Comparison of hemagglutination inhibition and hemagglutinin pseudovirus neutralization titres in relation to protection against influenza in a mouse model

The hemagglutination inhibition (HI) test has long been used as a standard measure of antibody response for inactivated influenza vaccines. However, the HI test has limitations, such as insensitivity when using some H3N2 virus strains and failure to detect neutralizing antibodies that target regions distant from the receptor binding site. We therefore examined a hemagglutinin pseudovirus neutralization (PVN) test as a possible supplement or alternative to the HI test. We evaluated the association of HI or PVN titres with protection against influenza infection in mice based on morbidity (where the illness was defined as 25% body weight loss). We assessed this relationship using dose–response models incorporating HI or PVN titres as a variable. The morbidity was correlated with the pre-exposure titres, and such a correlation was well described by a modified dose–response model. The mathematical modelling suggests that PVN titres consistently show a stronger association with in vivo protection as compared to HI titres in mice. Given our findings, the PVN test warrants further investigation as a tool for evaluating antibody responses to influenza vaccines containing hemagglutinin. The resulting models may also be useful for analyzing human clinical data to identify potentially protective antibody titres against influenza illness.     

The dynamics of antibody formation to a vaccinal strain of Francisella tularensis in different immunization regimens

The use of different schemes of albino mice immunization either by living or by killed preparations of the vaccine strain of Francisella tularensis when obtaining monoclonal antibodies to the tularemia microbe made it possible to reveal definite regularities in the dynamics of antibody formation. The highest titres of antibodies in sera of animals-donors of splenocytes were obtained during the daily (for 3 days) intraperitoneal immunization of mice with living vaccine or with its thrice administration to the spleen thrice with the interval of 10 days. Revaccination against a background of high titres of antibodies decreased their quantity in blood serum of mice, while that against a background of low titres increased them.     

Dengue E Protein Domain III-Based DNA Immunisation Induces Strong Antibody Responses to All Four Viral Serotypes

Dengue virus (DENV) infection is a major emerging disease widely distributed throughout the tropical and subtropical regions of the world affecting several millions of people. Despite constants efforts, no specific treatment or effective vaccine is yet available. Here we show a novel design of a DNA immunisation strategy that resulted in the induction of strong antibody responses with high neutralisation titres in mice against all four viral serotypes. The immunogenic molecule is an engineered version of the domain III (DIII) of the virus E protein fused to the dimerising CH3 domain of the IgG immunoglobulin H chain. The DIII sequences were also codon-optimised for expression in mammalian cells. While DIII alone is very poorly secreted, the codon-optimised fusion protein is rightly expressed, folded and secreted at high levels, thus inducing strong antibody responses. Mice were immunised using gene-gun technology, an efficient way of intradermal delivery of the plasmid DNA, and the vaccine was able to induce neutralising titres against all serotypes. Additionally, all sera showed reactivity to a recombinant DIII version and the recombinant E protein produced and secreted from mammalian cells in a mono-biotinylated form when tested in a conformational ELISA. Sera were also highly reactive to infective viral particles in a virus-capture ELISA and specific for each serotype as revealed by the low cross-reactive and cross-neutralising activities. The serotype specific sera did not induce antibody dependent enhancement of infection (ADE) in non-homologous virus serotypes. A tetravalent immunisation protocol in mice showed induction of neutralising antibodies against all four dengue serotypes as well.     

A comparison of antibody titres in mouse uterine fluid after immunization by several routes, and the effect of the uterus on antibody titres in vaginal fluid

Measurements of specific antibody titres in uterine fluid of mice immunized by different routes indicated that two immunizations in the pelvic presacral space using aluminium hydroxide as adjuvant was a simple and effective way to elicit a significant IgA and IgG response. Higher IgA and IgG titres were produced in uterine fluid by subcutaneous immunization with antigen in Freund's complete adjuvant followed by intravaginal boosting without adjuvant, but this immunization involved both a toxic adjuvant and repeated applications of large doses of antigen in the vagina. Intragastric immunization produced an IgA response in the uterus but no IgG. Local intravaginal priming and boosting with large doses of antigen without adjuvant produced an IgA response in uterine fluid, but was less effective for IgG and was inefficient in terms of time and the amount of antigen used. Hysterectomy reduced the concentration of specific IgA in vaginal fluid of immunized mice to no more than 5% of normal, indicating that most of the IgA in vaginal fluid originates in the uterus. In contrast, IgG titres were not significantly different in hysterectomized and intact mice. IgA titres in vaginal fluid were at least partly restored to normal levels in sham-hysterectomized mice.     

Viral infections in renal allograft recipients treated with long-term immunosuppression.

