Effect of passive immunization or maternally transferred immunity on the antibody response to a genetic vaccine to rabies virus

A plasmid vector, termed pSG5rab.gp, expressing the glycoprotein of rabies virus was tested in young adult or neonatal mice in the presence of maternally transferred immunity or passively administered antibodies to rabies virus for induction of an antibody response. Mice born to rabies virus-immune dams developed an impaired antibody response to genetic immunization at 6 weeks of age, as had been previously observed upon vaccination with an inactivated viral vaccine. Similarly, mice passively immunized with hyperimmune serum showed an inhibited B-cell response upon vaccination with the pSG5rab.gp vector, resulting in both cases in vaccine failures upon challenge with a virulent strain of rabies virus. In contrast, the immune responses of mice vaccinated as neonates in the presence of maternal immunity or upon passive immunization to rabies virus with the pSG5rab.gp construct were only marginally affected.     

Characterisation of a novel lyssavirus isolated from Pteropid bats in Australia

A novel lyssavirus isolated from Pteropid bats in Australia (Australian Bat Lyssavirus, ABLV) has been characterised using gene sequence analyses, electron microscopy and a panel of monoclonal antibodies. Electron microscopic examination of Pteropid bat and mouse brain material as well as virus isolated from tissue culture medium, showed the presence of bullet-shaped rhabdovirus particles and structures characteristic of lyssavirus. Analysis using nucleocapsid (N) specific monoclonal antibodies, showed a strong relationship between this new lyssavirus and serotype 1 rabies. The nucleotide sequence of the prototype strain of ABLV was determined from the initiator methionine codon for the nucleocapsid protein (N protein) to the amino terminus of the polymerase gene (L protein), a distance of 5344 nucleotides. Comparisons of the deduced N, phosphoprotein (P), matrix protein (M), and glycoprotein (G) proteins showed that ABLV was more closely related to serotype 1 classic rabies viruses than to other members of the Lyssavirus genus. The percent relatedness of the ABLV proteins when compared to the cognate proteins of PV (Pasteur vaccine strain) rabies was 92, 75, 87 and 75% for the N, P, M and G proteins, respectively. Phylogenetic studies of N protein sequences showed clearly that ABLV is an unrecognised member of the Lyssavirus genus and represents a new genotype, genotype 7.     

Norepinephrine inhibits a toxin resistant Ca2+ current in carotid body glomus cells: evidence for a direct G protein mechanism

The rabies virus glycoprotein molecule (G) can be divided into two parts separated by a flexible hinge: the NH2 half (site II part) containing antigenic site II up to the linear region (amino acids [aa] 253 to 275 encompassing epitope VI [aa 264]) and the COOH half (site III part) containing antigenic site III and the transmembrane and cytoplasmic domains. The structural and immunological roles of each part were investigated by cell transfection and mouse DNA-based immunization with homogeneous and chimeric G genes formed by fusion of the site II part of one genotype (GT) with the site III part of the same or another GT. Various site II-site III combinations between G genes of PV (Pasteur virus strain) rabies (GT1), Mokola (GT3), and EBL1 (European bat lyssavirus 1 [GT5]) viruses were tested. Plasmids pGPV-PV, pGMok-Mok, pGMok-PV, and pGEBL1-PV induced transient expression of correctly transported and folded antigens in neuroblastoma cells and virus-neutralizing antibodies against parental viruses in mice, whereas, pG-PVIII (site III part only) and pGPV-Mok did not. The site III part of PV (GT1) was a strong inducer of T helper cells and was very effective at presenting the site II part of various GTs. Both parts are required for correct folding and transport of chimeric G proteins which have a strong potential value for immunological studies and development of multivalent vaccines. Chimeric plasmid pGEBL1-PV broadens the spectrum of protection against European lyssavirus genotypes (GT1, GT5, and GT6).     

