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Background: The potency of inactivated and recombinant influenza vaccines is measured using the single‐radial immunodiffusion (SRID) assay. The strain‐specific antigen and antibody potency reagents required for the assay are prepared and distributed by regulatory agencies to ensure vaccine standardization, but timely reagent production is always challenging. This poses unique concerns for rapid pandemic responses. Alternative methods have been described for generating strain‐specific potency antibody reagents without the need for live influenza virus, but such methods are infrequently used, suggesting the need for additional antigen expression approaches. Methods: We describe a rapid process using a mammalian expression system to produce recombinant influenza hemagglutinin (rHA). This platform was used to generate rHA from two H5 clade 2.3.4.4 influenza viruses, in both soluble ectodomain or full‐length HA forms, and a soluble ectodomain rHA from an influenza H2 virus. Results: The purified rHAs were used as immunogens to produce HA antibody reagents that were tested for suitability in the SRID assay to accurately measure the potency of inactivated pandemic influenza vaccines. Antibody reagents generated to either ectodomain or full‐length rHA worked well in the SRID assay and resulted in vaccine potency values equivalent to those generated with standard reference antibodies. Conclusions: The results demonstrate that rHA produced from a simple mammalian cell transfection method can be used to generate HA antibody suitable for use in the influenza vaccine SRID potency assay and suggest a practical means by which an extensive library of pandemic reagents can easily be prepared in advance of and during an influenza emergency. [ABSTRACT FROM AUTHOR]

Objective(s): In this study, the adjuvant activity of aqueous and alcoholic extracts of propolis was examined on the inactivated herpes simplex virus-1 (HSV-1). Materials and Methods: BALB/C mice were administered with inactivated (HSV-1; the KOS strain) plus alcoholic and aqueous extracts, followed by assessment of the cellular and humoral immune responses. Results: Alcoholic and aqueous extracts, as an adjuvant, revealed a significant increase in lymphocyte proliferation and cytotoxic T lymphocyte (CTL) responses versus the HSV-1 group. In addition, HSV-1 plus alcoholic extract showed a remarkable increase in IFN-γ cytokine and IFN-γ/IL-4 ratio. On the other hand, both alcoholic and aqueous extracts in the HSV-1 vaccine suppressed the IL-4 cytokine response as compared with the HSV-1 vaccine. In addition, HSV-1 plus alcoholic extract showed a significant increment in IgG1, IgG2a, and IgG2b isotypes as compared with the HSV-1 vaccine. Conclusion: Propolis extracts seem to modulate the immune response against inactivated HSV-1 model and can be used as a suitable vaccine adjuvant or a component of a complex adjuvant against infectious diseases. [ABSTRACT FROM AUTHOR]

Acknowledging the significant impact of temperature deviations on vaccine efficacy, this study examines the prevalence of exposure to non-recommended temperatures, particularly focusing on freeze-sensitive vaccines. This study aims to identify critical points of temperature breaches during transportation from storage to last-mile delivery. In this cross-sectional study, the temperature integrity of vaccines has been examined during transportation across three states in India (Karnataka, Maharashtra, and Rajasthan), utilizing data loggers to track and record temperatures in real time. By analyzing instances of exposure to temperatures beyond the recommended 2–8 °C range, the study aims to identify the critical junctures within the vaccine supply chain susceptible to temperature excursions. The methodology entailed a systematic data logger analysis at each stage of transportation up to the last mile. In this study, descriptive statistics with the analysis of variance test has been applied to investigate the significance of variations in the duration of temperature breaches. The novelty of this study lies in its comprehensive examination of vaccine temperature integrity across critical transportation phases in India, utilizing real-time data-logging technology. Based on the results, the study provides recommendations using the ABCD strategic framework, encompassing awareness, best practices, continuous monitoring, and data management and documentation. This framework will be instrumental in addressing the identified challenges and significantly improving the temperature monitoring flexibility. The findings will inform targeted interventions, optimizing the cold chain to enhance the preservation of vaccine potency and identifying flexible solutions for maintaining the required temperatures from storage to transportation. [ABSTRACT FROM AUTHOR]

Objective(s): In this study, the adjuvant activity of aqueous and alcoholic extracts of propolis was examined on the inactivated herpes simplex virus-1 (HSV-1).Materials and Methods: BALB/C mice were administered with inactivated (HSV-1; the KOS strain) plus alcoholic and aqueous extracts, followed by assessment of the cellular and humoral immune responses. Results: Alcoholic and aqueous extracts, as an adjuvant, revealed a significant increase in lymphocyte proliferation and cytotoxic T lymphocyte (CTL) responses versus the HSV-1 group. In addition, HSV-1 plus alcoholic extract showed a remarkable increase in IFN-γ cytokine and IFN-γ/IL-4 ratio. On the other hand, both alcoholic and aqueous extracts in the HSV-1 vaccine suppressed the IL-4 cytokine response as compared with the HSV-1 vaccine. In addition, HSV-1 plus alcoholic extract showed a significant increment in IgG1, IgG2a, and IgG2b isotypes as compared with the HSV-1 vaccine.Conclusion: Propolis extracts seem to modulate the immune response against inactivated HSV-1 model and can be used as a suitable vaccine adjuvant or a component of a complex adjuvant against infectious diseases.

Analyzing vaccine stability under different storage and transportation conditions is critical to ensure that effectiveness and safety are not affected by distribution. In a simulation of the last mile in the supply chain, we found that shock and vibration had no effect on Ad26.ZEBOV/MVA-BN-Filo Ebola vaccine regimen quality under refrigerated conditions.

Background: Rabies, a potentially lethal virus, affects more than 150 countries. Although the rabies vaccine and immunoglobulin have been available since 1908, Bangladesh is new to vaccine manufacturing. We checked the quality of the local manufacturing rabies vaccine for substandard. Methods: The potency and immunogenicity of 20 vaccines were analyzed by three in vivo and in vitro methods from March 2020 to May 2023. Single radial immunodiffusion, fluorescent antibody virus neutralization, and national institutes of health tests were carried out to evaluate the vaccine's efficacy to provide sufficient protection against the rabies virus. Results: The potency of the rabies vaccine was determined by the in vitro SRID method by measuring glycoprotein content. An average of 16 articles from each batch was calculated. The minimum and maximum average mean values of the 20 batches were 5.058 and 5.346, respectively. The variance was calculated at 0.00566. We found a coefficient of variation (CV) between 9.36% and 14.80%. The 100% sample was satisfactory, as these samples had a potency of over 2.5 IU/mL. To observe immunogenicity, we applied the FAVN method for determining antibody titers. An average of 16 articles from every batch were counted to quantify antibody titers. The mean quantity of antibody titers ranged from 2.389 to 3.3875. The CV was slightly lower because of the dispersion of the data. At last, we performed an in vivo method, the NIH test method, to determine potency based on mortality rate. We found a mean value of 4.777 IU/SHD with a standard deviation of 1.13 IU/SHD. All 20 batches were found 100% satisfactory in the NIH test. Conclusion: The study implies that the rabies human vaccines manufactured in Bangladesh are potent enough to provide sufficient immunogenicity. Our research is warranted testimony for healthcare providers who work to extirpate rabies. [ABSTRACT FROM AUTHOR]

