Plasma Antibody Titers Research Articles

Outer membrane vesicles (OMVs) from Tenacibaculum maritimum as a potential vaccine against fish tenacibaculosis

Outer membrane vesicles (OMVs) have been gained increasing attention in vaccinology due to their ability to induce strong protective humoral and cell-mediated immunity. The Gram-negative bacterium Tenacibaculum maritimum, the causative agent of marine tenacibaculosis, poses a significant challenge to the global aquaculture industry due to its difficult prophylaxis. In previous studies, we demonstrated that OMV production is a key virulence mechanism in T. maritimum. Building on this, the present study aimed to evaluate the efficacy of a natural, encapsulated multi-antigen vaccine made from adjuvant-free, crude T. maritimum OMVs (Tm-OMVs). A vaccination experiment using SP9.1-OMVs was conducted in juvenile turbot (Scophthalmus maximus L.), followed by a T. maritimum bath challenge. Immune responses in the turbot were assessed by measuring anti-Tm antibody levels and analyzing the expression of eight key immune-related genes (il-1β, il-8, il-22, pcna, c3, cd4-1, ifng2, cd8α). The results showed that immunization with SP9.1-OMVs provided significant protection against T. maritimum infection (RPS = 70 %). Vaccinated fish exhibited a dose-dependent increase in anti-Tm antibody titers in blood plasma, along with rapid induction of both innate (il-1β, il-8, il-22, c3) and adaptive (cd4-1, ifng2, cd8α) immune genes as early as 4 h post-bath challenge. These findings offer new insights into the early immune response of turbot following T. maritimum infection and could serve as a foundation for developing novel OMV-based vaccines.

Metabolomic profiling of maternal plasma identifies inverse associations of acetate and urea with anti-SARS-CoV-2 antibody titers following COVID-19 vaccination during pregnancy.

We conducted a comprehensive metabolomic analysis of plasma samples obtained from pregnant women who displayed varying post-vaccination antibody titers after receiving mRNA-1273-SARS-CoV-2 vaccines. The study involved 62 pregnant women, all of whom had been vaccinated after reaching 24weeks of gestation. To quantify post-vaccination plasma antibody titers, we employed binding antibody units (BAU) in accordance with the World Health Organization International Standard. Subsequently, we classified the study participants into three distinct BAU/mL categories: those with high titers (above 2000), medium titers (ranging from 1000 to 2000), and low titers (below 1000). Plasma metabolomic profiling was conducted using 1H nuclear magnetic resonance spectroscopy, and the obtained data were correlated with the categorized antibody titers. Notably, in pregnant women exhibiting elevated anti-SARS-CoV-2 antibody titers, reduced plasma concentrations of acetate and urea were observed. A significant negative correlation between these compounds and antibody titers was also evident. An analysis of metabolomics pathways revealed significant inverse associations between antibody titers and four distinct amino acid metabolic pathways: (1) biosynthesis of phenylalanine, tyrosine, and tryptophan; (2) biosynthesis of valine, leucine, and isoleucine; (3) phenylalanine metabolism; and (4) degradation of valine, leucine, and isoleucine. Additionally, an association between the synthesis and degradation pathways of ketone bodies was evident. In conclusion, we identified different metabolic pathways that underlie the diverse humoral responses triggered by COVID-19 mRNA vaccines during pregnancy. Our data hold significant implications for refining COVID-19 vaccination approaches in expectant mothers. KEY MESSAGES : Anti-SARS-CoV-2 antibody titers decline as the number of days since COVID-19 vaccination increases. Anti-SARS-CoV-2 antibody titers are inversely associated with acetate, a microbial-derived metabolite, and urea. Amino acid metabolism is significantly associated with SARS-CoV-2 antibody titers.

Intravenous Bacille Calmette–Guérin vaccination protects simian immunodeficiency virus-infected macaques from tuberculosis

Tuberculosis, caused by Mycobacterium tuberculosis (Mtb), is the most common cause of death in people living with human immunodeficiency virus (HIV). Intra-dermal Bacille Calmette–Guérin (BCG) delivery is the only licensed vaccine against tuberculosis; however, it offers little protection from pulmonary tuberculosis in adults and is contraindicated in people living with HIV. Intravenous BCG confers protection against Mtb infection in rhesus macaques; we hypothesized that it might prevent tuberculosis in simian immunodeficiency virus (SIV)-infected macaques, a model for HIV infection. Here intravenous BCG-elicited robust airway T cell influx and elevated plasma and airway antibody titres in both SIV-infected and naive animals. Following Mtb challenge, all 7 vaccinated SIV-naive and 9 out of 12 vaccinated SIV-infected animals were protected, without any culturable bacteria detected from tissues. Peripheral blood mononuclear cell responses post-challenge indicated early clearance of Mtb in vaccinated animals, regardless of SIV infection. These data support that intravenous BCG is immunogenic and efficacious in SIV-infected animals.