Thirty-nine renal allograft recipients who had received continuous immunosuppression for six to 13 years were examined clinically and virologically for evidence of past or present viral infection. Twenty-five had common warts, usually on the hands. In most the warts had appeared about one year after transplantation; once present, they never disappeared. Six patients had had a zoster rash from two months to four years after transplantation. None had had jaundice, and there was no change in the frequency of colds or non-specific fibrile illness. Four patients had no cytomegalovirus complement-fixing antibodies throughout the observation period; in the other 35 the antibody titre had risen appreciably during the first three to four months after transplantation. Antibody titres were high (mean 64) at follow-up, being only slightly lower than the highest titres achieved during the immediate postoperative period. None of the patients had had symptomatic cytomegalovirus infection, and in only two was the virus isolated from the urine at follow-up; the titres were extremely low. No changes occurred in the frequency of herpes simplex eruptions. Although all patients had herpes simplex humoral antibody, none excreted the virus. Although cytomegalovirus antibody titres were high, virus excretion was rare, indicating that chronic cytomegalovirus infection in these patients is immunologically well controlled.     

Experimental protective of fraction I of the plague microbe]

The immunogenic properties of the freeze-dried preparation of fraction I tisolated from the culture of avirulent Pasteurella pestis strain were studied in experiments on hamadryas baboons, guinea-pigs and white mice. The preparation was found to have a pronounced protective effect in hamadryas baboons and white mice, especially when the preparation was used in mixture with incomplete Freund adjuvant. In hamadryas baboons resistance to infection was shown to correlate with hemagglutionation antibody titres.     

Infectivity period of mice inoculated with human adenoviruses

Due to non-productive infections, mice are not a good model to study some human adenoviruses. However, mice provide an excellent model to study the metabolic effects of human adenovirus, Ad36. Research interest in Ad36 is increasing rapidly, and consequently an increase in the use of mice as a model is anticipated. However, little is known about the transmission potential of Ad36 from infected mice to other laboratory animals or personnel. While underestimating the infectivity could promote inadvertent spread of Ad36, overstating it could drain valuable laboratory resources and animals. Therefore, we determined the duration of infectivity in female C57BL/6J mice that were experimentally infected with human adenoviruses Ad36 or Ad2. Other uninfected mice were co-housed for one week with the experimentally-infected animals, four or eight weeks postinfection. Additionally, uninfected mice were housed in the cages of mice that were infected with Ad36, 12 weeks earlier. The presence of viral DNA in tissues was used to indicate infection of mice. Although experimentally-infected mice harboured viral DNA at least up to 12 weeks, the horizontal transmission of infection was observed in co-housed mice only up to four weeks postinfection. Thus, Ad36-infected mice should be considered potentially infective for eight weeks and appropriate handling and barrier containment should be used. After eight week postinfection, horizontal transmission appears unlikely. This information may provide guidelines for animal handling, and experimental design using Ad36, which may increase safety for laboratory personnel and reduce the number of mice required for experiments.     

Echinococcus granulosus: induction of T-independent antibody response against protoscolex glycoconjugates in early experimental infection

In this work CD4-knockout mice were used as a model to analyse the role of CD4⁺ T cells in the antibody response against Echinococcus granulosus immunization or experimental infection. Results obtained with mice immunized with protoscolex antigens indicated that these contain T-independent antigens. After infection, CD4-knockout mice and C57Bl/6 mice showed similar titres of specific antibodies indicating that T-independent antibody production was quantitatively important in early infection. We have also identified an antigenic fraction from protoscoleces (E4⁺) which induces CD4 T cell independent antibody response in early stages of infection.In conclusion, the results presented here directly support the existence of T-independent immunogens in E. granulosus protoscoleces and suggest that T-independent antibody response may be quantitatively important in early infection.     

Transmission of human T-cell leukaemia virus type I into adult mice

Human T-cell leukaemia virus type I (HTLV-I)-transformed rabbit T-cells, F647a, were intraperitoneally injected into eight 10-week-old C3H/He and C3H/HeJ mice (1 x 10(7) F647a cells/mouse), respectively. Antibody titres against HTLV-I increased to a peak at 1-3 months after injection in both C3H/He and C3H/HeJ mice. At 12 months after injection, antibody titres of two of the eight C3H/HeJ mice became undetectable, whereas those of all the C3H/He mice still ranged from 1:10 to 1:40. Sera from both seropositive C3H/He and C3H/HeJ mice reacted with HTLV-I core proteins, but not with the env protein. HTLV-I proviral sequences were detected in two of eight C3H/He mice and three of the eight C3H/HeJ mice. These results suggest that HTLV-I is able to infect an adult mouse.     

