could be distinguished from true autoantibodies in that they reacted with denatured, minor, or foreign components of the preparations or to regions unexposed in the native form of the immunogen. This response contrasted with spontaneously arising autoantibodies accompanying graft-versus-host (GVH) disease in the same strain of mice which preferred native nucleoprotein conformations within nuclei, chromatin, or DNA-histone complexes. Histone beads elicited antihistone antibodies displaying a sustained IgM isotype in contrast to spontaneously arising autoantibodies in GVH disease which were predominantly IgG. These studies demonstrate that immunization with autoantigens does not usually elicit true autoantibodies and suggest that lymphocyte populations responsible for pseudoautoimmune responses are different from autoantibody-producing cells. We speculate that if autoimmunity is driven by particulate forms of in vivo self-materials, additional factors are required for breaking the natural tolerance to native conformations within the immunogen.

Animals↗

PubMed1987

Elevation of serum IgG levels and normalization of T4/T8 ratio after hepatitis in a patient with common variable hypogammaglobulinemia.

Acute hepatitis infection developed in a 47-year-old male patient with common variable hypogammaglobulinemia as a consequence of plasma transfusion therapy. Coincident with increases in serum transaminase activities indicative of acute hepatitis, serum IgG levels continued to rise to 506 mg/dl. When plasma replacement therapy was stopped, a transient decline in IgG level (to 371 mg/dl) was produced, followed by a sharp increase in IgG to 607 mg/dl. During this period, the patient's T4/T8 ratio, which had been inverted (0.89), exhibited significant normalization to 1.57. Nevertheless, the patient failed to produce specific antibody after immunization with a number of defined antigens. The mechanism whereby this presumed non-A, non-B hepatitis augmented endogenous IgG production in this patient remains unknown but may be related to diminished suppressor T cell activity. The patient's inability to produce specific antibody during this period suggests an underlying defect in one or more lymphocyte subsets involved in either helper T cell activity and/or immunologic memory.

Agammaglobulinemia↗

PubMed1984

The humoral immune response in the American cockroach, Periplaneta americana: reactivity to a defined antigen from honeybee venom, phospholipase A2.

Our previous experiments demonstrated that honeybee venom could induce a specific, adaptive humoral immune response in the American cockroach. Since honeybee venom is a complex substance made up of several proteins, a more defined antigen is needed for future characterization studies. One of the components of bee venom, phospholipase A2 (PA2) was found to be highly lethal and immunogenic in the roach. Roaches injected with PA2 generated a specific primary response that developed over a period of time, peaking within 10 days, and then gradually subsiding by the fifth week. Specificity of this response was demonstrated by the fact that immunized animals were protected against the original immunizing PA2, but not to PA2 from a heterologous source. In addition, a secondary response could be induced with PA2, demonstrating the existence of immunologic memory. Thus, we established that PA2 could induce as good, if not better, humoral responsiveness as whole bee venom, and therefore could be utilized as a more defined antigen in studies designed to characterize the inducible humoral factor in the roach.

Animals↗

PubMed1984

Immunogenicity of biotinylated hapten-avidin complexes.

The efficacy of avidin as a carrier for the generation of anti-hapten antibodies was assessed in mice by immunization with complexes of avidin and synthetic peptides containing biotin and an epsilon-dinitrophenyl (DNP) lysine residue. The synthetic haptens were constructed with 0, 1 or 2 6-aminocaproyl groups as spacers between the biotin and DNP-lysine moieties. Complexes without a spacer did not induce anti-DNP antibody responses, while those with two spacers induced stronger responses than those with only one spacer. However, the anti-DNP responses to avidin-biotinylated hapten complexes were considerably weaker than responses to a conventional hapten-protein conjugate (DNP-ovalbumin), and, like "T-independent" antigens, failed to induce significant immunological memory. The distribution of isotypes in the anti-DNP antibodies produced to avidin-biotin-6-aminocaproyl-epsilon-DNP-lysine-alanine and DNP-ovalbumin was similar, but the former antigen induced significantly lower levels of antibody in (CBA/N X BALB/c) F1 male mice with the xid defect than in phenotypically normal female littermates, and also induced significant responses in nu/nu mice, in contrast to DNP-ovalbumin. These findings suggest that there is at least a "T-independent" or "T-efficient" component in the response to avidin-biotin complexes, perhaps due to the tetrameric structure of the molecule. Estimates of the depth of the receptor site for biotin were obtained by using the complexes to competitively inhibit the binding of anti-DNP antibody to plates coated with DNP-protein. The findings were consonant with the data on immunogenicity (capacity to induce anti-DNP antibody responses) and suggested that the receptor site has a depth of 16-26 A.

Alanine↗

PubMed1995

Allograft rejection in cattle with bovine leukocyte adhesion deficiency.

