Hepatitis B Surface Antigen Concentration Research Articles

Mathematical Models of Early Hepatitis B Virus Dynamics in Humanized Mice

Analyzing the impact of the adaptive immune response during acute hepatitis B virus (HBV) infection is essential for understanding disease progression and control. Here we developed mathematical models of HBV infection which either lack terms for adaptive immune responses, or assume adaptive immune responses in the form of cytolytic immune killing, non-cytolytic immune cure, or non-cytolytic-mediated block of viral production. We validated the model that does not include immune responses against temporal serum hepatitis B DNA (sHBV) and temporal serum hepatitis B surface-antigen (HBsAg) experimental data from mice engrafted with human hepatocytes (HEP). Moreover, we validated the immune models against sHBV and HBsAg experimental data from mice engrafted with HEP and human immune system (HEP/HIS). As expected, the model that does not include adaptive immune responses matches the observed high sHBV and HBsAg concentrations in all HEP mice. By contrast, while all immune response models predict reduction in sHBV and HBsAg concentrations in HEP/HIS mice, the Akaike Information Criterion cannot discriminate between non-cytolytic cure (resulting in a class of cells refractory to reinfection) and antiviral block functions (of up to 99%\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$99\\%$$\\end{document} viral production 1–3 weeks following peak viral load). We can, however, reject cytolytic killing, as it can only match the sHBV and HBsAg data when we predict unrealistic levels of hepatocyte loss.

Accuracy of Saliva as a Diagnostic Medium for Hepatitis B Virus Infection: A Quantitative ELISA Analysis

Context: Diagnosis of hepatitis B virus infection (HBV) has been traditionally serum-based with its attendant risks and invasive procedure. Massive research interests are being directed to the use of saliva for screening, diagnosis, and monitoring of several infectious diseases, including HBV. Various reports suggest that saliva contains biomarkers that are comparable with that of sera. It can also be obtained with minimal risk to the clinician and patient as well. Aim: To determine the diagnostic accuracy of saliva in quantitative detection of hepatitis B surface antigen (HBsAg) using enzyme-linked immunosorbent assay (ELISA). Settings and Design: This cross-sectional analytical study was performed on HBV seropositive patients at the Gastroenterology clinic of the University of Benin Teaching Hospital, Benin City, Edo State. Materials and Methods: Under standardized conditions, equal amounts of blood and saliva samples of 43 HBsAg seropositive patients were analyzed using ELISA to quantitatively detect the concentration of HBsAg. Statistical Analysis Used: The analysis relied on Spearman’s correlation coefficient, linear regression analysis, and Bland–Altman plots to describe the correlational, predictive, and agreements between measurements of HBsAg in sera and saliva. The statistical significance was set at P < 0.05, while a 95% confidence level was used to construct intervals. Results: All participants had detectable levels of HBsAg in both saliva and serum with mean titers of 1.70 ± 0.35 ng/ml and 2.80 ± 0.77 ng/ml, respectively. Correlational and linear regression showed poor fit and predictive relationships of the HBsAg levels. Bland–Altman analysis showed good agreement and no significant bias in their diagnostic agreements. Conclusion: Saliva can be reliably used in screening and diagnosis of HBV infection. There was agreement in their levels independently and within their averages. It may be premature to rely on saliva for quantitative assay of HBsAg in treatment monitoring.

Highly Sensitive Hepatitis B Virus Identification by Antibody-Aptamer Sandwich Enzyme-Linked Immunosorbent Assay