Human diploid cell culture rabies vaccine (HDCV) and purified chick embryo cell culture rabies vaccine (PCECV) both confer protective immunity against infection with the silver-haired bat rabies virus strain (SHBRV)

The demonstration of extensive differences in the antigenic makeups of the silver-haired bat rabies virus (SHBRV) and canine rabies virus (COSRV) strains raised concerns as to whether current licensed rabies vaccines are sufficiently protective against SHBRV. NIH mouse protection test results show that both the human diploid cell culture rabies vaccine (HDCV) and the purified chicken embryo cell rabies vaccine (PCECV) protected against lethal infection with SHBRV as well as the canine rabies strain COSRV. However, in this investigation, the potencies of both vaccines in mice were found to be significantly higher for COSRV than for SHBRV. The in vivo protection data are confirmed by in vitro virus neutralizing antibody (VNA) test results which demonstrate that mice immunized with HDCV or PCECV develop significantly higher VNA titres against COSRV than against SHBRV. In contrast, VNA tests of sera from individuals immunized with HDCV or PCECV showed that humans, as opposed to mice, develop significantly higher VNA titres against SHBRV than against COSRV. These data suggest that HDCV and PCECV will protect humans against infection with the silver-haired but rabies virus strain in addition to canine rabies virus strains.     

Immunization against rabies with plant-derived antigen

We previously demonstrated that recombinant plant virus particles containing a chimeric peptide representing two rabies virus epitopes stimulate virus neutralizing antibody synthesis in immunized mice. We show here that mice immunized intraperitoneally or orally (by gastric intubation or by feeding on virus-infected spinach leaves) with engineered plant virus particles containing rabies antigen mount a local and systemic immune response. After the third dose of antigen, given intraperitoneally, 40% of the mice were protected against challenge infection with a lethal dose of rabies virus. Oral administration of the antigen stimulated serum IgG and IgA synthesis and ameliorated the clinical signs caused by intranasal infection with an attenuated rabies virus strain.     

Diffusing capacity of the lung for carbon monoxide

A phage-display technology was used to produce a single-chain Fv antibody fragment (scFv) from the 30AA5 hybridoma secreting anti-glycoprotein monoclonal antibody (MAb) that neutralizes rabies virus. ScFv was constructed and then cloned for expression as a protein fusion with the g3p minor coat protein of filamentous phage. The display of antibody fragment on the phage surface allows its selection by affinity using an enzyme-linked immunosorbent assay (ELISA); the selected scFv fragment was produced in a soluble form secreted by E. coli. The DNA fragment was sequenced to define the germline gene family and the amino-acid subgroups of the heavy (VH) and light (VL) chain variable regions. The specificity characteristics and neutralization capacity of phage-displayed and soluble scFv fragments were found to be identical to those of the parental 30AA5 MAb directed against antigenic site II of rabies glycoprotein. Phage-display technology allows the production of new antibody molecule forms able to neutralize the rabies virus specifically. The next step could be to engineer and produce multivalent and multispecific neutralizing antibody fragments. A cocktail of multispecific neutralizing antibodies could contain monovalent, bivalent or tetravalent scFv fragments, for passive immunoglobulin therapy.     

Enhanced virus clearance by early inducible lymphocytic choriomeningitis virus-neutralizing antibodies in immunoglobulin-transgenic mice

Following infection of mice with lymphocytic choriomeningitis virus (LCMV), virus-neutralizing antibodies appear late, after 30 to 60 days. Such neutralizing antibodies play an important role in protection against reinfection. To analyze whether a neutralizing antibody response which developed earlier could contribute to LCMV clearance during the acute phase of infection, we generated transgenic mice expressing LCMV-neutralizing antibodies. Transgenic mice expressing the immunoglobulin mu heavy chain of the LCMV-neutralizing monoclonal antibody KL25 (H25 transgenic mice) mounted LCMV-neutralizing immunoglobulin M (IgM) serum titers within 8 days after infection. This early inducible LCMV-neutralizing antibody response significantly improved the host's capacity to clear the infection and did not cause an enhancement of disease after intracerebral (i.c.) LCMV infection. In contrast, mice which had been passively administered LCMV-neutralizing antibodies and transgenic mice exhibiting spontaneous LCMV-neutralizing IgM serum titers (HL25 transgenic mice expressing the immunoglobulin mu heavy and the kappa light chain) showed an enhancement of disease after i.c. LCMV infection. Thus, early-inducible LCMV-neutralizing antibodies can contribute to viral clearance in the acute phase of the infection and do not cause antibody-dependent enhancement of disease.     