The BCG vaccines on the market have employed a Mycobacterium bovis (M. bovis) sub-strains derived from the initial strain. To date, there has been no recommendation regarding the sub-strains with the highest effectiveness when administered to humans. Because it remains the standard for Tuberculosis treatment, the quality of the BCG vaccine must be verified. The viability test is one of the parameters for BCG vaccine quality control. The culture method has become the gold standard for viability testing with various testing media. The present study aimed to evaluate the performance of Lowenstein Jensen (LJ) and Ogawa media for the viability test of Pasteur 1173P2 and Russian (Moscow) - 384 sub-strains of M. bovis in the BCG vaccine. The number of culturable particles of each sub-strain in the BCG vaccine was estimated and statistically evaluated using the t-test. The colonies of the Pasteur 1173P2 have characteristics; tended to clump on both mediums with tiny, rough, and pale yellow/cream colors. Although the colony character of the Russian (Moscow) - 384 generally has similar feature, it did not cluster and had a smooth texture. In terms of growth rate, LJ and Ogawa media performed similarly for Pasteur 1173P2 and Russian (Moscow) - 384 sub-strains. Maximum growth is reached by the fifth week. The culturable particles of Pasteur P1173P2 sub-strains did not differ between mediums. Whereas the growth of the Russian (Moscow) - 384 sub-strains was statistically better on Ogawa media. The results of this study reveal that the performance of the media used for determining the number of culturable particles is based on the sub-strains of M. bovis present in the BCG vaccine. [ABSTRACT FROM AUTHOR]

Foot and mouth disease is a highly contagious viral disease that poses a significant economic threat to cloven-hoofed animals, including cattle and sheep. The emergence of a novel foot and mouth disease virus-A isolate, FMDV-A-Egy-AHRI-RL385-Ven-2022, in Egypt in 2022 has raised concerns about its potential impact on existing vaccination programs. Given that vaccination is a key strategy for foot and mouth disease virus control, the present study was aimed to assess the cross-protective efficacy of both local and imported inactivated vaccines against this new threat. Through challenge experiments and serum neutralization tests, we observed limited effectiveness of both vaccine types. The calculated r1-values at 28 days post-vaccination indicated a minimal immune response to FMDV-A-Egy-AHRI-RL385-Ven-2022 (0.176 and 0.175 for local and imported vaccines, respectively). Challenge experiments further confirmed these findings, revealing 0% protection from the local vaccine and only 20% protection from imported vaccines by day 7 post-challenge. These results underscore the urgent need to update existing foot and mouth disease virus vaccines in Egypt by incorporating the newly circulating FMDV-A-Egy-AHRI-RL385-Ven-2022 strain. This proactive measure is crucial to prevent future outbreaks and ensure effective disease control.

The present work aims to establish a new alternative protocol to evaluate in vitro potency of inactivated Newcastle disease virus vaccine using Real Time PCR. Aqueous phases of seven inactivated Newcastle disease virus vaccines batches of different manufacturers were extracted by isopropyl myristate. The Newcastle disease virus antigen of each vaccine sample was determined by a standard Real Time PCR assay. Vaccines were inoculated into separate groups of 3-week-old specific pathogen free chickens using the recommended dose of vaccine. The immunogenicity was assessed for each vaccine by the Newcastle disease virus hemagglutination inhibition antibody titers. Individual serum samples were collected 4 weeks post vaccination, then vaccine efficacy and protection rates were recorded after challenge test of birds vaccinated with the virulent Newcastle disease virus. There is the possibility of using the Real Time PCR as an in vitro assay for vaccine evaluation. The Cycle Threshold values were ranged between 21.17 and 25.23. On the other hand, the hemagglutination inhibition titers ranged between 7.1 log2 to 6.2. The comparison between the Cycle Threshold values of the antigen extracts and the corresponding results of challenge test and in vivo hemagglutination inhibition assays using sera of vaccinated birds proved a strong correspondence between the in vitro and in vivo results.

Despite the great success of vaccines in various infectious diseases, most current vaccines are not effective enough, and on the contrary, clinically approved alum adjuvants cannot induce sufficient immune responses, including a potent cellular immune response to confer protection. In this study, we used Nanochelating Technology to develop novel nanoadjuvants to boost the potency of the alum-adjuvanted inactivated severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) vaccine. BALB/c mice were immunized twice over 2 weeks with different doses of adjuvanted-vaccine formulations and immune responses were assessed. The analysis results of IFN-γ and IL-17 cytokines demonstrated the effectiveness of the nanoadjuvants produced by the Nanochelating Technology in shifting the alum-based vaccine toward a stronger Th1 pattern. In addition, these nanoadjuvants improved IL-2 cytokine response, which shows the efficacy of these novel formulations in inducing specific T lymphocyte proliferation. Using these nanoadjuvants increased IL-10 cytokine secretion that may be representative of a better immunoregulatory impact and may also potentially prevent immunopathology responses. Moreover, specific IgG titer analysis revealed the potency of these nanoadjuvants in improving humoral immune responses. The enzyme-linked immunosorbent assay of receptor-binding domain (RBD)-specific IgG response showed that the developed novel formulations induced strong IgG responses against this protein. This study shows that the nanostructures produced by the Advanced Nanochelating Technology have potent adjuvant effects on alum-based SARS-CoV-2 vaccines to not only compensate for alum weakness in inducing the cellular immune responses by smart regulation of the immune system but also significantly improve the humoral and cellular immune responses simultaneously. [ABSTRACT FROM AUTHOR]