Anti-FVIII antibodies in Black and White hemophilia A subjects: do F8 haplotypes play a role?

The most common complication in hemophilia A (HA) treatment, affecting 25% to 30% of patients with severe HA, is the development of alloimmune inhibitors that foreclose the ability of infused factor VIII (FVIII) to participate in coagulation. Inhibitors confer significant pathology on affected individuals and present major complexities in their management. Inhibitors are more common in African American patients, and it has been hypothesized that this is a consequence of haplotype (H)-treatment product mismatch. F8 haplotypes H1 to H5 are defined by nonsynonymous single-nucleotide polymorphisms encoding sequence variations at FVIII residues 1241, 2238, and 484. Haplotypes H2 to H5 are more prevalent in individuals with Black African ancestry, whereas 80% to 90% of the White population has the H1 haplotype. This study used an established multiplex fluorescence immunoassay to determine anti-FVIII antibody titers in plasma from 394 individuals with HA (188 Black, 206 White), measuring their binding to recombinant full-length H1 and H2 and B-domain-deleted (BDD) H1/H2, H3/H5, and H4 FVIII proteins. Inhibitor titers were determined using a chromogenic assay and linear B-cell epitopes characterized using peptide microarrays. FVIII-reactive antibodies were readily detected in most individuals with HA, with higher titers in those with a current inhibitor, as expected. Neither total nor inhibitory antibody titers correlated with F8 haplotype mismatches, andpeptides with D1241E and M2238V polymorphisms did not comprise linear B-cell epitopes. Interestingly, compared with the full-length FVIII products, the BDD-FVIII proteins were markedly more reactive with plasma antibodies. The stronger immunoreactivity of BDD-FVIII suggests that B-domain removal might expose novel B-cell epitopes, perhaps through conformational rearrangements of FVIII domains.

PB2270: CLEARANCE OF PERSISTENT SARS-COV2 INFECTION WITH ANTIBODY CONTAINING PLASMA IN LEUKEMIA AND LYMPHOMA PATIENTS

Background: Patients with hematological malignancies or other reasons for inadequate immune response are at high risk for poor outcome of COVID-19 disease. In addition, a significant fraction of patients unable to mount an antibody response fail to clear SARS-Cov2 virus putting them at risk for clinical deterioration. Also, persistent COVID-19 infection impedes further treatment of the underlying disease. So far, no treatment has been established for persistent COVID-19 infection in immunocompromised patients. Aims: Here, we treated eight immunocompromised patients with active SARS-COV2 infection with plasma from convalescent or/and vaccinated donors in a case series of individual treatments. Methods: Patients were eligible for treatment if active SARS-Cov2 infection was confirmed at least 14 days after initial detection. In two out of eight patients sequencing results confirmed the omicron variant. Therapies before plasma treatment included antivirals and monoclonal antibodies. Underlying diseases were B-ALL (n=3; including one patient after allogeneic stem cell transplantation), multiple myeloma (n=2), lymphoma (n=2) and immunosuppression after kidney transplantation (n=1). Median age was 48 years (range 7 – 78). Antibody containing plasma was obtained from healthy donors either after confirmed SARS-COV2 infection or after mRNA vaccination. Overall, 22 treatment cycles were administered in 8 patients (median 2, range 1 to 7). For each patient the available plasma with the highest titers of neutralizing antibodies and matching blood type was administered. Results: Neutralizing antibody titers in plasma were median 1:320 (range 1:40 to 1:2560). Titers of neutralizing antibodies in convalescent plasma (n=10 treatments; median titer: 1:80) were lower than in vaccinated donor plasma (median 1:320; p=0.058, Mann-Whitney U test) (n=12 treatments). Median Ct-values of viral load before treatment were 20.5 (range 14.9 to 37.3). After treatment (median 5 days), the Ct-values changed to median 26.9 (range 19.5 to non-detectable). The median fold decrease in viral load was estimated to be 127-fold with a range from 0.8 to >2x104 (p<10-4, paired t-test). Similar reductions in viral load were achieved after administration of either plasma from convalescent (median 109) or vaccinated donors. SARS-COV2 virus was cleared in 7 of 8 (87.5%) patients with ongoing treatment in one patient. No specific adverse events were observed. Figure 1: Fold reduction in viral load after a single treatment with anti-SARS-COV2 containing plasma. Image:Summary/Conclusion: Plasma with high titers of neutralizing SARS-COV2 antibodies can clear persistent infection in patients with hematological malignancies and immunocompromised patients. Plasma obtained either from convalescent patients or vaccinated donors might constitute an effective and inexpensive treatment in this situation.