Neutralizing antibodies protect against lethal flavivirus challenge but allow for the development of active humoral immunity to a nonstructural virus protein

Antibody-mediated neutralization of viruses has been extensively studied in vitro, but the precise mechanisms that account for antibody-mediated protection against viral infection in vivo still remain largely uncharacterized. The two points under discussion are antibodies conferring sterilizing immunity by neutralizing the virus inoculum or protection against the development of disease without complete inhibition of virus replication. For tick-borne encephalitis virus (TBEV), a flavivirus, transfer of neutralizing antibodies specific for envelope glycoprotein E protected mice from subsequent TBEV challenge. Nevertheless, short-term, low-level virus replication was detected in these mice. Furthermore, mice that were exposed to replicating but not to inactivated virus while passively protected developed active immunity to TBEV rechallenge. Despite the priming of TBEV-specific cytotoxic T cells, adoptive transfer of serum but not of T cells conferred immunity upon naive recipient mice. These transferred sera were not neutralizing and were predominantly specific for NS1, a nonstructural TBEV protein which is expressed in and on infected cells and which is also secreted from these cells. Results of these experiments showed that despite passive protection by neutralizing antibodies, limited virus replication occurs, indicating protection from disease rather than sterilizing immunity. The protective immunity induced by replicating virus is surprisingly not T-cell mediated but is due to antibodies against a nonstructural virus protein absent from the virion.     

Primary influenza A virus infection induces cross-reactive antibodies that enhance uptake of virus into Fc receptor-bearing cells

Sera of young children who had had a primary infection with influenza A virus or were immunized with a live attenuated influenza A virus vaccine were examined for antibody responses that neutralized virus or enhanced uptake of virus into Fc receptor-bearing cells, because antibodies that enhance uptake of influenza virus into Fc receptor-bearing cells have been reported using mouse immune serum and monoclonal antibodies. The neutralizing antibody titers to the homologous infecting virus and to another H1N1 virus isolated several years later were higher after natural infection than after infection with the live attenuated virus. Natural infection and the attenuated vaccine induced antibodies that enhanced uptake of homologous virus and H1N1 virus isolated several years later. These results demonstrate that primary influenza A virus infection results in the induction of infection-enhancing antibodies.     

Nephrologists'' and internal medicine physicians'' expectations of renal dietitians and general clinical dietitians

How well antibodies can protect against disease due to HIV-1 infection remains a pivotal but unresolved issue with important implications for vaccine design and the use of prophylactic antibody to prevent infection after accidental exposure to the virus and to interrupt transmission of virus from mother to child. Strong doubts about the possible utility of antibodies in vivo have been raised because of the relative resistance of primary viruses to antibody neutralization in vitro. Primary viruses are likely to be close to the viruses transmitted during natural infection in humans. Vaccine studies have been of little value in assessing antibody efficacy in vivo because none of the strategies described to date have elicited significant neutralizing antibody responses to primary viruses. Passive immunization studies are similarly hindered by the paucity of reagents able to neutralize primary viruses effectively and a single study has suggested some benefit. Here we describe experiments to explore the ability of passive antibody to protect against primary virus challenge in hu-PBL-SCID mice. In this model, severe combined immunodeficient (SCID) mice are populated with human peripheral blood mononuclear cells (PBMCs) and infected with HIV-1. We find that the potent neutralizing human monoclonal antibody IgG1b12 at high dose is able to completely protect even when given several hours after viral challenge. The results are encouraging for antibody-based postexposure prophylaxis and support the notion that antibody induction could contribute to an effective vaccine.     