Classical Swine Fever (CSF) is still one of the most economically important viral diseases of pigs. The disease is controlled by vaccination in the endemic countries. Hence, availability or supply of efficacious and potent vaccine in the field settings is of utmost importance. Currently, as per requirement of any Pharmacopoea, a CSF vaccine must contain 100 PD 50 /dose which is determined by vaccinating pigs at 1/40th and 1/160th dilution of each dose followed by virulent challenge at 28 days post vaccination (dpv). Here, the control and the unprotected groups succumb to disease and need to be euthanized. Moreover, such challenge experiments are not feasible for each batch of the vaccine. In this communication, an alternate method of PD 50 dose calculation of live-attenuated CSF vaccines by measuring Serum Neutralizing Titre i. e Fluorescent Antibody Virus Neutralization (FAVN) titre of the vaccinated pigs at 28 dpv was established. This alternative method do not require the vaccinated pigs to be challenged. Serum samples, generated out of QC testing of eight batches of CSF vaccines in the laboratory, were tested and found that pigs having FAVN titre ≥10 were protected against challenge. Initially this test was optimized in serum samples of 12 animals and then validated with another 56 serum samples. It was found that the alternate method is 100% correlating with the challenge experiment. Thus, based on FAVN titre of the vaccinated animal serum, it can be predicted whether the pigs would or would not come through the challenge infection. Using the predicted status (protected/succumbed), PD50 can be calculated by applying Reed and Muench formula, hence alternate method can be used as routine QC test for potency of CSF vaccines. The newly developed assay was specific since no signal was observed in controls. • Fluorescent Antibody Virus Neutralization (FAVN) test is gold standard for measuring protective antibody titer of classical swine fever virus. • Animal challenge method of PD 50 determination does not follow 4R principle of animal use. • FAVN titer of vaccinated animal sera has been correlated with animal protection titer obtained by challenge method. • Significant agreement was found between two methods, supporting FAVN as alternate. • This test can replace the challenge experiment for PD50 determination of CSF vaccines. [ABSTRACT FROM AUTHOR]

Inactivated Polio Vaccines (IPV) and live Oral Polio Vaccine (OPV) were introduced in the mid-20th century, and their coordinated worldwide use led to almost complete elimination of the disease, with only one serotype of poliovirus remaining endemic in just two countries. Polio eradication will lead to discontinuation of OPV use and its replacement with IPV or other vaccines that are currently under development that will need to be tested in clinical trials. Despite decades of research, questions remain about the serological correlates of polio vaccine efficacy, specifically whether the vaccines are equally protective against immunologically different strains of the same serotype. The absence of significant morbidity does not allow use of a protection endpoint in clinical trials, so the answer could be obtained only by using surrogate markers such as immunogenicity. In this study, a panel of wild and vaccine-derived polioviruses of serotype 1 were tested in neutralization assays with sera from vaccine-immunized individuals. The results demonstrated that there was a significant difference in titers of neutralizing antibodies in human sera when measured against different strains. When measured with a homologous strain used for vaccine manufacture all subjects had detectable levels of antibodies, while neutralization tests with some heterologous strains failed to detect neutralizing antibodies in a number of subjects. Administration of a booster dose of IPV led to a significant increase in neutralizing titers against all strains. Results of the experiments using animal sera, performed to obtain more information on protectivity of neutralizing antibodies against heterologous strains, were consistent with the results obtained in the assays using human sera. These results are discussed in the context of serological biomarkers of protection against poliomyelitis, suggesting that potency of vaccines made from serologically different strains should be determined against both homologous and heterologous challenge viruses. [ABSTRACT FROM AUTHOR]

We compared 2019 influenza seasonality and vaccine effectiveness (VE) in four southern hemisphere countries: Australia, Chile, New Zealand and South Africa. Influenza seasons differed in timing, duration, intensity and predominant circulating viruses. VE estimates were also heterogeneous, with all-ages point estimates ranging from 7-70% (I2: 33%) for A(H1N1)pdm09, 4-57% (I2: 49%) for A(H3N2) and 29-66% (I2: 0%) for B. Caution should be applied when attempting to use southern hemisphere data to predict the northern hemisphere influenza season.

BACKGROUND:As a dominant seasonal influenza virus, H3N2 virus rapidly evolves in humans and is a constant threat to public health. Despite sustained research efforts, the efficacy of H3N2 vaccine has decreased rapidly. Even though antigenic drift and passage adaptation (substitutions accumulated during vaccine production in embryonated eggs) have been implicated in reduced vaccine efficacy (VE), their respective contributions to the phenomenon remain controversial. METHODS:We utilized mutational mapping, a powerful probabilistic method for studying sequence evolution, to analyze patterns of substitutions in different passage conditions for an unprecedented amount of H3N2 hemagglutinin sequences (n = 32 278). RESULTS:We found that passage adaptation in embryonated eggs is driven by repeated convergent evolution over 12 codons. Based on substitution patterns at these sites, we developed a metric, adaptive distance (AD), to quantify the strength of passage adaptation and subsequently identified a strong negative correlation between AD and VE. CONCLUSIONS:The high correlation between AD and VE implies that passage adaptation in embryonated eggs may be a strong contributor to the recent reduction in H3N2 VE. We developed a computational package called MADE (Measuring Adaptive Distance and vaccine Efficacy based on allelic barcodes) to measure the strength of passage adaptation and predict the efficacy of a candidate vaccine strain. Our findings shed light on strategies for reducing Darwinian evolution within the passaging medium in order to potentially restore an effective vaccine program in the future.

Several inactivated SARS-CoV-2 vaccines have been approved for human use, but are not highly potent. In this study, different formulations of the inactivated SARS-CoV-2 virus were developed in Alum, Montanide 51VG, and Montanide ISA720VG adjuvants, followed by assessment of immune responses. The SARS-CoV-2 virus was inactivated with formalin and formulated in the adjuvants. BALB/c mice were immunized subcutaneously with 4 μg of vaccines on days 0 and 14; (IL-4) and (IFN-g), cytotoxic T lymphocyte (CTL) activity, and specific immunoglobulin G (IgG) titer and IgG1, IgG2a, and IgG2a/IgG1 ratio, and anti-receptor-binding domain (RBD) IgG response were assessed 2 weeks after the final immunization. Immunization with SARS-CoV-2-Montanide ISA51VG showed a significant increase in the IFN-γ cytokine versus SARS-CoV-2-Alum, SARS-CoV-2-Montanide ISA720VG, and control groups (p < 0.0033). Cytokine IL-4 response in SARS-CoV-2-Alum group showed a significant increase compared with SARS-CoV-2-Montanide ISA51VG, SARS-CoV-2-Montanide ISA720VG, and control groups (p < 0.0206). In addition, SARS-CoV-2-Montanide ISA51VG vaccine induced the highest IFN-γ/IL-4 cytokine ratio versus other groups (p < 0.0004). CTL activity in SARS-CoV-2-Montanide ISA51VG and SARS-CoV-2-Montanide ISA720VG groups showed a significant increase compared with SARS-CoV-2-Alum and control groups (p < 0.0075). Specific IgG titer in SARS-CoV-2-Montanide ISA51 VG and SARS-CoV-2-Montanide ISA720VG showed a significant increase compared with SARS-CoV-2-Alum and control groups (p < 0.0143). Results from specific IgG1and IgG2a in SARS-CoV-2-Alum, SARS-CoV-2-Montanide ISA51VG, and SARS-CoV-2-Montanide ISA720VG vaccine showed a significant increase compared with phosphate buffer saline (PBS) group (p < 0.0001), but SARS-CoV-2-Montanide ISA51VG and SARS-CoV-2-Montanide ISA 720VG groups showed the highest IgG2a/IgG1 ratio and a significant increase compared with SARS-CoV-2-Alum group (p < 0.0379). Moreover, inactivated SARS-CoV-2+Alum and SARS-CoV-2-Montanide ISA 720VG groups demonstrated a significant increase in anti-RBD IgG response versus the SARS-CoV-2-Montanide ISA51VG group. It seems that the type of vaccine formulation is a critical parameter, influencing the immunologic pattern and vaccine potency and human-compatible oil-based adjuvants were more potent than Alum adjuvant in the vaccine formulation. [ABSTRACT FROM AUTHOR]