Hepatitis B virus (HBV) infection is considered a major global health problem, causing various health issues, including cirrhosis, hepatitis, and liver cancer. The risk of developing such complications increases when the viral load is above 105 copies/mL. Early identification of HBV infection is imperative to preventing the spread of infection to other parts of the body. Although various sensing methods have been developed to identify HBV, researchers are still working toward developing cheap, easy, and sensitive detection methods. We developed a highly sensitive and rapid HBV detection method using nanomaterials on enzyme-linked immunosorbent assay (ELISA). Aptamer-antibody was utilized as the detection probe and immobilized on a zeolite-modified ELISA plate to detect the HBV biomarker hepatitis B surface antigen (HBsAg). To enhance the detection of HBsAg, aptamer and antibody were attached to gold nanoparticle through electrostatic interaction and immobilized on the zeolite-modified ELISA plate through amine linker. This probe-modified ELISA plate detected low levels of HBsAg, with a detection limit of 0.1 ng/mL. Furthermore, serum spiked experiments showed increment of absorbance with increasing HBsAg concentration, but control trials with other biomolecules showed no increment of absorbance, showing the specific and selective detection of HBsAg. This nanomaterial-modified ELISA plate can detect low levels of HBsAg and help in the diagnosis of HBV infection in its early stages.

Study of influence of metal ions in the diagnosis of recombinant hepatitis B surface antigen (HBsAg) using ELISA technique

ObjectivesPreliminary diagnosis of hepatitis B is performed by measuring the levels of serological markers using enzyme-linked immunosorbent assay (ELISA). Errors in diagnosis of the disease using serum markers might occur due to the presence of metallic salts in diagnostic samples. The present study was designed to evaluate the effects of various metal ions on the diagnosis of recombinant hepatitis B surface antigen (HBsAg). MethodsA simple step sandwich ELISA technique was used to evaluate the effects of metal ions on the diagnosis of HBsAg. Further, the zeta potential, electrical conductivity and redox potential of solutions with proteins and metal salts were measured to evaluate the interaction of these metal ions with the protein. ResultsThe results showed that recombinant HBsAg significantly interacted with the tested metal ions, with lithium ion showing the highest interaction, followed by barium, aluminium, and nickel as the percent decrease in the free HBsAg concentration in presence of these metal ions were found to be 95.5, 94.1, 93.5, and 91.6%, respectively. Zeta potential of the protein in reaction mixture also changed considerably and marked decrease in the concentration of free HBsAg was observed. This decrease in the concentration of free HBsAg resulted into error in the ELISA test which was negative in presence of these metal ions due to strong interaction with the HBsAg protein. ConclusionsThe findings of this study clearly demonstrated that the presence of metal ions in pathological samples can result in interaction with the antigen leading to possible diagnostic errors.

Correlations between serum hepatitis B core-related antigen and hepatitis B surface antigen in patients with hepatitis B cirrhosis and a hepatitis B virus-DNA-negative status: a retrospective study.

ObjectiveThis study aimed to examine the correlations between serum hepatitis B core-related antigen (HBcrAg) and hepatitis B surface antigen (HBsAg) titers in patients with hepatitis B cirrhosis and a hepatitis B virus (HBV)-DNA-negative status.MethodsWe retrospectively analyzed the data and blood samples of patients who were diagnosed with HBV liver cirrhosis and an HBV-DNA negative status. These patients were hospitalized between October 2018 and October 2019 at one hospital.ResultsA total of 180 patients were included. The median (interquartile range) HBsAg and HBcrAg concentrations were 2.77 log10 IU/mL (1.60–3.15) and 3.96 log10 U/mL (2.70–4.97), respectively. A non-linear significant relationship was found between HBsAg and HBcrAg concentrations. The inflection point was 0.58. The effect size and confidence interval on the left and right sides of the inflection point were 0.10 (−0.23–0.42) and 0.62 (0.46–0.78), respectively. When HBsAg concentrations were ≥0.58 log10 IU/mL, HBsAg concentrations were positively correlated with HBcrAg concentrations. When HBsAg concentrations increased by 1 log10 IU/mL, HBcrAg concentrations increased by 0.62 log10 U/mL (95% confidence interval: 0.46, 0.78).ConclusionsThere might be a non-linear relationship between HBcrAg and HBsAg concentrations in patients with hepatitis B cirrhosis and an HBV-DNA-negative status.