The importance of MHC-I and MHC-II responses in vaccine efficacy against lethal herpes simplex virus type 1 challenge

To investigate the importance of major histocompatability complex (MHC) class I- and MHC class II-dependent immune responses in herpes simplex virus-1 (HSV-1) vaccine efficacy, groups of beta 2% (MHC I-) and Ab% (MHC II-) mice were inoculated with various vaccines, and then challenged intraperitoneally with HSV-1. Following vaccination with either live avirulent HSV-1, expressed HSV-1 glycoprotein D (gD), or a mixture of seven expressed HSV-1 glycoproteins (7gPs), Ab% (MHC-II-) mice developed no enzyme-linked immunosorbent assay (ELISA) or neutralizing antibody titres. In contrast, significant ELISA and neutralizing antibody titres were induced in beta 2m% (MHC-I-) mice by all three vaccines. The neutralizing antibody titres were similar for all three vaccines, but were only approximately 1/4 to 1/3 of that developed in C57BL/6 (parental) mice vaccinated with the same antigens. All three vaccines protected 100% of the wild-type C57BL/6 mice against lethal challenge with 2 x 10(7) plaque-forming units (PFU) of HSV-1. The live virus vaccine and the 7gPs vaccine also protected 80% of the beta 2m% mice against the same lethal HSV-1 challenge dose. In contrast, in Abo/o mice, none of the vaccines provided significant protection against the same lethal challenge dose of HSV-1. However, at a lower challenge dose of 2 x 10(6) PFU, all three vaccines protected 70-80% of the vaccinated Ab% mice (compared to only 10% survival in mock vaccinated controls). Thus, vaccination provided some protection against lethal HSV-1 challenge in both beta 2m% and Ab% mice; however, the protection was less than that seen in the parental C57BL/6 mice. In addition, Ab% mice were less well protected by vaccination than were beta 2m% mice. Our results suggest that (1) both MHC-I and MHC-II are involved in vaccine efficacy against HSV-1 challenge; (2) both types of responses must be present for maximum vaccine efficacy: and (3) the MHC-II-dependent immune response appeared to be more important than the MHC-I-dependent immune response for vaccine efficacy against HSV-I challenge.     

Relations in families with a mentally retarded child from the perspective of the siblings

Five monoclonal antibodies (MoAbs) against Indian reference/vaccine strain of foot-and-mouth disease (FMD) virus subtype A22 (IND17/77) and a guinea pig antibody against a synthetic peptide representing amino acids (aa) 136-151 of VP1 polypeptide of A22 virus were used in the study. All the antibodies either failed to react or showed a reduced reactivity with trypsin-treated (TT)-146 S virus particles in enzyme-linked immunosorbent assay (ELISA), and could neutralize the infectivity of the reference virus. The antibodies were hence identified as specific to a trypsin-sensitive neutralizable antigenic site of the virus. Using the antibodies we isolated mutants which showed either no or reduced reactivity with the homologous as well as heterologous antibodies in ELISA. The mutants could not be neutralized with the respective antibodies but were efficiently neutralized with the serum from vaccinated cattle (BVS). These results indicated that the antibodies elicited in cattle following vaccination protected them adequately against the mutants selected and that the trypsin-sensitive neutralizable antigenic site of FMD A22 virus as identified by the MoAbs may not be dominant in eliciting a neutralizing antibody response in vaccinated cattle.     