Foot and mouth disease virus is a highly infectious and contagious pathogen. Recently the topotype VII, Lib‐12 lineage of serotype SAT2 was reported through outbreaks in Egypt during 2018. Vaccination is an effective way to control and combat the foot and mouth disease virus outbreaks especially in endemic areas like Egypt. The present study was aimed to evaluate the efficacy of the current produced foot and mouth disease vaccine, against the recently isolated field strain foot and mouth disease virus SAT2 topotype VII, Lib-12 lineage (SAT2 Libya), by applying in vitro and in vivo studies. Two batches of the current foot and mouth disease virus vaccine were inoculated in calves. At the 28th day post-vaccination serum samples were collected and tested against tissue culture adapted foot and mouth disease virus SAT2 Libya and SAT2/EGY/2/2012 using virus neutralization test to determine serological relationship (r1-value). The challenge test for vaccinated calves was carried out against the virulent foot and mouth disease virus SAT2 Libya. It was found that neutralizing antibody titers induced by the two vaccine batches (1 and 2) and those in unvaccinated animals were 0.48, 0.39 and 0.15 log10 TCID50/mL, respectively, while the challenge revealed protection values of 20%, 0% and 0%, respectively. Furthermore, the r1 values were 0.195 and 0.186 for vaccine batches (1 and 2), respectively. It was concluded that the available local commercial inactivated foot and mouth disease virus vaccine batches (SAT2 SAT2/EGY/2/2012) are unable to protect calves against the current circulating foot and mouth disease virus field isolate SAT2 topotype VII, Lib-12 lineage, thus it is highly recommended to update the existing vaccines with the present isolated strain.

Animal (in vivo) potency tests have been utilized for over a century in support of vaccine development and for quality testing. This is a legacy of the best science at the time of their introduction. Advances in knowledge and technology, however, have provided opportunities to utilize more sensitive assays during development and replace legacy animal tests with in vitro alternatives. This coupled with initiatives such as replacement, reduction, and refinement (the 3-R's) and quality by design (QbD) have brought industry and regulators together in the introduction of advanced vaccine control strategies. This article examines historical and current uses of animals in vaccines technical development and control, and their replacement with in vitro alternatives from a risk point of view. An overarching risk is that a vaccine tested with an alternative potency assay fails to protect its target recipient. This can be addressed from the perspective of the assay's association with the vaccine mechanism of action, and the rules used to introduce the vaccine into the patient population (e.g., specifications). Commonly understood concepts such as analytical precision play a role in risk evaluation based on its impact on the sensitivity of a test to detect meaningful product changes caused by variations in manufacture or over a vaccine's shelf life. This should be considered when evaluating solutions such as the reduction of multi-concentration (or dilution) in vivo assays to a single concentration test. While the use of animals in vaccine development will not go away all together, the paradigm must shift from in vivo tests to in vivo models. To help ensure success, principles and practices related to introduction of in vitro alternatives require global collaboration among industry, regulators, pharmacopeias, and supporting organizations., (Copyright © 2024 International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.)

Due to the genetic diversity of low pathogenic avian influenza (LPAI) viruses of subtype H9N2, it deemed appropriate to study the potency of the vaccines based on the antigens of strains А/chicken/Amursky/03/12 and A/chicken/Chelyabinsk/314-1/20 that represent currently circulating in the Russian Federation genetic lineages Y280 and G1, respectively. While low pathogenicity of the agent does not allow demonstrating the vaccine protective properties by the direct methods generally used for potency assessment (e.g. morbidity and mortality), the indirect methods were used: determination of antigenic relatedness of the strains, level of the postvaccinal homologous and heterologous humoral immunity, analysis of the virus genome synthesis inhibition (reduction) in vaccinated birds following their challenge. The strains used in the vaccines were determined to have some antigenic differences, which were demonstrated in the hemagglutination inhibition (HI) assay during control of the postvaccinal immunity in birds. Both vaccines generally induced strong humoral immunity in vaccinated birds (9–10 log2 determined using HI assay) with some difference in the levels of the immune response following the use of homologous or heterologous antigens. It was also reliably determined that homologous immunity facilitated more expressed inhibition of the virus reproduction after the challenge. The level of inhibition (reduction) of the virulent LPAI virus genome synthesis in vaccinated birds following their challenge with H9N2 virus of genetic lineage G1 was higher in birds following homologous vaccination, while the time periods of the genome detection in the biomaterial samples were the same. It was demonstrated that due to antigenic and immunogenic differences between LPAI H9N2 strains, use of both antigenic components in the inactivated vaccines is appropriate.

In 2017, influenza seasonal activity was high in the southern hemisphere. We present interim influenza vaccine effectiveness (VE) estimates from Australia. Adjusted VE was low overall at 33% (95% confidence interval (CI): 17 to 46), 50% (95% CI: 8 to 74) for A(H1)pdm09, 10% (95% CI: -16 to 31) for A(H3) and 57% (95% CI: 41 to 69) for influenza B. For A(H3), VE was poorer for those vaccinated in the current and prior seasons.