Single-cell RNA sequencing reveals immunological rewiring at the maternal-fetal interface following asymptomatic/mild SARS-CoV-2 infection.

While severe coronavirus 2019 (COVID-19) is associated with immune activation at the maternal-fetal interface, responses to asymptomatic/mild severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection during pregnancy remain unknown. Here, we assess immunological adaptations in blood and term decidua in response to asymptomatic/mild disease in pregnant women. We report attenuated antigen presentation and type I interferon (IFN) signaling pathways, loss of tissue-resident decidual macrophages, and upregulated cytokine/chemokine signaling in monocyte-derived decidual macrophages. Furthermore, we describe increased frequencies of activated tissue-resident T cells and decreased abundance of regulatory T cells with infection while frequencies of cytotoxic CD4/CD8 T cells are increased in the blood. In contrast to decidual macrophages, type I IFN signaling is higher in decidual T cells. Finally, infection leads to a narrowing of T cell receptor diversity in both blood and decidua. Collectively, these observations indicate that asymptomatic/mild COVID-19 during pregnancy results in remodeling of the immunological landscape of the maternal-fetal interface, with a potential for long-term adverse outcomes for the offspring.

A possible dose-response equation: Viral load after plasma infusion in COVID-19 patients and anti-SARS-CoV-2 antibody titers in convalescent plasma.

Clinical trials of convalescent plasma therapy for coronavirus disease 2019 (COVID-19) are extensive, but the relationship between antibody titers, infused volume of plasma and virus clearance in patients remains unknown. This study proposed a possible estimating equation for clinical use of high antibody titer convalescent plasma. A total of 38 patients were recruited in the Guanggu District Maternal and Child Health Hospital of Hubei Province from March 1 to 30, 2020. COVID-19 convalescent plasma was collected and high-titer (≥1:640) anti-S-RBD units used. The SARS-CoV-2 nucleic acid viral load was measured 24 h before and 72 h after convalescent plasma infusion. Convalescent plasma therapy was associated with reduced viral load in patients with moderate and severe severity. The viral negative rate at 72 h was 65.8%. The disappearance of viral nucleic acid in study patients was positively correlated with infuscate antibody titer and volume (r=0.3375, p=0.04). A possible estimation equation was as follows: Log10 (Reduction in viral load)=0.18 + 0.001 × (Log2 S-RBD antibody titer × Plasma infusion volume) (r=0.424, p=0.009). In a single case, the viral nucleic acid persisted 14 days after the fourth plasma infusion. This study proposes a potential dose-response equation that adds a convenient way to estimate the dose of convalescent plasma product. It is beneficial to facilitate the rational allocation of plasma with high antibody titers and provide an individualised use strategy for convalescent plasma therapy.

Induction of Functional Specific Antibodies, IgG-Secreting Plasmablasts and Memory B Cells Following BCG Vaccination.

Tuberculosis (TB) is a major global health problem and the only currently-licensed vaccine, BCG, is inadequate. Many TB vaccine candidates are designed to be given as a boost to BCG; an understanding of the BCG-induced immune response is therefore critical, and the opportunity to relate this to circumstances where BCG does confer protection may direct the design of more efficacious vaccines. While the T cell response to BCG vaccination has been well-characterized, there is a paucity of literature on the humoral response. We demonstrate BCG vaccine-mediated induction of specific antibodies in different human populations and macaque species which represent important preclinical models for TB vaccine development. We observe a strong correlation between antibody titers in serum versus plasma with modestly higher titers in serum. We also report for the first time the rapid and transient induction of antibody-secreting plasmablasts following BCG vaccination, together with a robust and durable memory B cell response in humans. Finally, we demonstrate a functional role for BCG vaccine-induced specific antibodies in opsonizing mycobacteria and enhancing macrophage phagocytosis in vitro, which may contribute to the BCG vaccine-mediated control of mycobacterial growth observed. Taken together, our findings indicate that the humoral immune response in the context of BCG vaccination merits further attention to determine whether TB vaccine candidates could benefit from the induction of humoral as well as cellular immunity.