Sensitive Aptasensing of Hepatitis B Viral Antigen Gold Dielectrode by Current-Volt Measurements

Hepatitis B virus (HBV) is a serious human health issue, primarily leading to cirrhosis, chronic hepatitis, and primary liver cancer. Patients are at higher risk of a poor prognosis if the viral load exceeds >105 copies/mL; hence, it is crucial to identify the early stages of HBV infection. This research aimed to identify hepatitis B surface antigen (HBsAg) on an interdigitated gold electrode sensor assisted by HBsAg-specific aptamer and antibody interactions. The aptamer-HBsAg-antibody sandwich assay was carried out on an interdigitated electrode using aptamer as the capture molecule and anti-HBsAg antibody as the detection molecule. Higher capture probe immobilization was achieved through the gold nanourchin conjugated capture aptamer. On the aptamer-attached electrode, HBsAg interacted with and was sandwiched by the detection antibody. The sandwich assay lowered the limit of HBsAg detection to 0.1 ng/mL, as indicated by the calculated R2 value of 0.9915 on a linear regression graph presenting data for concentrations of HBsAg from 0.1 to 60 ng/mL. Furthermore, specific HBsAg detection was confirmed by complementarity of the capture aptamer sequence and nonspecific proteins (hepatitis C-the core antigen (HCVcoreAg) and albumin). A selective experiment revealed a gradual increase in current responses in a HBsAg-spiked serum sample, demonstrating the interference-free detection of HBsAg for the real-life sample.

Bushen Jianpi Formula Combined with Entecavir for the Treatment of HBeAg-Negative Chronic Hepatitis B: A Multicenter, Randomized, Double-Blind, Placebo-Controlled Trial.

Background Bushen Jianpi formula (BSJPF, also known as Lingmao formula) is a traditional Chinese medicine for chronic hepatitis B (CHB). The previous study has suggested that the treatment combination of BSJPF and entecavir (ETV) can achieve a significant loss of hepatitis B e antigen (HBeAg) and a significant decrease in serum level of hepatitis B virus (HBV) DNA in HBeAg-positive CHB patients with mildly elevated alanine aminotransferase. Objective This study aimed to evaluate the efficacy and safety of BSJPF combined with ETV for treating HBeAg-negative CHB patients. Methods A total of 640 patients were assigned randomly to the treatment group (receiving BSJPF combined with ETV for 96 weeks) or the control group (receiving a placebo combined with ETV for 96 weeks) in a 1 : 1 ratio. The primary endpoints are the rate of loss of hepatitis B surface antigen (HBsAg). The secondary outcomes included the rate of decrease in the HBsAg concentration to ≥1 lg·IU/mL, the HBV DNA suppression, the decline of the level of covalently closed circular DNA (cccDNA) in the liver, histological improvements, and the rate of ALT normalization. Results The rate of HBsAg loss in the treatment group was significantly higher than that of the control group (5.5% versus 1.8%, P=0.031). There were 11.1% of patients in the treatment group who recorded a reduction in HBsAg ≥1 lg·IU/mL, which is better than 5.9% of patients in the control group (P=0.043). There was no significant difference between the two groups with regard to the rate of HBV DNA clearance, the reduction in intrahepatic cccDNA, and the rate of ALT normalization (P > 0.05). The rate of liver fibrosis improvement in the treatment group was better than that of the control group (35.5% versus 11.8%, P=0.031), but there was no difference in necroinflammatory improvement (P > 0.05). The adverse events (AEs) were similar between the two groups, except for the abnormal kidney function, with 2.2% in the control group and 0.0% in the treatment group (P=0.028). Conclusion The combination of BSJPF and ETV can increase the rate of HBsAg loss and the rate of histological fibrosis improvement without serious adverse events in CHB patients. Trial Registration. This trial is registered with ChiCTR-IOR-16009880 on November 16, 2016—retrospectively registered, http://www.chictr.org.cn/showproj.aspx?proj=16836.

Silicon Nanowires Length and Numbers Dependence on Sensitivity of the Field-Effect Transistor Sensor for Hepatitis B Virus Surface Antigen Detection.