In Foot-and-mouth disease (FMD) enzootic countries, periodic vaccination is the key tool in controlling the disease incidence. Active seromonitoring of the vaccinated population is critical to assess the impact of vaccination. Virus neutralization test (VNT) and enzyme-linked immunosorbent assays (ELISA) are commonly used for antibody detection. Assays like liquid phase blocking ELISA (LPBE) or solid phase competition ELISA (SPCE) are preferred as they do not require handling of live FMDV and are routinely used for seromonitoring or for vaccine potency testing; however, false positives are high in LPBE. Here we report, a monoclonal antibody (mAb) based SPCE as a potential alternate assay for antibody titration. From a panel of 12 mAbs against FMDV serotype A, two mAbs were chosen for the development of SPCE. Based on a set of 453 sera, it was demonstrated that mAb 2C4G11, mAb 6E8D11and polyclonal antibody (pAb) based SPCE had a relative sensitivity of 86.1, 86.1 and 80.3 %; and specificity of 99.6, 99.1 and 99.1 %, respectively. The correlation, repeatability, and level of agreement of the assays were high demonstrating the potential use of mAb in large scale surveillance studies and regular vaccine potency testing. • mAb ELISA was applied for detection of antibodies to FMDV serotype A. • mAb based SPCE results correlated with polyclonal antibody (pAb) based assay. • mAb and pAb based assays had relative sensitivity of 86 % and 80 %, respectively. • Both the assays showed relative specificity of 99 % on large scale evaluation. • The assay has a greater potential in surveillance and vaccine potency studies. [ABSTRACT FROM AUTHOR]

GSK is currently working to improve the commercial presentation of the licensed quadrivalent conjugate vaccine (Menveo) for use against meningococcal serogroup A, C, W, Y (MenACWY) infections. Menveo consists of a primary, lyophilized vial, containing the serogroup A antigen that is reconstituted with the content of a second, liquid, vial that contains the serogroup C, W, Y antigens, to give the final liquid MenACWY product. Since the MenA structure is prone to hydrolytic degradation in liquid formulations, we used mathematical models to rationally design a clinical Phase 2 development plan and provide end of shelf-life (EoSL) and release specification setting for the MenACWY liquid product. By using development and clinical stability data, statistical models were built and used to predict both the MenA free saccharide (FS) and O-Acetyl (OAc) content during long-term storage conditions at 5 °C and stressed (accelerated) stability studies at 15 °C, 22.5 °C, 25 °C, 37 °C and 50 °C. This approach allowed us to define an aging plan for the clinical material to reach at least the required levels of MenA FS and OAc levels at product EoSL. The clinical material was then exposed to a temperature of 22.5 ± 2.5 °C for 59 days to generate FS OAc content of about 35% and 40%, respectively, which was then delivered to the patients in the clinical trial. To the best of our knowledge, this work represents the first example in the field of vaccine research where statistical models have been used to rationally design tailored lots, with the goal of setting EoSL and release specification limits based on data collected on artificially aged clinical material, in which the FS and OAc levels tested were intended to support a product shelf-life of at least 24 months. [ABSTRACT FROM AUTHOR]

Several glycoconjugate vaccines have been licensed or are currently in clinical development to prevent bacterial infections. Here we report the development of a single analytical assay to quantify the conjugated saccharide content, as alternative to two separated total and free (unconjugated) saccharide assays used so far, for a quadrivalent conjugate vaccine containing meningococcal serogroup A polysaccharide (α-1,6-linked N-acetylmannosamine phosphate repeating unit partly O-acetylated at position C 3 or C 4) coupled with CRM 197 protein. The results confirm a high linear correlation among the two approaches (conjugated saccharide content vs. difference of total saccharide and free saccharide). Conjugated saccharide content estimation is therefore demonstrated to be a suitable method to monitor the product quality of vaccines containing meningococcal serogroup A conjugate antigen, in the final filled presentation as demonstrated here and potentially on the bulk conjugate before formulation. [ABSTRACT FROM AUTHOR]

Foot and mouth disease is a highly contagious viral disease of cloven-hoofed animals that has a significant economic impact on livestock. A recent outbreak was detected and recorded as exotic strain of foot and mouth disease virus SAT2 (Serotype SAT2, topotype VII, Lib-12 lineage). The emergency vaccine was produced and assessed in vivo and large number of vaccine batches were urgently needed. The present work was aimed to provide a rapid evaluation of inactivated foot and mouth disease SAT2 oily vaccine to exclude the unsatisfactory batches during emergency circumstances and to reduce time, effort and cost. The extraction of foot and mouth disease antigen content from oily adjuvanted vaccine was carried out using isopropyl myristate and benzyl alcohol methods. The extracted viral antigen was identified by foot and mouse disease serotyping ELISA and 146S content was quantified using sucrose density gradient analysis. Evaluations were carried out instantly and at 2h, 6h and 24h. The results indicated the efficiency of benzyl alcohol to breakdown the oil emulsion either MONTANIDE™ ISA 206 VG or MONTANIDE™ ISA 50 V2, while the isopropyl myristate was efficient for MONTANIDE™ ISA 50 V2 only. The identification and quantification of 146S for extracted antigen using benzyl alcohol indicated significant stable records at different time intervals for the vaccine batches, while the extraction using isopropyl myristate indicated unstable records at different time intervals. It was concluded that the evaluation of monovalent foot and mouse disease vaccine could be conducted in vitro, using serotyping ELISA and quantification of 146S for the extracted antigen, either using benzyl alcohol or isopropyl myristate (MONTANIDE™ ISA50 V2 only), with the consideration that 146S content should not less than 4 μg/mL.

This study assessed three Ad26.RSV.preF/RSV preF protein combinations, combining different Ad26.RSV.preF doses and naturally aged preF protein, representing the expected critical vaccine quality attributes close to release, around intermediate shelf-life (ISL) and near-presumed end-of-shelf-life (EoSL), as a way to evaluate the vaccine immunogenicity and safety throughout its shelf-life. A single dose of Ad26.RSV.preF/RSV preF protein vaccine was administered to adults 60-75 years of age. Solicited adverse events (AEs), unsolicited AEs, and serious AEs (SAEs) were assessed for 7-day, 28-day, and 6-month periods after vaccination, respectively. RSV preF-binding antibody concentrations and RSV neutralizing titers were measured 14 days post-vaccination as primary and secondary endpoints, respectively; binding antibodies were also measured 6 months post-vaccination. The RSV preF-binding antibody responses induced by Ad26.RSV.preF/RSV preF protein vaccine lots representing the critical quality attributes around ISL and near presumed EoSL were noninferior to the responses induced by the vaccine lot representing the critical quality attributes near release. The RSV preF-binding and RSV neutralizing antibody levels measured 14 days post-vaccination were similar across the 3 groups. RSV preF-binding antibody concentrations were also similar 6 months post-vaccination. Solicited AEs were mostly mild to moderate in intensity, and a decreased reactogenicity was observed from the Release group to the ISL and EoSL group. None of the reported SAEs were considered related to study vaccination. The study provided evidence of sustained immunogenicity and safety over the intended shelf-life of the Ad26.RSV.pref/RSV preF protein vaccine. The three vaccine lots had acceptable safety profiles.