Different adjuvanted pediatric HIV envelope vaccines induced distinct plasma antibody responses despite similar B cell receptor repertoires in infant rhesus macaques.

Different HIV vaccine regimens elicit distinct plasma antibody responses in both human and nonhuman primate models. Previous studies in human and non-human primate infants showed that adjuvants influenced the quality of plasma antibody responses induced by pediatric HIV envelope vaccine regimens. We recently reported that use of the 3M052-SE adjuvant and longer intervals between vaccinations are associated with higher magnitude of antibody responses in infant rhesus macaques. However, the impact of different adjuvants in HIV vaccine regimens on the developing infant B cell receptor (BCR) repertoire has not been studied. This study evaluated whether pediatric HIV envelope vaccine regimens with different adjuvants induced distinct antigen-specific memory B cell repertoires and whether specific immunoglobulin (Ig) immunogenetic characteristics are associated with higher magnitude of plasma antibody responses in vaccinated infant rhesus macaques. We utilized archived preclinical pediatric HIV vaccine studies PBMCs and tissue samples from 19 infant rhesus macaques immunized either with (i) HIV Env protein with a squalene adjuvant, (ii) MVA-HIV and Env protein co-administered using a 3-week interval, (iii) MVA-HIV prime/ protein boost with an extended 6-week interval between immunizations, or (iv) with HIV Env administered with 3M-052-SE adjuvant. Frequencies of vaccine-elicited HIV Env-specific memory B cells from PBMCs and tissues were similar across vaccination groups (frequency range of 0.06-1.72%). There was no association between vaccine-elicited antigen-specific memory B cell frequencies and plasma antibody titer or avidity. Moreover, the epitope specificity and Ig immunogenetic features of vaccine-elicited monoclonal antibodies did not differ between the different vaccine regimens. These data suggest that pediatric HIV envelope vaccine candidates with different adjuvants that previously induced higher magnitude and quality of plasma antibody responses in infant rhesus macaques were not driven by distinct antigen-specific memory BCR repertoires.

Effect of time and titer in convalescent plasma therapy for COVID-19

SummaryThe clinical benefit of convalescent plasma (CP) for patients with coronavirus disease (COVID)-19 is still debated. In this systematic review and meta-analysis, we selected 10 randomized clinical trials (RCTs) and 15 non-randomized studies (total number of patients = 22,591) of CP treatment and evaluated two different scenarios: (1) disease stage of plasma recipients and (2) donated plasma antibody titer, considering all-cause mortality at the latest follow-up. Our results show that, when provided at early stages of the disease, CP significantly reduced mortality: risk ratio (RR) 0.72 (0.68, 0.77), p < 0.00001, while provided in severe or critical conditions, it did not (RR: 0.94 [0.86, 1.04], p = 0.22). On the other hand, the benefit on mortality was not increased by using plasma with a high-antibody titer compared with unselected plasma. This meta-analysis might promote CP usage in patients with early-stage COVID-19 in further RCTs to maximize its benefit in decreasing mortality, especially in less affluent countries.

STAT6 signaling pathway controls germinal center responses promoted after antigen targeting to conventional type 2 dendritic cells

Conventional dendritic cells (cDCs) are antigen-presenting cells specialized in naïve T cell priming. Mice splenic cDCs are classified as cDC1s and cDC2s, and their main functions have been elucidated in the last decade. While cDC1s are specialized in priming type 1 helper T cells (TH1) and in cross presentation, cDC2s prime T follicular helper (TFH) cells that stimulate germinal center (GC) formation, plasma cell differentiation and antibody production. However, less is known about the molecular mechanisms used by cDCs to prime those responses. Here, using WT and STAT6-deficient mice (STAT6 KO), we targeted a model antigen to cDC1s and cDC2s via DEC205 and DCIR2 receptors, respectively, in an attempt to study whether the STAT6 signaling pathway would modulate cDCs’ ability to prime helper T cells. We show that the differentiation and maturation of cDCs, after stimulation with an adjuvant, were comparable between WT and STAT6 KO mice. Besides, our results indicate that, in STAT6 KO mice, antigen targeting to cDC2s induced reduced TFH and GC responses, but did not alter plasma cells numbers and antibody titers. Thus, we conclude that the STAT6 signaling pathway modulates the immune response after antigen targeting to cDC2s via the DCIR2 receptor: while STAT6 stimulates the development of TFH cells and GC formation, plasma cell differentiation occurs in a STAT6 independent manner.