An inverse relationship between heparin content and antibody response in genetically selected mice. Sex effect and evidence of a polygenic control for skin heparin concentration

The heparin content of genetically selected mice with high and low antibody response to bacterial antigens is reported. An inverse relationship between antibody titres and concentration of heparin was observed for both male and female mice. The lower-antibody-responder line contains twice as much heparin as the higher-responder ones. Furthermore, the female mice also contained twice as much heparin as the male mice. Genetic analysis of the parental and interline hybrids has shown a partial dominance for the character 'heparin content' in favour of the high-heparin phenotype and this character appears to be subjected to polygenic control. The possible biological role of heparin and/or mast cells in the surveillance of the organism against some pathogens is discussed in the light of these and other findings.     

Lymphoid tissue responses to perfluorocarbon emulsion in rats: time course effects relative to immune challenge

1. The effects of either intraperitoneal (i.p.) or intravenous (i.v.) injection of low doses (10 ml/kg) of the commercial emulsion, Fluosol-DA 20% (F-DA), on lymphoid tissues and antibody production against sheep red blood cells (SRBC) have been studied relative to the timing of immunization in rats. 2. Spleen and liver weights were significantly increased in response to injection of F-DA although no consistent pattern was observed. 3. Thymus weight was decreased following F-DA injection in some experimental groups whereas mesenteric lymph node (MLN) weights were unchanged throughout. 4. The mean plasma antibody titre to SRBC was significantly increased in some groups of animals injected with F-DA both before or after immunization; maximum titres were observed following injection of emulsion simultaneously with SRBC. 5. These results show that lymphoid tissue weights and plasma antibody titres in rats immunized with SRBC vary according to the timing and route of a previous or subsequent injection of F-DA.     

Haemagglutination inhibition antibody levels one year after natural measles infection and vaccination.

An assessment of haemagglutination inhibition antibody (HAI) titres of 1,163 children, comprising 739 recipients of live measles vaccines and 424 patients with natural measles infection after 1 year was made in this investigation. Statistical analysis revealed a significant difference in the levels of HAI antibodies. Of the vaccinated children a significant 67.45% showed antibody titres of less than or equal to 1:16, while only 23.48% of children with natural measles showed these antibody titres. The importance and implication of such HAI antibody titres is discussed.     

The SCID/Beige mouse as a model to investigate protection against Yersinia pestis

In this study, we have shown that severe combined immunodeficient/beige mice reconstituted with hyperimmune Balb/c lymphocytes can be used as a model to demonstrate adoptive and passive protection against plague infection. Reconstitution of severe combined immunodeficient/beige mice was successful in nine out of ten mice as demonstrated by spleen colonisation and sustained circulating immunoglobulin titres. Furthermore, an increase in antibody titre was evident after a booster immunisation of reconstituted mice. Presence of circulating antibody correlated with protection against a systemic plague challenge and indicated that in reconstituted mice adoptive transfer of a functional immune system had occurred. The severe combined immunodeficient/beige mouse was also used to demonstrate passive protection against inhaled and systemic plague infection. The reconstituted severe combined immunodeficient/beige mouse model demonstrating protective immunity against plague will be further developed to identify the immune cell subsets responsible for this protection.     

Cell-mediated and humoral immune responses in mice during experimental infection with Mycoplasma pulmonis

Cell-mediated and humoral immune responses in mice after challenge exposure with Mycoplasma pulmonis were investigated. The cell-mediated immune response was determined by means of the delayed-type footpad swelling and the humoral immune response by means of the indirect haemagglutination test. Delayed-type footpad swelling and serum antibody titres were detected at one week after the challenge exposure and persisted for 7 weeks until the end of the experiment. However, there was a poor correlation between the degree of delayed-type footpad swelling and that of serum antibody titre. Delayed-type footpad swelling in mice with gross pneumonic lesions was less than that of mice with no gross lesions. A weak negative linear correlation was observed between the delayed-type footpad swelling and the number of M. pulmonis isolated from lungs.     

Effects of a novel perfluorochemical emulsion on lymphoid tissues and immunocompetence in rats: time course effects relative to immunological challenge

1. The effects of intravenous (i.v.) injection of low doses of a novel perfluorochemical (PFC) emulsion on lymphoid tissues and antibody production against i.v.-injected sheep red blood cells (SRBC) have been studied relative to the timing of immunization in rats. 2. Spleen and, to a lesser extent, liver weights were significantly increased in response to injection of emulsion but no consistent pattern was observed. 3. Thymus weight was consistently decreased following injection of emulsion; in contrast, mesenteric lymph node (MLN) weights were unchanged throughout except for a significant increase in animals injected with emulsion 24 hr prior to immunization. 4. The mean plasma antibody titre to SRBC showed a variable response in animals receiving the emulsion: titres were significantly increased in rats injected with emulsion 1 hr prior to immunization whereas they were significantly reduced in animals injected with emulsion 7 days before SRBC. 5. These results show that lymphoid tissue weights and plasma antibody titres to SRBC vary according to the time of a previous or subsequent injection of PFC emulsion.     