Background: Potency evaluation of rabies vaccine is a cheap, fast, high precision and consistent with ethical values is critical, so researchers have modified a variety of methods such as: National Institute of Health (NIH) method, Single Radial Immunodiffusion (SRID) and so on. The purpose of the present study was to replace an in vitro method consistent with medical ethics criteria instead of an in vivo method. By recognizing that the potency of the rabies vaccine depends on the amount of glycoprotein antigen content and the monoclonal antibody detect the correct folding of antigen of the rabies virus, then the glycoprotein content could be represent of vaccine potency. Methods: In this study, we designed an immune-capture enzyme-linked immunosorbent assay (ELISA) with three antibodies (capture, primary and secondary) to determine the existent amount of viral glycoprotein in different rabies vaccines, and compared the results at the same time with measuring potency of those vaccines using the NIH method. This applied study was conducted from September 2016 to September 2018 at the Research Laboratory of the World Health Organization Collaborating Center for reference and research on rabies at the Pasteur Institute of Iran in Tehran. Results: The slope of the standard line was calculated to R2=0.98 (P=0.0013). In the humans’ vaccines, the mean lied between 5.554-7.336 (SD=0.0463-0.1039) and the coefficient of variation was 0.778-2.436 (SD=0.0041-0.2724), at the same time in the animals’ vaccines the mean were 2.293-5.993 (SD=0.0041-0.2724) and the coefficient of variation was calculated 0.182-4.546. For animal vaccines the Pearson correlation coefficient is 0.99 and for the human vaccines this coefficient was 0.95. Also, the concordance correlation coefficient for animal vaccines was 0.98 and for human vaccines is 0.95, indicating a moderate to high concordance in both animals and humans vaccines. Conclusion: The designed Immuno-capture ELISA kit had a proper acceptance criterion, intermediate precision, good linearity and robustness for measuring the glycoprotein level of the vaccine, which was directly related to the vaccine potency.

Objectives: Influenza is associated with significant morbidity and mortality, especially in older and immunocompromised patients. Few data are available on the clinical benefit of high dose trivalent influenza vaccine (TIV). We aimed to assess the clinical efficacy and safety of high dose TIV.Methods: We conducted a systematic review and meta-analysis of randomized controlled trials (RCTs), evaluating high dose versus standard dose TIV for prevention of seasonal influenza in adult population. Primary outcome was laboratory-confirmed influenza. Subgroups analyses included older adults and immunocompromised patients.Results: We included 16 trials, 47,857 patients; 10 included older adults and three immunocompromised patients. Laboratory confirmed influenza was significantly reduced with high dose TIV (relative risk 0.76, 95% confidence interval 0.64 to 0.9). This outcome stemmed mainly from one trial in older adults. Specifically, A(H3N2) laboratory confirmed influenza, but not A(H1N1) or B lineages, was reduced. No difference in mortality or hospitalizations was demonstrated. Immunological response was significantly higher with high dose vaccine. Serious adverse events were significantly less common in the high dose group.Conclusions: High dose TIV lowers the rates of laboratory confirmed influenza, mainly A (H3N2), in older adults vs. standard dose. Further studies should address immunocompromised patients and report clinical outcomes. [ABSTRACT FROM AUTHOR]

BackgroundThe Shingles Prevention Study (SPS) demonstrated zoster vaccine efficacy through 4 years postvaccination. A Short-Term Persistence Substudy (STPS) demonstrated persistence of vaccine efficacy for at least 5 years. A Long-Term Persistence Substudy (LTPS) was undertaken to further assess vaccine efficacy in SPS vaccine recipients followed for up to 11 years postvaccination. Study outcomes were assessed for the entire LTPS period and for each year from 7 to 11 years postvaccination.MethodsSurveillance, case determination, and follow-up were comparable to those in SPS and STPS. Because SPS placebo recipients were offered zoster vaccine before the LTPS began, there were no unvaccinated controls. Instead, SPS and STPS placebo results were used to model reference placebo groups.ResultsThe LTPS enrolled 6867 SPS vaccine recipients. Compared to SPS, estimated vaccine efficacy in LTPS decreased from 61.1% to 37.3% for the herpes zoster (HZ) burden of illness (BOI), from 66.5% to 35.4% for incidence of postherpetic neuralgia, and from 51.3% to 21.1% for incidence of HZ, and declined for all 3 outcome measures from 7 through 11 years postvaccination. Vaccine efficacy for the HZ BOI was significantly greater than zero through year 10 postvaccination, whereas vaccine efficacy for incidence of HZ was significantly greater than zero only through year 8.ConclusionsEstimates of vaccine efficacy decreased over time in the LTPS population compared with modeled control estimates. Statistically significant vaccine efficacy for HZ BOI persisted into year 10 postvaccination, whereas statistically significant vaccine efficacy for incidence of HZ persisted only through year 8.

Analyzing vaccine stability under different storage and transportation conditions is critical to ensure that effectiveness and safety are not affected by distribution. In a simulation of the last mile in the supply chain, we found that shock and vibration had no effect on Ad26.ZEBOV/MVA-BN-Filo Ebola vaccine regimen quality under refrigerated conditions.