Clinical characteristics and plasma antibody titer of patients with COVID-19 in Zhejiang, China.

Coronavirus disease 2019 (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which first affected humans in China on December 31, 2019 (Shi et al., 2020). Coronaviruses generally cause mild, self-limiting upper respiratory tract infections in humans, such as the common cold, pneumonia, and gastroenteritis (To et al., 2013; Berry et al., 2015; Chan et al., 2015). According to the Report of the World Health Organization (WHO)-China Joint Mission on COVID-19 (WHO, 2020), the case fatality rate of COVID-19 increases with age, while the rate among males is higher than that among females (4.7% and 2.8%, respectively). Since an effective vaccine and specific anti-viral drugs are still under development, passive immunization using the convalescent plasma (CP) of recovered COVID-19 donors may offer a suitable therapeutic strategy for severely ill patients in the meantime. So far, several studies have shown therapeutic efficacy of CP transfusion in treating COVID-19 cases. A pilot study first reported that transfusion of CP with neutralizing antibody titers above 1:640 was well tolerated and could potentially improve clinical outcomes through neutralizing viremia in severe COVID-19 cases (Chen et al., 2020). Immunoglobulin G (IgG) and IgM are the most abundant and important antibodies in protecting the human body from viral attack (Arabi et al., 2015; Marano et al., 2016). Our study aimed to understand the aspects of plasma antibody titer levels in convalescent patients, as well as assessing the clinical characteristics of normal, severely ill, and critically ill patients, and thus provide a basis for guiding CP therapy. We also hoped to find indicators which could serve as a reference in predicting the progression of the disease.

Convalescent Plasma for the Treatment of COVID-19: Perspectives of the National Institutes of Health COVID-19 Treatment Guidelines Panel.

In the United States, the efficacy and safety of convalescent plasma for treating coronavirus disease 2019 (COVID-19) is currently being tested in randomized placebo-controlled clinical trials. Treatment of individual patients with COVID-19 with convalescent plasma outside such trials is also now permitted through U.S. Food and Drug Administration Emergency Use Authorization. Here, members of the National Institutes of Health COVID-19 Treatment Guidelines Panel provide their views regarding use of convalescent plasma for treating COVID-19.

Recruitment Strategy for Potential COVID-19 Convalescent Plasma Donors

Recruitment Strategy for Potential COVID-19 Convalescent Plasma Donors

Investigation on compliance with sepsis bundle and outcomes in patients with septic shock in Changshu area

Objective: To perform a statistical analysis of the ABO blood group distribution of COVID-19 convalescents, and further analyze the ABO blood group distribution in COVID-l9 convalescents with different plasma antibody titer against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-Z).

Role of serology in covid 19

SARS CoV2 infection is spreading all over the world and had caused a pandemic of unheard proportions. Accurate and rapid diagnosis of cases is important for treatment, isolation and epidemiological purposes. So far RT-PCR is the gold standard for providing diagnosis, however detection of antibodies specific to SARS CoV2 can provide another means of diagnosis and also serve as an important epidemiological tool. As the world is grappling with the COVID19 pandemic, there is increasing global interest in the role of serological testing for population monitoring, detecting asymptomatic infections and for plasma antibody titres to be used for therapeutics. Many commercial serological assays have become available, however, for interpretation of these assays it is important to know the kinetics of antibody development. In this review an overview on the role of serology in COVID 19 has been provided.