In the present investigation cell-mediated immunity in animals with bovine leukocyte adhesion deficiency (BLAD) was studied by means of skin transplantation experiments. Autograft and allograft behaviour in animals with BLAD was compared with the behaviour of simultaneously transplanted autografts and allografts in healthy controls. Allograft survival time was prolonged in three BLAD cattle (28, 30, and 72 days) compared to six healthy controls (12-14 days). When transplantations were repeated on one animal with BLAD using skin grafts from the same donor, accelerated rejection was observed (allograft survival time decreased from 72 days at primary to 35 days at secondary and to 21 days at tertiary transplantation), suggesting the development of immunological memory. Graft-infiltrating lymphocytes that were obtained from allograft biopsies during the period of rejection, were shown to be from recipient origin (beta 2-integrin negative). Our findings demonstrate that, although prolonged allograft survival is observed in cattle with BLAD, skin allografts are ultimately rejected.

Animals↗

PubMed1991

Pneumococcal conjugate vaccines.

We have prepared conjugates of pneumococcal type 4 polysaccharides (PS4) or oligosaccharides to tetanus toxoid using the carbodiimide method. The use of a spacer, 6-aminohexanoic acid, resulted in higher incorporation of carrier protein. Conjugates contained up to 10% free polysaccharide, but no free protein. In general, polysaccharide conjugates induced higher anti-PS4 IgG antibody titers than oligosaccharide conjugates. Conjugates with the highest amount of incorporated protein were the most immunogenic. The response to conjugated PS4 does show characteristics of a T cell-dependent antibody response, in terms of both isotype distribution and induction of immunological memory. Repeated immunization with high doses of PS4TT conjugate resulted in a virtually negative anti-PS4 IgG response, suggestive of the induction of high dose tolerance.

Animals↗

PubMed1994

Immunological responses from non-exposed donors to malaria antigens: implications for immunity and pathology.

An approach to identification of epitopes suitable for vaccine development has been to locate regions of malaria target antigens that are recognized by individuals with clinical immunity. This has applied to identification of T- and B-cell epitopes. It is now realized, however, that T cells from individuals without prior exposure to malaria can respond to malaria parasites, malaria proteins, and peptides copying protein sequences. Such observations raise questions about which epitopes we should be targeting for vaccine development, but also challenge our understanding of immunological memory. Such responses from non-exposed individuals may also be important in expression of disease symptoms.

Antigens, Protozoan↗

PubMed1989

Is booster injection with hepatitis B vaccine necessary in healthy responders? A study of the immune response.

Loss of protective anti-HBs levels (less than 10 IU/l) was noted in 5 (13%) of 38 well documented healthy responders to hepatitis B vaccine 30 months after completing the initial standard vaccination series. Revaccination with a single booster injection of 20 micrograms hepatitis B vaccine intramuscularly resulted in anti-HBs levels well above those initially obtained, thus confirming considerable immunological memory. Both decline prior to and rise after booster injection were proportional to the anti-HBs level obtained initially. The antibody production after a single booster injection was closely monitored in 13 individuals. A swift response was observed from day 4 onwards in all subjects. Based on passive immunization data and in vitro infection of human hepatocytes, this time delay is likely to permit infection of hepatocytes. Therefore, until further data on longterm follow-up of vaccinated in individuals in whom anti-HBs levels have dropped to less than 10 IU/l reveal compelling evidence to the contrary, booster injections remain mandatory for those individuals at risk.

Adult↗

PubMed1992

Is immunity to malaria really short-lived?

Protection against Plasmodium falciparum malaria is usually considered to be the cumulative product of repeated exposure to parasites, and thus a function of age, in endemic areas. The recent outbreak of malaria in the central highlands of Madagascar gave Philippe Deloron and Claire Chougnet the opportunity to compare the incidence of malaria in children and young adults exposed to malaria for the first time, with that in older adults who spent their childhood in the study area before malaria control was introduced. Protection, as well as immune responses to two major P. falciparum antigens, was not related to age. Individuals older than 40 years were more protected than were younger adults. This increased protection was probably due to immunological memory.

Journal Article↗

PubMed1988

Immunogenicity of a recombinant vaccinia virus expressing envelope a glycoprotein of bovine leukaemia virus.

We constructed a recombinant vaccinia virus (RVV) expressing envelope (env) glycoprotein (gp51) of bovine leukaemia virus (BLV): the expression of gp51 was detected by Western blot of the lysates of rabbit kidney cells infected with the RVV. The rabbits inoculated intradermally with the RVV alone failed to induce detectable anti-gp51 antibodies even 10 weeks after immunization. However, when these animals were boosted with inactivated BLV virion in saline, significant levels of anti-gp51 antibodies were induced as shown both in Western blot and immunodiffusion analyses. In these animals, antibodies against gag product (p24) were not detected. On the other hand, the rabbits inoculated with wild-type vaccinia virus and boosted similarly three times with the BLV virion in saline did not induce detectable anti-gp51 antibodies at all. The present experiment revealed that the RVV possessed the capability to endow immunological memory without inducing apparent anti-gp51 antibody responses, meaning that the RVV activated helper T cells far more strongly than B cells. The applicability of the RVV to vaccine development is discussed.