Aim: The present study was aimed to establish a protocol for the evaluation of the in vitro potency of commercial inactivated Newcastle disease virus (NDV) oil-adjuvanted vaccines using hemagglutination test (HA) and blocking ELISA (B-ELISA) based on polyclonal antibodies. Materials and Methods: Aqueous phases from a total of 47 batches of inactivated NDV vaccines manufactured by 20 different companies were extracted with isopropyl myristate. The viral antigen in each sample was detected and quantified by a standard HA test and a B-ELISA assay. To verify the efficiency of the antigen extraction method used in the batches which showed HA and to test the validity of using in vitro antigen quantification by HA and B-ELISA tests, a subset of 13 batches (selected from the total 47 batches) was inoculated in groups of 3-4-week-old specific pathogen-free chickens using the recommended vaccine dose. The immunogenicity of the selected vaccine batches was assessed by the NDV-hemagglutination inhibition antibody titers in individual serum samples collected 4 weeks after vaccination. Further, the efficacy of the vaccines and their protection rates were determined by a challenge test carried out for the vaccinated chickens with the Egyptian 2012 isolate of the virulent NDV genotype VII. Results: A strong correlation was observed between HA titers and B-ELISA mean titers in the tested 47 batches (R2=0.817). This indicated the possibility of using the latter in vitro assays for vaccine potency assessment. The recommended protective NDV antigen titer measured by B-ELISA was determined to be 28 ELISA units per dose. The comparison between the HA titers of the aqueous extracts of test vaccines and the corresponding results of in vivo potency assays (i.e., immunogenicity and efficacy), including antibody titers in the serum of vaccinated birds, indicated that the efficiency of the antigen extraction used may interfere with obtaining a strong correlation between the in vitro and in vivo results. Conclusion: HA or B-ELISA tests can be used as rapid and cost-effective alternatives to traditional in vivo potency tests for vaccine potency assessment by quantifying the NDV antigen present in aqueous phase extracts of the tested vaccines. The latter in vitro protocol, however, requires efficient extraction of the antigen to be able to obtain good correlation with the traditional in vivo potency tests.

Replacement of the potency tests for diphtheria vaccines is a high priority for the international initiative to reduce, refine, and replace animal use in vaccine testing. Diphtheria toxoid containing vaccine products marketed in the US currently require potency testing by the United States Public Health Service (USPHS) test, which includes an in vivo passive protection test with a diphtheria toxin challenge. Here we describe an in vitro Diphtheria Vero Cell (DVC) assay which combines the immunization approach from the USPHS test and the use of a cell based neutralization assay for serological testing of vaccine potency. The DVC assay reduces the overall number of animals used compared to other serological potency tests and eliminates the in vivo toxin challenge used in the US test. The DVC assay can be used to test vaccine products with a low or high diphtheria toxoid dose. It has been optimized and validated for use in a quality control testing environment. Results demonstrate similar sera antibody unitage as well as agreement between the serum neutralization values determined using the USPHS test and the DVC assay and thus support the use of the DVC assay for routine and stability testing for diphtheria toxoid containing vaccine products. [ABSTRACT FROM AUTHOR]

In Japan, the practical application of completely cell-based seasonal influenza vaccines is under consideration. Considering the good correlation between the immunogenicity of egg-based influenza vaccines and the hemagglutinin (HA) content determined by the single radial immunodiffusion (SRD) assay, we determined the potency of the first cell-based quadrivalent vaccine experimentally generated in Japan using the SRD assay in this study. A primary liquid standard (PLS) and reference antigen were generated from the purified vaccine virus, and a sheep antiserum was produced against the HA of the vaccine virus. Since the purity of the PLS affects the reliability of vaccine potency testing, the purification steps are significant. We successfully prepared a purified PLS nearly free of cell debris. The HA content in the PLS was first estimated from the total amount of viral protein and the percentage of HA content determined by SDS-PAGE analysis. The HA content in the reference antigen was calibrated to that in the PLS via the SRD assay. The vaccine potency, that is, the HA content in each vaccine, was finally measured using the corresponding reference antigen. Ultimately, the measured vaccine potency of the monovalent vaccine was similar to that of the quadrivalent vaccine. [ABSTRACT FROM AUTHOR]

Background: Potency evaluation of rabies vaccine is a cheap, fast, high precision and consistent with ethical values is critical, so researchers have modified a variety of methods such as: National Institute of Health (NIH) method, Single Radial Immunodiffusion (SRID) and so on. The purpose of the present study was to replace an in vitro method consistent with medical ethics criteria instead of an in vivo method. By recognizing that the potency of the rabies vaccine depends on the amount of glycoprotein antigen content and the monoclonal antibody detect the correct folding of antigen of the rabies virus, then the glycoprotein content could be represent of vaccine potency. Methods: In this study, we designed an immune-capture enzyme-linked immunosorbent assay (ELISA) with three antibodies (capture, primary and secondary) to determine the existent amount of viral glycoprotein in different rabies vaccines, and compared the results at the same time with measuring potency of those vaccines using the NIH method. This applied study was conducted from September 2016 to September 2018 at the Research Laboratory of the World Health Organization Collaborating Center for reference and research on rabies at the Pasteur Institute of Iran in Tehran. Results: The slope of the standard line was calculated to R2=0.98 (P=0.0013). In the humans’ vaccines, the mean lied between 5.554-7.336 (SD=0.0463-0.1039) and the coefficient of variation was 0.778-2.436 (SD=0.0041-0.2724), at the same time in the animals’ vaccines the mean were 2.293-5.993 (SD=0.0041-0.2724) and the coefficient of variation was calculated 0.182-4.546. For animal vaccines the Pearson correlation coefficient is 0.99 and for the human vaccines this coefficient was 0.95. Also, the concordance correlation coefficient for animal vaccines was 0.98 and for human vaccines is 0.95, indicating a moderate to high concordance in both animals and humans vaccines. Conclusion: The designed Immuno-capture ELISA kit had a proper acceptance criterion, intermediate precision, good linearity and robustness for measuring the glycoprotein level of the vaccine, which was directly related to the vaccine potency. [ABSTRACT FROM AUTHOR]

Background: Following the declaration of wild-type 2 poliovirus eradication in 2015, the type 2 component was removed from the live-attenuated oral polio vaccine (OPV). This change implies a need to improve global coverage through routine immunization with inactivated polio vaccine (IPV), to ensure type 2 immunity. Several manufacturers use Sabin OPV strains for IPV production (sIPV), rather than the usual wild-type strains used for conventional IPV (cIPV). However, in contrast to cIPV, potency assays for sIPV have not been standardized, no international references exist, and no antigen units have been defined for a sIPV human dose. Thus, sIPV products from different manufacturers cannot be compared, and the relationship between antigenicity and immunogenicity of sIPV is not well understood.Methods: A collaborative study was conducted in which laboratories used different methods to measure the antigen content of a set of sIPV and cIPV samples with an aim to identify a suitable reference for sIPV products.Results: The study revealed differences in the reactivity of antibody reagents to cIPV and sIPV products.Conclusions: Homologous references are required to measure the antigen content of IPV products consistently. The first World Health Organization international standard for sIPV was established, with new, specific Sabin D-antigen units assigned. [ABSTRACT FROM AUTHOR]