Efficacy of two adjuvant systems to promote humoral immunity to the pre-proghrelin peptide obestatin in pigs: consequences for the growth of piglets to weaning

The poor antigenicity of peptide antigens demands the selection of effective adjuvants to induce humoral immunity. The peptides obestatin and ghrelin from the pro-hormone pre-proghrelin were initially identified as antagonistic in regulating feeding behaviour, with obestatin being suppressive. The efficacy of two adjuvant systems, DEAE with the oil polysorbate emulsion of BP85:Span80 and the surfactant-oil system Montanide (ISA 50v) were therefore assessed with an obestatin-ovalbumin conjugate injected into late pregnant sows. This enabled the supply of antibodies directed against obestatin to newborn piglets through colostrum with the objective of promoting ghrelin secretion and therefore increasing feeding behaviour. Pregnant Landrace × Large White sows (n = 28) were immunised with 0.5 mg obestatin-ovalbumin in 2 mL DEAE:BP85:Span80 (DEAE; n = 14) or with 2 mL Montanide (ISA 50v: n = 14) as adjuvants at days 91 and 105 of gestation. After farrowing, piglets remained with their mothers during the lactation period and were weighed after weaning at Day 28. Antibody titres (unitless) in colostrum were assessed by ELISA as 5543 ± 2388 and 3139 ± 1151 for the DEAE and Montanide adjuvants respectively. These were associated with total IgG of 67.7 ± 3 and 82.3 ± 4.8 mg/mL respectively (P = 0.018). Piglet plasma titres were 5100 ± 1576 and 5762 ± 1688 for DEAE and Montanide respectively at Day 5 postpartum. These titres were still detectable through to Day 28 (titres of 1213 ± 389 and 665 ± 203 respectively (P = 0.176). However, sow colostral antibody titres were not related to piglet antibody concentrations on D5 (r = –0.225, P = 0.341). Sow plasma antibody titres were not related to titres at Day 28 in piglets across treatments (r = 0.198, P = 0.402). The concentration of ghrelin in colostrum was 672 ± 78 and 666 ± 39 pg/mL for the DEAE and Montanide groups, respectively, leading to piglet plasma concentrations on Day 5 of 1105 ± 164 and 530 ± 84 pg/mL (P = 0.002). Animals grew from birthweights of 1.7 ± 0.1 and 1.8 ± 0.1 (P = 0.993) to 7.7 ± 1.2 and 7.8 ± 1.0 kg (P = 0.295) at weaning, representing growth rates of 200.5 ± 52.9 and 225.5 ± 53.4 g/day (P = 0.181). There was a significant negative correlation between piglet D28 antibody titre and growth rate to weaning with the Montanide adjuvant (r = 0.116, P = 0.035) but not for the DEAE (r = –0.118, P = 0.411). Although both adjuvants were capable of generating high antibody titres, the DEAE dextran was likely to be the most effective adjuvant to induce a humoral immune response to develop further with a commercial vaccine.

Neutralizing and Non-Neutralizing Anti-FVIII Antibodies in Black and White Hemophilia A Subjects: A Natural History Profile

The most common complication in hemophilia A (HA) treatment, affecting 25-30% of severe HA patients, is the development of alloimmune inhibitors that foreclose the ability of infused factor VIII (FVIII) to participate in coagulation. Inhibitors confer significant pathology on affected individuals and present major complexities in their management. Inhibitors are more common in African American patients, and it has been hypothesized that this is a consequence of haplotype (H)-treatment product mismatch. F8 gene haplotypes H1-H5 are defined by combinations of nonsynonymous SNPs encoding FVIII sequence variants D1241E, M2238V and R484H. F8 haplotypes H2-H5 are more prevalent in individuals with black African ancestry, while &gt;90% of the white population has the H1 haplotype. This study used a validated Luminex-based assay to determine total anti-FVIII antibody titers in plasma from 395 HA (189 black, 206 white) and 23 non-HA control subjects, measuring their binding to recombinant full-length H1 and H2 and B-domain-deleted (BDD) H1/H2, H3 and H4 FVIII proteins. Inhibitor titers were determined using a chromogenic Bethesda assay with the Nijmegen modification. Linear B-cell epitopes recognized by antibodies in human plasma samples were characterized using commercial peptide microarrays with imprinted 15-mer peptides spanning the FVIII A1, A2, C1 and C2 domains, with binding interactions detected using fluorescent-labeled anti-human IgG antibodies. Neither total nor inhibitory antibody titers correlated with F8 haplotype. FVIII peptides with the D1241E and M3348V polymorphisms showed low antibody reactivity, indicating they do not comprise linear B-cell epitopes. Similarly, antibodies from subjects with H3 and H5 haplotypes, who were necessarily infused with FVIII products having a different haplotype than that of their endogenous, (dysfunctional) F8 sequence, did not show haplotype-correlated differential binding to the three BDD-FVIII or full-length FVIII proteins, indicating the polymorphic M2238V or D1241E sites do not correspond to immunodominant, conformational B-cell epitopes. Interestingly, the BDD-FVIII proteins were significantly more reactive with antibodies in plasma than were two commercial full-length recombinant FVIII products. Overall, results of this study indicated that low-titer FVIII-reactive antibodies are readily detected in most HA subjects and in a majority of healthy non-HA controls. The observed stronger immunoreactivity of BDD-FVIII suggests that B-domain removal exposes novel B-cell epitopes, perhaps through conformational rearrangements of FVIII domains. Disclosures Pratt: Bloodworks NW: Patents &amp; Royalties: inventor on patents related to FVIII immunogenicity; Grifols, Inc: Research Funding. Peters:Sanofi: Employment. Mann:Haematologic Technologies: Other: Owner; Stago: Consultancy; Novo Nordisk: Consultancy; Takeda: Consultancy; Shire: Consultancy; Baxalta: Consultancy.