Silicon nanowire field effect transistor (NWFET) sensors have been demonstrated to have high sensitivity, are label free, and offer specific detection. This study explored the effect of nanowire dimensions on sensors’ sensitivity. We used sidewall spacer etching to fabricate polycrystalline silicon NWFET sensors. This method does not require expensive nanoscale exposure systems and reduces fabrication costs. We designed transistor sensors with nanowires of various lengths and numbers. Hepatitis B surface antigen (HBsAg) was used as the sensing target to explore the relationships of nanowire length and number with biomolecule detection. The experimental results revealed that the sensor with a 3 µm nanowire exhibited high sensitivity in detecting low concentrations of HBsAg. However, the sensor reached saturation when the biomolecule concentration exceeded 800 fg/mL. Sensors with 1.6 and 5 µm nanowires exhibited favorable linear sensing ranges at concentrations from 800 ag/mL to 800 pg/mL. The results regarding the number of nanowires revealed that the use of few nanowires in transistor sensors increases sensitivity. The results demonstrate the effects of nanowire dimensions on the silicon NWFET biosensors.

Magnetic separation and enzymatic catalysis conjugated colorimetric immunosensor for Hepatitis B surface antigen detection

Hepatitis B Surface Antigen (HBsAg) is usually recognized as an earliest diagnostic markers for Hepatitis B Virus (HBV) infections. The precise and sensitive determination of HBsAg is crucial for the diagnostic of HBV infection. In this paper, a colorimetric immunosensor has been successfully designed and fabricated based on conjugating magnetic separation and enzymatic catalysis, for point-of-care detection of HBsAg. The HBsAg was simultaneously captured by the biotinylated HBsAg primary antibody (biotin-Ab1) labeled immunomagnetic nanoparticles and detection antibody (Ab2). In conjugation with Ab2, horseradish peroxidase (HRP) labeled rabbit anti-goat secondary antibody was linked with chromogenic agents to induce the colorimetric signal. As a result, a good linear relation was achieved between the concentration of HBsAg and OD450 nm (absorbance values at 450 nm) in the 0.1–20 ng mL−1 ranges, demonstrating a great specificity to HBsAg with a limit of detection of 0.012 ng mL−1. Furthermore, considering its succeed in real human serum sample test, this immunosensor could make contribute to potential applications in rapid diagnosis, effective prevention of epidemic and infectious diseases.

Amine-functionalized Cu-MOF nanospheres towards label-free hepatitis B surface antigen electrochemical immunosensors.

Metal-organic framework (MOF) nanomaterials offer a wide range of promising applications due to their unique properties, including open micro- and mesopores and richness of functionalization. Herein, a facile synthesis via a solvothermal method was successfully employed to prepare amine-functionalized Cu-MOF nanospheres. Moreover, the growth and the morphology of the nanospheres were optimized by the addition of PVP and TEA. By functionalization with an amine group, the immobilization of a bioreceptor towards the detection of hepatitis B infection biomarker, i.e., hepatitis B surface antigen (HBsAg), could be realized. The immobilization of the bioreceptor/antibody to Cu-MOF nanospheres was achieved through a covalent interaction between the carboxyl group of the antibodies and the amino-functional ligand in Cu-MOF via EDC/NHS coupling. The amine-functionalized Cu-MOF nanospheres act not only as a nanocarrier for antibody immobilization, but also as an electroactive material to generate the electrochemical signal. The electrochemical sensing performance was characterized using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and differential pulse voltammetry (DPV). The results showed that the current response proportionally decreased with the increase of HBsAg concentration. More importantly, the sensing performance of the amine-functionalized Cu-MOF nanospheres towards HBsAg detection was found to be consistent in real human serum media. This strategy successfully resulted in wide linear range detection of HBsAg from 1 ng mL-1 to 500 ng mL-1 with a limit of detection (LOD) of 730 pg mL-1. Thus, our approach provides a facile and low-cost synthesis process of an electrochemical immunosensor and paves the way to potentially utilize MOF-based nanomaterials for clinical use.