Neutralizing antibodies from the sera of human immunodeficiency virus type 1-infected individuals bind to monomeric gp120 and oligomeric gp140

Antibodies that neutralize primary isolates of human immunodeficiency virus type 1 (HIV-1) appear during HIV-1 infection but are difficult to elicit by immunization with current vaccine products comprised of monomeric forms of HIV-1 envelope glycoprotein gp120. The limited neutralizing antibody response generated by gp120 vaccine products could be due to the absence or inaccessibility of the relevant epitopes. To determine whether neutralizing antibodies from HIV-1-infected patients bind to epitopes accessible on monomeric gp120 and/or oligomeric gp140 (ogp140), purified total immunoglobulin from the sera of two HIV-1-infected patients as well as pooled HIV immune globulin were selectively depleted of antibodies which bound to immobilized gp120 or ogp140. After passage of each immunoglobulin preparation through the respective columns, antibody titers against gp120 and ogp140 were specifically reduced at least 128-fold. The gp120- and gp140-depleted antibody fraction from each serum displayed reduced neutralization activity against three primary and two T-cell line-adapted (TCLA) HIV-1 isolates. Significant residual neutralizing activity, however, persisted in the depleted sera, indicating additional neutralizing antibody specificities. gp120- and ogp140-specific antibodies eluted from each column neutralized both primary and TCLA viruses. These data demonstrate the presence and accessibility of epitopes on both monomeric gp120 and ogp140 that are specific for antibodies that are capable of neutralizing primary isolates of HIV-1. Thus, the difficulties associated with eliciting neutralizing antibodies by using current monomeric gp120 subunit vaccines may be related less to improper protein structure and more to ineffective immunogen formulation and/or presentation.     

Virus-neutralizing antibodies of immunoglobulin G (IgG) but not of IgM or IgA isotypes can cure influenza virus pneumonia in SCID mice

The ability of monoclonal antibodies (MAbs) to passively cure an influenza virus pneumonia in the absence of endogenous T- and B-cell responses was investigated by treating C.B-17 mice, homozygous for the severe combined immunodeficiency (SCID) mutation, with individual monoclonal antiviral antibodies 1 day after pulmonary infection with influenza virus PR8 [A/PR/8/34 (H1N1)]. Less than 10% of untreated SCID mice survived the infection. By contrast, 100% of infected SCID mice that had been treated with a single intraperitoneal inoculation of at least 175 micrograms of a pool of virus-neutralizing (VN+) antihemagglutinin (anti-HA) MAbs survived, even if antibody treatment was delayed up to 7 days after infection. The use of individual MAbs showed that recovery could be achieved by VN+ anti-HA MAbs of the immunoglobulin G1 (IgG1), IgG2a, IgG2b, and IgG3 isotypes but not by VN+ anti-HA MAbs of the IgA and IgM isotypes, even if the latter were used in a chronic treatment protocol to compensate for their shorter half-lives in vivo. Both IgA and IgM, although ineffective therapeutically, protected against infection when given prophylactically, i.e., before exposure to virus. An Fc gamma-specific effector mechanism was not an absolute requirement for antibody-mediated recovery, as F(ab')2 preparations of IgGs could cure the disease, although with lesser efficacy, than intact IgG. An anti-M2 MAb of the IgG1 isotype, which was VN- but bound well to infected cells and inhibited virus growth in vitro, failed to cure. These observations are consistent with the idea that MAbs of the IgG isotype cure the disease by neutralizing all progeny virus until all productively infected host cells have died. VN+ MAbs of the IgA and IgM isotypes may be ineffective therapeutically because they do not have sufficient access to all tissue sites in which virus is produced during influenza virus pneumonia.     