SHIV Virus-Like Particles (VLPs) Vaccination Induces Partial Protection from SHIV Challenge in a Rhesus Macaque Model

Simian/Human immunodeficiency virus (SHIV) virus-like particles (VLPs) composed of SIV Gag, HIVsf162 gp120/ gp41 envelope, and human CD40L are whole pseudovirion vaccines capable of eliciting both humoral and cellular immunity. We immunized four rhesus macaques by intranasal prime and four sub-cheek boosts with VLPs adjuvanted with conjugatable adjuvant lipid vesicles containing monophosphoryl lipid A (MPLA), and compared their immune parameters to those in five unimmunized control macaques. Increased plasma antibody titers to SIV Gag were observed in all four immunized macaques and increased sf162 gp140 titers were observed in three of the four with one macaque (10-195) maintaining sustained anti-Env antibody levels. Compared to controls, a significant increase in memory B cells and CD4+ central memory T cells was detected in the immunized group. Among these, elevated Gagspecific CD107a membrane localization in the CD8+ central memory T cells was detected in one macaque (10-195). All nine macaques were subsequently challenged intrarectally with SHIVsf162.P3. After challenge, eight of the nine macaques became infected with SHIV, while the macaque 10-195 was protected. Another immunized macaque (10-189) that got infected but consequently generated high Gag-specific IFN-γ and CD107a CD8+ T cells; and Envspecific IL-2 and CD107a CD8+ T cells controlled virus and had undetectable levels of plasma SIV copy number by the termination of the study. Our VLPs vaccine strategy represents a promising immunogenic conformationally intact HIV vaccine that may lead to possible prevention and control of HIV infection.

A time-course study of gene expression and antibody repertoire at early time post vaccination of Atlantic salmon

The majority of studies of vaccine responses in Atlantic salmon have focused on several weeks after vaccination, and employed a limited number of marker genes. In this study, novel techniques were used to examine a broad panel of expressed genes and antibody repertoire of Atlantic salmon following vaccination. Salmon parr were vaccinated with a multivalent oil-based vaccine, and blood plasma and head kidney were sampled at several time-points between 0–35 days post vaccination. Saline-injected fish were used as control at all time-points. Microarray analyses showed increased expression of immune genes from the first day to the end of study in the head kidney of vaccinated fish. Genes up-regulated in the late phase included several leukocyte markers and components of the oxidative burst complex. A suite of genes that can take part in B cells differentiation were up-regulated from day 14, at which time secretory IgM transcripts also peaked. This coincided with marked increased plasma titres of non-vaccine specific antibodies binding to a hapten-carrier antigen DNP-KLH, while antibodies to bacterial components of the vaccine, Moritella viscosa and Aeromonas salmonicida, first showed significantly elevated antibody levels at day 21, and at a markedly lower magnitude than the non-vaccine specific titres. Sequencing of the variable region of IgM heavy chain (CDR3) revealed higher cumulative frequencies of unique clonotypes in vaccinated salmon starting from day 14 when specific antibodies were first detected. Reduced sequence variance of CDR3 suggested expansion of recently emerged clonotypes. Overall, the results presented here follow a broad panel of gene expression, immunoglobulin sequencing and plasma antibody titres in the first few weeks after vaccination of Atlantic salmon, pointing to a potentially important contribution of non-vaccine specific antibody responses early in the vaccine response.