Hydrazide mediated oriented coupling of antibodies on quantum dot beads for enhancing detection performance of immunochromatographic assay

Immunochromatographic assays (ICAs) are one of the most popular on-site diagnostic tools. The bio-activity of the probe holds great promise for the detection performance of ICA. The orientation of antibodies on nanoparticle surface plays a vital role in improving the bio-activity of the probe. Hydrazide mediated oriented coupling (HDZ-O) strategy can erect the antibodies on the particle surface via the nucleophilic addition reaction between the hydrazide group of particles and the aldehyde group of the Fc region of antibodies. In this work, we synthesized the hydrazide and carboxylic group modified quantum dot beads (QBs-NH-NH2, and QBs-COOH) and compared two coupling strategies for preparing QB probes systematically. Results showed the reaction time for producing QB probe by using HDZ-O method was 20 min, and the optimal labeled antibody content was 80 μg per mg of QBs. Those for carbodiimide method were 180 min and 320 μg per mg of QBs. Moreover, the QB probe by HDZ-O showed a higher bio-activity than that by carbodiimide method under their optimal labeled antibody concentrations. Subsequently, the QB probe by HDZ-O strategy was applied further in sandwich ICA for hepatitis B surface antigen (HBsAg) detection. The results indicated that the strip has a widely dynamic linearity with HBsAg concentration ranging from 0.05 ng/mL to 3200 ng/mL. The limit of detection of the strip is 0.05 ng/mL, which is approximately fourfold lower than that of the probe prepared by carbodiimide method. In addition, the ICA strip also showed acceptable accuracy and precision for HBsAg quantitative detection in real serum samples, which have average recoveries in intra- and inter-assays that vary from 89.16% to 97.63% with the variable coefficients less than 6.98%. The proposed ICA strip showed good agreement (R2 > 0.94) with a commercial chemiluminescence kit for 40 of clinical serum samples. All in all, this study proposed a robust strategy to immobilize the antibodies with proper orientation on the surface of QBs, and the resultant QB probe can improve the detection performance of sandwich LFIA platform remarkably.

Bayesian network to predict hepatitis B surface antigen seroclearance in chronic hepatitis B patients.

There is a need for an interpretable, accurate and interactions-considered model for predicting hepatitis B surface antigen (HBsAg) seroclearance. We aimed to construct a Bayesian network (BN) model using available medical records to predict HBsAg seroclearance in chronic hepatitis B (CHB) patients, and to evaluate the model's performance. This was a case-control study. A total of 1966 consecutive CHB patients (mean age 39.04±11.23years) between January 2006 and June 2015 were included. The demographic and clinical characteristics, laboratory data and imaging parameters were obtained and used to build a BN model to estimate the probability of HBsAg seroclearance. Baseline serum HBsAg and hepatitis Be antigen (HBeAg) levels, virological response and HBeAg seroclearance were the most significant predictors of HBsAg seroclearance. The post-test probability table showed that patients with baseline HBsAg concentrations ≤2000IU/mL, negative baseline HBeAg, an initial virological response and without HBeAg seroclearance (i.e. no recurrence of HBeAg positivity during follow-up) were most likely to have HBsAg seroclearance. The constructed BN model had an area under the receiver operating characteristic curves of 0.896 (95% confidence interval [CI]: 0.892, 0.899), a sensitivity of 0.840 (95% CI: 0.833, 0.846), a specificity of 0.880 (95% CI: 0.876, 0.884) and an accuracy of 0.878 (95% CI: 0.874, 0.882) for predicting HBsAg seroclearance. The established BN model accurately estimated the probability of HBsAg seroclearance and is a promising tool to assist clinical decision-making.