Immunoglobulin G is the main protective antibody in mouse vaginal secretions after vaginal immunization with attenuated herpes simplex virus type 2

We investigated the protective role of antibodies in vaginal secretions of mice that were immune to vaginal challenge with herpes simplex virus type 2 (HSV-2). Unfractionated vaginal immunoglobulins from immune and nonimmune mice and affinity-purified immunoglobulin G (IgG) and secretory IgA (S-IgA) from immune secretions were adjusted to their concentrations in vivo. Wild-type HSV-2 was incubated in the immunoglobulin preparations for 15 min in vitro, followed by inoculation into vaginae of nonimmune mice. HSV-2 was neutralized by unfractionated antibody and purified IgG from immune secretions but not by unfractionated nonimmune antibody or by purified immune S-IgA. The protective effect of IgG in vivo was investigated by passively transferring purified serum IgG from immune and nonimmune donors to nonimmune recipients before vaginal challenge infection. Immune IgG significantly reduced the percentage of vaginal epithelium infected, concentrations of shed virus protein in the vaginal lumen, and illness scores, even though the viral antibody titers in serum and vaginal secretions of recipient mice at the time of challenge were only 29 and 8%, respectively, of those in actively immunized mice. Additionally, removal of vaginal secretions from immune mice 10 min before vaginal challenge with HSV-2 significantly increased the concentration of shed virus protein in the vaginal lumen after challenge. Collectively, the data indicate that IgG antibody in vaginal secretions of immune mice provides early protection against vaginal challenge infection, probably by neutralizing virus in the vaginal lumen. In contrast, S-IgA antibody contributed relatively little to immune protection of the vagina.     

Comparative analysis of humoral immune responses to HIV type 1 envelope glycoproteins in mice immunized with a DNA vaccine, recombinant Semliki Forest virus RNA, or recombinant Semliki Forest virus particles

The Semliki Forest virus (SFV) system seems to be a useful new approach for generating effective immune responses against HIV-1 in animal models. We evaluated this system by comparing the humoral immune responses raised in mice immunized against the HIV-1 envelope with the SFV system, a DNA vaccine, and a recombinant Env glycoprotein. gp160 ELISA antibody titers (204,800) were highest in the sera from mice immunized with recombinant Semliki Forest virus particles. These sera contained antibodies to the CD4-binding site and recognized linear epitopes on gp120 and gp41 that were also recognized by a pool of sera from HIV1-infected individuals. This demonstrates that the HIV-1 envelope produced in vivo by the SFV system does not fold aberrantly. A low level of neutralizing antibodies against the HIV-1LAI strain was also detected in the serum of one mouse immunized with recombinant SFV particles, suggesting that booster injections should be given to achieve a more effective immune response. SFV recombinant particles induced the strongest humoral responses to the HIV-1 envelope of all the potential HIV env vaccines tested.     

Passive immunization with a human immunodeficiency virus type 1-neutralizing monoclonal antibody in Hu-PBL-SCID mice: isolation of a neutralization escape variant

A monoclonal antibody (694/98-D) directed toward the V3 loop of human immunodeficiency virus type 1 (HIV-1) was evaluated for pre- and postexposure prophylaxis in SCID mice reconstituted with human peripheral blood lymphocytes (Hu-PBL-SCID). Fifty percent protection against the HIV-1LAI strain was obtained by preexposure administration of 1.32 mg/kg antibody. However, virus isolated from 1 mouse 3 weeks after passive immunization with 13.2 mg/kg antibody proved resistant to subsequent in vitro neutralization by 694/98-D. V3 loop sequence analysis of cloned virus revealed amino acid changes within the linear core epitope recognized by 694/98-D and in one flanking amino acid. Further evaluation of 694/98-D for postexposure prophylaxis in mice revealed that 694/ 98-D was effective when given 15 min after virus. However, efficacy declined to 50% if treatment was delayed to 1 h after virus inoculation. These studies point out some potential drawbacks of passive immunization with monoclonal antibodies.     