The development of combined vaccines is a very useful project for all the advantages that this type of vaccine provides. However, complexities in its execution due to their technological challenges make difficult its development. In this sense, the pentavalent vaccine (DPT-HB-Hib), branded as Heberpenta®, has been produced in Cuba containing five antigens: diphtherial and tetanus anatoxins, whole cells of Bordetella pertussis, recombinant hepatitis B virus surface antigen and the synthetic polysaccharide (polyribosyl ribitol phosphate, PRP) of Haemophilus influenzae conjugated with tetanus anatoxin (PRP-T). A pre-formulation study was carried out using a mixture of adjuvants defining the optimum absorption time for each antigen. Three lots formulated at a pilot scale with a defined technology were studied using different physical-chemical and biological tests. The technology for the formulation of the pentavalent vaccine produced in Cuba was defined according to these results, which must later undergo stability studies and preclinical and clinical studies. [ABSTRACT FROM AUTHOR]

Background and Objectives: In recent years, due to the eradication of type 2 poliovirus, a bivalent vaccine containing types 1 and 3 is used in Iran. Since it is a highly thermolabile vaccine, it should be stored at a recommended temperature. Since cold chain may not be applied in Iran's tropical weather conditions, potency of the vaccine may be subjected to change. Therefore, we evaluated the effective factors on the potency of this vaccine. Methods: We evaluated stability of a bivalent oral poliovirus vaccine produced by Razi Institute of Iran to ensure consistency of virus at different temperatures (-20, 2-8, 22-25 and 35-37 °C), time intervals and freeze/thaw cycles. Three consecutive batches produced during full-scale production were randomly sampled. Results: At -20 °C, there was no change in vaccine vial monitor (VVM). The potency of the samples exposed to 2-8 oC for 60 days and to 22-25 oC for five days met the specification. The mean potency of the samples was 6.17, 6.00, 5.83, 5.75 and 5.54 (log CCID50/dose) after 10, 20, 30, 40 and 50 freeze/thaw cycles, respectively. In addition, the mean degradation of VVM was 0, 23.33, 60 and 100% for samples exposed to -20, 2-8, 22-25 and 35-37 oC, respectively. Conclusion: The results indicate the effects of environmental factors on the potency of the vaccines and the correlation between the VVM grade, color change and the vaccine potency for programming vaccine distribution networks at different transit levels. Based on the results of our study, the best temperature for maintenance and transportation of bivalent oral poliovirus vaccine is -20 oC. Furthermore, freeze/thaw cycles lower the potency of the vaccine and change the VVM grade significantly. [ABSTRACT FROM AUTHOR]

• Potency of agitated and static Ebolavirus vaccine samples were similar for 7 days at 2–8 °C. • These results suggest no impact to the Ebolavirus vaccine potency from agitation. • The Ebolavirus vaccine may be transported at 2–8 °C where transport at −70 °C is challenging. The Ebolavirus vaccine (rVSVΔG-ZEBOV-GP) is stored at −80 to −60 °C and should be kept frozen for transport. Due to significant logistical challenges, frozen transport is not feasible for some remote locations. To determine if local distribution at 2–8 °C is a potential option for these locations, a study was conducted to evaluate the impact of agitation on the thawed vaccine. After up to 7 days of constant agitation, no impact on vaccine potency was evident for the agitated vaccine versus the corresponding vaccine kept stationary at 2–8 °C. In conclusion, in-country transport of the Ebolavirus vaccine, rVSVΔG-ZEBOV-GP, at 2–8 °C appears to be a feasible option for those remote locations where significant logistical challenges prohibit transporting the vaccine at −80 to −60 °C. [ABSTRACT FROM AUTHOR]

Aim: The present study was aimed to establish a protocol for the evaluation of the in vitro potency of commercial inactivated Newcastle disease virus (NDV) oil-adjuvanted vaccines using hemagglutination test (HA) and blocking ELISA (B-ELISA) based on polyclonal antibodies. Materials and Methods: Aqueous phases from a total of 47 batches of inactivated NDV vaccines manufactured by 20 different companies were extracted with isopropyl myristate. The viral antigen in each sample was detected and quantified by a standard HA test and a B-ELISA assay. To verify the efficiency of the antigen extraction method used in the batches which showed HA and to test the validity of using in vitro antigen quantification by HA and B-ELISA tests, a subset of 13 batches (selected from the total 47 batches) was inoculated in groups of 3-4-week-old specific pathogen-free chickens using the recommended vaccine dose. The immunogenicity of the selected vaccine batches was assessed by the NDVhemagglutination inhibition antibody titers in individual serum samples collected 4 weeks after vaccination. Further, the efficacy of the vaccines and their protection rates were determined by a challenge test carried out for the vaccinated chickens with the Egyptian 2012 isolate of the virulent NDV genotype VII. Results: A strong correlation was observed between HA titers and B-ELISA mean titers in the tested 47 batches (R2=0.817). This indicated the possibility of using the latter in vitro assays for vaccine potency assessment. The recommended protective NDV antigen titer measured by B-ELISA was determined to be 28 ELISA units per dose. The comparison between the HA titers of the aqueous extracts of test vaccines and the corresponding results of in vivo potency assays (i.e., immunogenicity and efficacy), including antibody titers in the serum of vaccinated birds, indicated that the efficiency of the antigen extraction used may interfere with obtaining a strong correlation between the in vitro and in vivo results. Conclusion: HA or B-ELISA tests can be used as rapid and cost-effective alternatives to traditional in vivo potency tests for vaccine potency assessment by quantifying the NDV antigen present in aqueous phase extracts of the tested vaccines. The latter in vitro protocol, however, requires efficient extraction of the antigen to be able to obtain good correlation with the traditional in vivo potency tests. [ABSTRACT FROM AUTHOR]

Objectives: To circumvent the limitations of the current golden standard method, colony-forming unit (CFU) assay, for viability of Bacille Calmette-Guérin (BCG) vaccines, we developed a new method to rapidly and accurately determine the potency of BCG vaccines. Methods: Based on flow cytometry (FACS) and fluorescein diacetate (FDA) as the most appropriate fluorescent staining reagent, 17 lots of BCG vaccines for percutaneous administration and 5 lots of BCG vaccines for intradermal administration were analyzed in this study. The percentage of viable cells measured by flow cytometry along with the total number of organisms in BCG vaccines, as determined on a cell counter, was used to quantify the number of viable cells. Results: Pearson correlation coefficients of FACS and CFU assays for percutaneous and intradermal BCG vaccines were 0.6962 and 0.7428, respectively, indicating a high correlation. The coefficient of variation value of the FACS assay was less than 7%, which was 11 times lower than that of the CFU assay. Conclusion: This study contributes to the evaluation of new potency test method for FACS-based determination of viable cells in BCG vaccines. Accordingly, quality control of BCG vaccines can be significantly improved. [ABSTRACT FROM AUTHOR]