Sequential injection-differential pulse voltammetric immunosensor for hepatitis B surface antigen using the modified screen-printed carbon electrode

A label-free electrochemical immunosensor was developed for the determination of hepatitis B surface antigen (HBsAg), a biomarker of hepatitis B virus (HBV). It is based on the specific binding of antigen-antibody on the electrode surface, which hindered the oxidation of silver nanoparticles (AgNPs) used as redox probe and hence reduced its oxidation current. The screen-printed carbon electrode (SPCE) was modified with carbon nanotube decorated with gold nanoparticles (AuNPs-CNT) and silver nanoparticles. The AuNPs provide good biocompatibility and large surface area for densely immobilizing the hepatitis B surface antibody (HBsAb) on the electrode, which helps to improve the signal. AgNPs act as a sensing probe in differential pulse voltammetric (DPV) detection. In addition, a sequential injection (SI) system was employed as an automated solution handling and minimizing chemical consumption. Characterization of the developed immunosensor by scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) confirmed the successful modification/immobilization in various steps of immunosensor fabrication. The percentage of the decrease in oxidative peak current of AgNPs was directly proportional to the HBsAg concentration. The linear range of 1–40 ng mL−1 with a detection limit of 0.86 ng mL−1 HBsAg were achieved. The proposed method is economical, efficient, and useful for clinical analysis.

Real-time detection of hepatitis B surface antigen using a hybrid graphene-gold nanoparticle biosensor

A hybrid biosensor based on a graphene resistor functionalized with self-assembled Graphene-AuNPs (Gold Nanoparticles) is demonstrated for the real-time detection of hepatitis B surface antigen (HBsAg). The hybrid biosensor consists of a ssDNA sequence attached to a graphene resistor device via π–π stacking interactions in combination with a ssDNA functionalized AuNP. The ssDNA has complementary sequences which through hybridization, yield the graphene-AuNP hybrid biosensor. Real-time 2-point resistance measurements, performed using varying concentrations of HBsAg, show a linear dependence of resistance change against the logarithm of HBsAg concentration (log[HBsAg]). A limit of detection of 50 pg ml−1 was observed. Moreover, the hybrid biosensor platform has potential to be applied to any biomarker of interest.

Development of a multiplex immunochromatographic strip test and ultrasensitive electrochemical immunosensor for hepatitis B virus screening

This research proposes two methods for hepatitis B diagnosis including rapid testing and electrochemical assay. For the first method, a multiplex hepatitis B test strip was fabricated to serve as a rapid test for hepatitis B screening. It was developed to simultaneously test three essential serological markers of hepatitis B virus infection including hepatitis B surface antigen (HBsAg), hepatitis B surface antibody (Anti-HBs) and hepatitis B core antibody (Anti-HBc). Gold nanoparticles (GNPs) were used as the signal generator on the test strip. Furthermore, a part of a paper network was incorporated on the strip for the gold-silver enhancement process. This paper network helped in decreasing the analysis time of enhancement and makes the enhancement process easier for rapid testing. The developed test strip was specific for each serological marker. The detection limits of HBsAg, Anti-HBs and Anti-HBc were obtained at 0.5, 0.3 and 0.1 μg mL−1, respectively. For the second method, electrochemical impedance spectroscopy (EIS) was applied for HBsAg detection. This method was proposed for quantitative hepatitis B detection. Anti-HBs antibodies were immobilized on a carbon screen printed electrode (SPCE) via the N-ethyl-N′-(3-(dimethylamino)propyl)carbo-diimide/N-hydroxy succinimide (EDC/NHS) couple reaction which reacted with the carboxyl group of the BSA cross-linked film on the electrode. The electrode modification process was characterized by EIS. A linear relationship between delta charge transfer resistance (ΔRct) and HBsAg concentration was obtained in the range of 5–3000 ng mL−1 with a detection limit of 2.1 ng mL−1. This work is appropriate for quantitative analysis because it is a simple and low-cost method to implement as the SPCE is disposable. Therefore, we hope that this research will be useful to improve hepatitis B detection in the future.

Accurate and cost-effective prediction of HBsAg titer in industrial scale fermentation process of recombinant Pichia pastoris by using neural network based soft sensor.