Comparison of serum and lung extracts for surveys of wild animals for antibodies to Francisella tularensis biovar palaearctica

A comparative study of the antibody titer against Francisella tularensis biovar palaearctica in serum and lung extract was made using different immunoassays. Samples were taken from experimentally-infected goshawks (Accipiter gentilis) and European beavers (Castor fiber), and in a field survey of wild beavers. Good accordance between the antibody titer in serum and in lung extract was found in the experimental studies. However, the antibody titer was generally one- to three-fold lower in lung extract than was the titer of serum. Results in the field survey were less reliable. Estimating antibody titer in lung extract is a practical method for surveys for antibody levels, and an alternative to serological surveys, despite lower sensitivity as compared to serum.     

Study of persistence of maternal antibodies to varicella-zoster virus by indirect haemagglutination, with result control by radioimmunoassay

The persistence of maternal antibody against varicella-zoster virus was studied by the method of indirect haemagglutination (IH) and a control of the results was performed by radioimmunoassay (RIA). The findings were analysed with reference to the constant half-life of passively acquired IgG and the titre range and frequency of antibodies in 220 persons of fertile age representative of the normal population. Statistical analysis of expected and the experimentally obtained data was performed. Among the normal population, the expectancy is a gradual decrease in maternal IH antibody titres to negativity during the first 10 months of life and 100% sero-negativity by the 11th and 12th month. The experimental data showed a decrease in IH seropositivity from the initial 100% in 1-month infants to 27% in 6-month and 7% in 12-month infants. Nine out of twelve of the experimentally found positive titres in infants during the second half of their first year of life deviated from the normal population sample. The results obtained by both of the methods used were in good agreement.     

The effect of protein-deprivation on the susceptibility to influenza virus infection: a murine model system.

The effect of a 4% albumin diet initiated at weaning on the susceptibility to influenza virus infection was studied in C57Bl mice. Protein-deprivation was found to enhance markedly the susceptibility to a lethal infection with both mouse-virulent and avirulent strains of virus. Viraemia was observed more frequently in protein-deprived mice, and virus persisted longer in the lungs. The humoral immune response following intranasal infection was depressed, with normal levels of IgG antibody but reduced levels of IgM antibody. No difference was found in the seroconversion frequencies between well-nourished and protein-deprived mice. Pre-immunization did not affect the virus titres in the lungs of protein-deprived mice after challenge with the homologous virus, nor did it prevent the spread of virus to the thymus and brain. The results were discussed in terms of the immunocompetence of the malnourished host and of the potential risk of epidemic influenza in children suffering from severe forms of protein-energy malnutrition.     

High titres of antibody to murine leukaemia virus in lymphocyte (Ly) alloantisera

The radioimmune precipitation (RIP) assay was used to examine the antibody titres against endogenous AKR murine leukaemia virus (MuLV) in a number of antisera to lymphocyte (Ly) alloantigens. The sera from normal donor and unimmunized recipient mice used in raising the alloantisera were also examined for anti-MuLV activity. It was found that all the antisera had high anti-MuLV titres and that in all but one case alloantigen immunization augmented the anti-viral titres. The degree of augmentation did not appear to be related to the anti-MuLV titre in the donor strain sera. Three I-region antisera were also examined for anti-MuLV antibodies and were found to have lower anti-viral titres than the Ly antisera even though immunization to I-region products greatly augmented the anti-viral titre. These results caution against the use of Ly antisera in characterizing the phenotype of lymphoid tumour cells without prior virus absorption.     

High titres of antibody to murine leukaemia virus in lymphocyte (Ly) alloantisera

The radioimmune precipitation (RIP) assay was used to examine the antibody titres against endogenous AKR murine leukaemia virus (MuLV) in a number of antisera to lymphocyte (Ly) alloantigens. The sera from normal donor and unimmunized recipient mice used in raising the alloantisera were also examined for anti-MuLV activity. It was found that all the antisera had high anti-MuLV titres and that in all but one case alloantigen immunization augmented the anti-viral titres. The degree of augmentation did not appear to be related to the anti-MuLV titre in the donor strain sera. Three I-region antisera were also examined for anti-MuLV antibodies and were found to have lower anti-viral titres than the Ly antisera even though immunization to I-region products greatly augmented the anti-viral titre. These results caution against the use of Ly antisera in characterizing the phenotype of lymphoid tumour cells without prior virus absorption.