Animals↗

PubMed1991

Active immunity is seen as a reduction in the cell response to oral live vaccine.

Oral immunization elicits a response of antibody-secreting cells (ASC) in the peripheral blood; these cells are believed to originate in the mucosa and hence give information on the mucosal immune response. We have shown earlier that oral booster immunization is followed by an elevated ASC response reflecting an immunologic memory. In the present study we show that a booster dose of a live bacterial vaccine given at a time of active mucosal immunity elicits a low ASC response only. This is probably because the multiplication of the live vaccine is inhibited in the gut, resulting in a low actual dose of the antigen. This situation may be an example of the protective immunity manifested when an orally immunized person encounters the pathogen in nature, and could be used to assess the protective immunity.

Administration, Oral↗

PubMed1991

Experience in Finland with Haemophilus influenzae type b vaccines.

The importance of Haemophilus influenzae type b (Hib) as the leading cause of bacteraemic infections in children was recognized in the early 1970s in Finland. An efficacy trial with the capsular polysaccharide vaccine demonstrated the efficacy of this first generation vaccine, but only from ages 18-24 months onwards. For this reason, since 1983 new polysaccharide-protein conjugate vaccines (PRP-D, HbOC, and PRP-T) have been extensively tested. These studies have shown that conjugate vaccines are immunogenic in early infancy and are capable of generating a lasting immunological memory. A second Hib vaccine efficacy trial in 1986-1987 showed that the conjugate vaccine (PRP-D) was 90% efficacious after the primary immunization series at 3, 4 and 6 months. After a booster dose at 14-18 months, no failure cases have occurred during the follow-up period. The same level of protection seems to be true also in a subsequent trial, where two Hib conjugate vaccines (PRP-D or HbOC) were given at 4, 6 and 14-18 months. These vaccinations have led to a significant decline in the number of invasive Hib infections in young children.

Antibodies, Bacterial↗

PubMed1993

Protection of mice against respiratory Bordetella pertussis infection by intranasal immunization with P.69 and FHA.

Intranasal immunization of adult female Balb/c mice with the Bordetella pertussis antigens FHA or P.69, greatly enhanced their ability to clear B. pertussis from their lungs following aerosol challenge compared with ovalbumin-immunized controls. Low numbers of lymphocytes secreting antibodies (IgG, IgA and IgM) against the immunizing antigens could be isolated from the lungs of immunized mice. Following aerosol challenge with B. pertussis there was a large increase in the numbers of FHA or P.69-specific antibody-secreting cells in the lungs of mice immunized with these antigens. Intranasal immunization, particularly with FHA, also primed mice to develop a systemic serum anti-pertussis antibody response subsequent to challenge. However, pulmonary clearance of B. pertussis correlated most closely with the local antibody response. A strong anti-FHA response was demonstrated in the lungs of mice that received a booster dose of FHA 9 months after their previous exposure to FHA, demonstrating that long immunological memory can develop in the murine respiratory tract following direct application of pertussis antigens to the respiratory tract mucosa.

Adhesins, Bacterial↗

PubMed1996

Protective immunization against Helicobacter stimulates long-term immunity.

Immunization with an oral vaccine composed of whole cell sonicates of Helicobacter felis plus cholera toxin (CT) can protect mice from H. felis challenge. The aim of this study was to determine whether this protective immunity was long-lived. Mice were given the vaccine then left for up to 15 months before challenge. After 15 months, all mice were still protected from H. felis infection, indicating that oral immunization using bacterial antigens plus CT can stimulate long-term local immunological memory.

Animals↗

PubMed1995

Persistence of anti-HBs antibodies in health care personnel vaccinated with plasma-derived hepatitis B vaccine and response to recombinant DNA HB booster vaccine.

Long-term persistence of specific antibodies after primary immunization against HBV infection has been reported. In this study, we evaluated the persistence of anti-HBs in vaccinees 6 years after primary immunization and the response to a booster dose using a recombinant DNA yeast-derived HB vaccine. An 85.4% seroprotection rate was observed after 6 years with a significantly higher seroprotective rate in those subjects who received four doses of vaccine primary immunization as compared with those who received three doses (93.9% versus 67.2%, p < 0.001). One month after receiving the booster dose, 98.6% of the subjects had an anamnestic type of response. The GMTs were found to decrease progressively with increasing age. The antibody levels after booster dose were higher than those attained at the end of primary immunization and reflected the trend seen before the administration of the booster. These results are consistent with the existence of an effective immunological memory in HB vaccine responders. Subjects who received four doses during primary immunization were better seroprotected and had a higher seroprotection rate after the booster dose.

Adolescent↗