In the current work, the attempt was made to apply best-fitted artificial neural network (ANN) architecture and the respective training process for predicting final titer of hepatitis B surface antigen (HBsAg), produced intracellularly by recombinant Pichia pastoris Mut+ in the commercial scale. For this purpose, in large-scale fed-batch fermentation, using methanol for HBsAg induction and cell growth, three parameters of average specific growth rate, biomass yield, and dry biomass concentration-in the definite integral form with respect to fermentation time-were selected as input vectors; the final concentration of HBsAg was selected for the ANN output. Used dataset consists of 38 runs from previous batches; feed-forward ANN 3:5:1 with training algorithm of backpropagation based on a Bayesian regularization was trained and tested with a high degree of accuracy. Implementing the verified ANN for predicting the HBsAg titer of the five new fermentation runs, excluded from the dataset, in the full-scale production, the coefficient of regression and root-mean-square error were found to be 0.969299 and 2.716774, respectively. These results suggest that this verified soft sensor could be an excellent alternative for the current relatively expensive and time-intensive analytical techniques such as enzyme-linked immunosorbent assay in the biopharmaceutical industry.

Safety and efficacy of stopping tenofovir disoproxil fumarate in patients with chronic hepatitis B following at least 8 years of therapy: a prespecified follow-up analysis of two randomised trials

Effective and well tolerated nucleos(t)ide analogue treatment exists for patients with chronic hepatitis B, although treatment is generally anticipated to be life-long, with concomitant costs and treatment-related side-effects. We aimed to characterise the outcomes of patients with persistent viral suppression who discontinued nucleotide analogue use after extended treatment. The primary objective of this prespecified analysis was to evaluate the safety of stopping long-term tenofovir disoproxil fumarate therapy in patients enrolled in two (completed) randomised controlled studies, GS-US-174-0102 (ClinicalTrials.gov, number NCT00117676) and GS-US-174-0103 (ClinicalTrials.gov, number NCT00116805). In those studies, patients who had completed 8 years or more of nucleotide analogue treatment, were hepatitis B surface antigen (HBsAg)-positive with hepatitis B virus (HBV) DNA concentration of less than 29 IU/mL, and were unwilling or unable to continue therapy were required by protocol to enter a 24-week treatment-free follow-up (TFFU) phase. We present data for patients in the TFFU phase who were assessed at baseline and monitored every 4 weeks for changes in qualitative serum HBsAg, HBV DNA, and alanine aminotransferase (ALT) concentrations in addition to standard safety assessments. Of 124 patients who entered the TFFU phase, 54 (44%) patients did not complete 24 weeks of follow-up (median 12 weeks; IQR 0-20). Overall, 32 (26%) patients reported an adverse event. Serious adverse events occurred in five (4%) patients, including elevated ALT concentrations in two patients, hepatic flare in two patients, and increased lipase in one patient. 38 (31%) of patients had grade 3 or higher laboratory abnormalities, the majority of which were ALT elevations (36 patients). Of the 106 hepatitis B e antigen (HBeAg)-negative patients who entered the TFFU phase, 63 (59%) were followed for 24 weeks. HBsAg loss was observed in five (5%) of the 106 HBeAg-negative patients who entered the TFFU phase, and 37 (35%) had both HBV DNA concentrations of less than 2000 IU/mL and ALT concentrations less than the ULN at TFFU week 24. 18 HBeAg-positive patients entered the TFFU phase, of whom seven (39%) were followed up for 24 weeks. Of these seven patients, none had HBsAg loss or HBV DNA of less than 2000 IU/mL and one (14%) had an ALT less than the ULN at week 24. Within 24 weeks of stopping 8 years or more of nucleotide analogue therapy almost a third of patients experienced a grade 3 or higher laboratory abnormality. Although few patients achieved HBsAg loss, a subgroup of HBeAg-negative patients can achieve a low-replicative state within a short duration of follow-up. Gilead Sciences, Inc.