Monoclonal anti thy 1.2 antibodies from hybridoma HO13-4 do not react with mouse natural killer cells

The susceptibility of NK cells and immune cytotoxic T-cells to treatment with (a) monoclonal anti thy 1.2 antibodies from hybridoma HO13-4, (b) rabbit anti-mouse T-cell antiserum and (c) gamma globulins prepared from AKR/J anti C3H/HeJ antiserum was studied in the presence of rabbit complement. Monoclonal anti thy 1.2 antibody treatment completely abolished the cytotoxic activity of immune T-cells derived from C57BL/6J mice (H-2b) immunized with (C57BL/6J x DBA/2)F1 spleen cells (H-2bd) against P815 (H-2d) target cells. The same treatment had no significant effect on the NK activity of spleen cells from unimmunized mice against YAC target cells. Rabbit anti-mouse T-cell and mouse anti theta antisera also abrogated completely the immune T cell activity of spleen cells. This treatment however also resulted in a partial loss of NK activity. These results indicate that conventional anti theta antisera contain antibodies which recognize antigenic specificities on T-cells as well as on a population of NK cells. The cross reactivity is not a result of thy 1.2 antigen expression on NK cells and T-cells as recognized by the monoclonal antibodies. The specificity recognized by the monoclonal antibody (HO13-4) is only expressed on T-cells.     

Suppression of immunological response against a transgene product delivered from microencapsulated cells

A potential obstacle to successful gene therapy for some patients is the in vivo production of neutralizing antibodies against the recombinant therapeutic product delivered. To mimic this clinical situation, we implanted microencapsulated recombinant cells producing human growth hormone into C57B1/6 mice to provoke antihuman growth hormone antibody production. We then investigated the efficacy of different immunosuppressive treatments to inhibit the development of neutralizing antibodies. The experimental mice were treated with either an immunosuppressive drug (FK506 or cyclophosphamide), a cytokine (interferon-gamma [IFN-gamma] or interleukin-12 [IL-12], or a monoclonal antibody (anti-CD4, anti-gp39, or CTLA4-Ig). Serum human growth hormone and mouse anti-human growth hormone antibody levels were measured by enzyme-linked immunosorbent assay (ELISA) for 4 weeks. There were three patterns of response noted among the seven treatment groups. First, the mice receiving IFN-gamma, IL-12, anti-gp39, or CTLA4-Ig were similar to the untreated controls-no suppression of anti-hGH antibodies and no improvement in delivery of hGH. Next, the mice receiving FK506 or cyclosphosphamide showed > or = 90% suppression of antibodies but also no improvement in product delivery. Last, the mice receiving anti-CD4 showed almost complete antibody suppression over 1 month postimplantation. Furthermore, only anti-CD4 permitted a sustained level of human growth hormone delivery to day 28, in contrast to the controls whose human growth hormone delivery was undetectable by day 14 postimplantation. Hence, the use of anti-CD4 inhibited formation of neutralizing antibodies against a recombinant gene product delivered in vivo, and allowed prolonged delivery of a foreign protein. Its role as adjunct treatment for appropriate patients receiving gene therapy should be examined further.     

Influenza virus neutralizing antibodies and IgG isotype profiles after immunization of mice with influenza A subunit vaccine using various adjuvants

The influence of various adjuvants on the development of influenza virus neutralizing antibodies and distribution of anti-influenza virus IgG isotypes after immunization of mice with influenza A (H3N2) subunit vaccine was investigated. Serum titres of influenza virus neutralizing antibodies and titres of influenza specific IgG isotypes were determined by a neutralization enzyme immunoassay (N-EIA) and a cell-associated antigen enzyme immunoassay (CA-EIA), respectively. Serum antibody titres as measured by the two tests correlated highly (r = 0.82; P < 0.001). N-EIA titres were enhanced by 38- and 34-fold, when L180.5/RaLPS and FCA, respectively, were administered with 1 microgram of vaccine. The adjuvants Q-VAC, L180.5 [W/O/W], L180.5 alone and Montanide ISA 740 were only moderately or not effective in enhancing the immune response to the 1 microgram dose of vaccine. The Q-VAC and L180.5/RaLPS adjuvants favoured IgG2a and IgG2b isotype responses to influenza compared to the other adjuvants. We suggest that N-EIA and CA-EIA may be valuable tools to monitor the effects of adjuvants on the neutralizing antibody and antibody isotype responses after influenza vaccination.