PACKAGING OF HEPATITIS B SURFACE ANTIGEN VACCINE AND MORINGA OLEIFERA EXTRACT INTO CHITOSAN NANOPARTICLES

Objective: Oftentimes, the recombinant antigen for the use of vaccines is less immunogenic than live attenuated or inactive vaccines. Hence, a potent adjuvant is needed to enhance the immune response. Moreover, the role of vector design is also important to facilitate the improvement of the immune response. The aim of this research was to develop hepatitis B surface antigen (HBsAg)-loaded nanoparticles and Moringa oleifera aqueous leaf extracts as an adjuvant using chitosan polymer. Methods: Chitosan nanoparticles were prepared by the ionic gelation method using sodium tripolyphosphate as the cross-linking agent. A system was composed of chitosan core in which HBsAg and M. oleifera extracts were incorporated. The concentration of HBsAg used in this combination was 10 μg/ml, and the concentrations of extracts were 10, 50, and 100 μg/ml, respectively. In this study, three types of nanoparticles were produced: HBsAg-loaded nanoparticles, M. oleifera-loaded nanoparticles, and combination of HBsAg–M. oleifera-loaded nanoparticles. The nanoparticles formed were characterized by the particle size, HBsAg entrapment efficiency using sodium dodecyl sulfate polyacrylamide gel electrophoresis, and the entrapment efficiency of extracts using the total flavonoid method. Results: The results showed that the particle size was between 111 and 245 nm. The entrapment efficiency of HBsAg in the separate formula was 79%, while that in the combined formula was approximately 96–98%. Furthermore, the entrapment efficiency of the extracts in the separate formula was around 64–91%, while that in the combined formula was 55–82.5%. Particularly, HBsAg–M. oleifera-loaded chitosan nanoparticles with the extract concentrations of 50 μg/ml showed the highest entrapment efficiencies of HBsAg and M. oleifera extracts of approximately 98 and 82.5%, respectively. Conclusion: Collectively, the system has been successfully developed, so it is then plausible to determine the function of the devices to enhance the immune response in the future.

PACKAGING OF HEPATITIS B SURFACE ANTIGEN VACCINE AND MORINGA OLEIFERA EXTRACT INTO CHITOSAN NANOPARTICLES

Objective: Oftentimes, the recombinant antigen for the use of vaccines is less immunogenic than live attenuated or inactive vaccines. Hence, a potent adjuvant is needed to enhance the immune response. Moreover, the role of vector design is also important to facilitate the improvement of the immune response. The aim of this research was to develop hepatitis B surface antigen (HBsAg)-loaded nanoparticles and Moringa oleifera aqueous leaf extracts as an adjuvant using chitosan polymer. Methods: Chitosan nanoparticles were prepared by the ionic gelation method using sodium tripolyphosphate as the cross-linking agent. A system was composed of chitosan core in which HBsAg and M. oleifera extracts were incorporated. The concentration of HBsAg used in this combination was 10 μg/ml, and the concentrations of extracts were 10, 50, and 100 μg/ml, respectively. In this study, three types of nanoparticles were produced: HBsAg-loaded nanoparticles, M. oleifera-loaded nanoparticles, and combination of HBsAg–M. oleifera-loaded nanoparticles. The nanoparticles formed were characterized by the particle size, HBsAg entrapment efficiency using sodium dodecyl sulfate polyacrylamide gel electrophoresis, and the entrapment efficiency of extracts using the total flavonoid method. Results: The results showed that the particle size was between 111 and 245 nm. The entrapment efficiency of HBsAg in the separate formula was 79%, while that in the combined formula was approximately 96–98%. Furthermore, the entrapment efficiency of the extracts in the separate formula was around 64–91%, while that in the combined formula was 55–82.5%. Particularly, HBsAg–M. oleifera-loaded chitosan nanoparticles with the extract concentrations of 50 μg/ml showed the highest entrapment efficiencies of HBsAg and M. oleifera extracts of approximately 98 and 82.5%, respectively. Conclusion: Collectively, the system has been successfully developed, so it is then plausible to determine the function of the devices to enhance the immune response in the future.

A study on false positive results of hepatitis B surface antigen (HBsAg) test: Frequency of samples considered false positives in relation to the HBsAg concentration

A study on false positive results of hepatitis B surface antigen (HBsAg) test: Frequency of samples considered false positives in relation to the HBsAg concentration