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A preparative purification process for recombinant hepatitis B core antigen using online capture by expanded bed adsorption followed by size-exclusion chromatography

Ho CW, Tan WS, Chong FC, Ling TC, Tey BT

J Microbiol Biotechnol, 2009 Apr;19(4):416-23.
PMID: 19421000

Hepatitis B core antigen (HBcAg) is an important serological marker used in the diagnosis of hepatitis B virus (HBV) infections. In the current study, a fast and efficient preparative purification protocol for truncated HBcAg from Escherichia coli disruptate was developed. The recombinant HBcAg was first captured by anion exchange expanded bed adsorption chromatography integrated with a cell disruption process. This online capture process has shortened the process time and eliminated the "hold-up" period that may be detrimental to the quality of target protein. The eluted product from the expanded bed adsorption chromatography was subsequently purified using size-exclusion chromatography. The results showed that this novel purification protocol achieved a recovery yield of 45.1% with a product purity of 88.2%, which corresponds to a purification factor of 4.5. The recovered HBcAg is still biologically active as shown by ELISA test.

Matched MeSH terms: Hepatitis B Core Antigens/genetics; Hepatitis B Core Antigens/isolation & purification*; Hepatitis B Core Antigens/metabolism
Negative chromatography purification of hepatitis B virus-like particles using poly(oligo(ethylene glycol) methacrylate) grafted cationic adsorbent

Lee MF, Chan ES, Tan WS, Tam KC, Tey BT

J Chromatogr A, 2015 Oct 9;1415:161-5.
PMID: 26358561 DOI: 10.1016/j.chroma.2015.08.056

Poly(oligo(ethylene glycol) methacrylate) (POEGMA), an inert polymer was grafted onto an anion exchange adsorbent for the exclusion of relatively larger hepatitis B virus-like particles (HB-VLPs) from the anion exchange ligand (Q) and at the same time this process allowed the selective adsorption of smaller size Escherichia coli host cell proteins (HCPs). The chain lengths of the POEGMA grafted were modulated by varying the amount of monomers used in the polymer grafting. The purification factor and yield of the HB-VLPs obtained from the flow-through of negative chromatography were 2.3 and 66.0±3.1%, respectively, when shorter chain length of POEGMA (SQ) was grafted. Adsorbent grafted with longer chain of POEGMA (LQ) excluded some HCPs that are larger in size together with the HB-VLPs, reducing the purity of the recovered HB-VLPs. Further heat-treatment of the flow-through pool from SQ followed by centrifugation increased the purity of heat stable HB-VLPs to 87.5±1.1%. Heat-treatment of the flow through sample resulted in thermal denaturation and aggregation of HCPs, while the heat stable HB-VLPs still remained intact as observed under a transmission electron microscope. The performance of the negative chromatography together with heat treatment in the purification of HB-VLPs is far better than the reported bind-and-elute techniques.

Matched MeSH terms: Hepatitis B Core Antigens/genetics; Hepatitis B Core Antigens/isolation & purification*; Hepatitis B Core Antigens/metabolism
Hepatitis B virus peptide inhibitors: solution structures and interactions with the viral capsid

Muhamad A, Ho KL, Rahman MB, Tejo BA, Uhrín D, Tan WS

Org Biomol Chem, 2015 Jul 28;13(28):7780-9.
PMID: 26100394 DOI: 10.1039/c5ob00449g

Hepatitis B virus (HBV) infection remains a health problem globally despite the availability of effective vaccines. In the assembly of the infectious virion, both the preS and S regions of the HBV large surface antigen (L-HBsAg) interact synergistically with the viral core antigen (HBcAg). Peptides preS and S based on the L-HBsAg were demonstrated as potential inhibitors to block the viral assembly. Therefore, the objectives of this study were to determine the solution structures of these peptides and study their interactions with HBcAg. The solution structures of these peptides were solved using (1)H, (13)C, and (15)N NMR spectroscopy. Peptide preS has several structured regions of β-turns at Ser7-Pro8-Pro9, Arg11-Thr12-Thr13 and Ser22-Thr23-Thr24 sequences whereas peptide S has only one structured region observed at Ser3-Asn4-His5. Both peptides contain bend-like structures surrounding the turn structures. Docking studies revealed that both peptides interacted with the immunodominant region of HBcAg located at the tip of the viral capsid spikes. Saturation Transfer Difference (STD) NMR experiments identified several aromatic residues in peptides preS and S that interact with HBcAg. This study provides insights into the contact regions of L-HBsAg and HBcAg at atomic resolution which can be used to design antiviral agents that inhibit HBV morphogenesis.

Matched MeSH terms: Hepatitis B Core Antigens/isolation & purification; Hepatitis B Core Antigens/pharmacology; Hepatitis B Core Antigens/chemistry*
N-terminally His-tagged hepatitis B core antigens: construction, expression, purification and antigenicity

Yap WB, Tey BT, Ng MY, Ong ST, Tan WS

J Virol Methods, 2009 Sep;160(1-2):125-31.
PMID: 19433111 DOI: 10.1016/j.jviromet.2009.04.038

The core antigen of the hepatitis B virus (HBcAg) has been used widely as a diagnostic reagent for the identification of the viral infection. However, purification using the conventional sucrose density gradient ultracentrifugation is time consuming and costly. To overcome this, HBcAg particles displaying His-tag on their surface were constructed and produced in Escherichia coli. The recombinant His-tagged HBcAgs were purified using immobilized metal affinity chromatography. Transmission electron microscopy and enzyme-linked immunosorbent assay (ELISA) revealed that the displayed His-tag did not impair the formation of the core particles and the antigenicity of HBcAg.

Matched MeSH terms: Hepatitis B Core Antigens/genetics; Hepatitis B Core Antigens/immunology*; Hepatitis B Core Antigens/isolation & purification*
Recombinant hepatitis B virus core particles: association, dissociation and encapsidation of green fluorescent protein

Lee KW, Tan WS

J Virol Methods, 2008 Aug;151(2):172-180.
PMID: 18584885 DOI: 10.1016/j.jviromet.2008.05.025

The recombinant hepatitis B virus (HBV) core antigen (HBcAg) expressed in Escherichia coli self-assembles into icosahedral capsids of about 35 nm which can be exploited as gene or drug delivery vehicles. The association and dissociation properties of the C-terminally truncated HBcAg with urea and guanidine hydrochloride (GdnHCl) were studied. Transmission electron microscopy (TEM) revealed that the dissociated HBcAg was able to re-associate into particles when the applied denaturing agents were physically removed. In order to evaluate the potential of the particles in capturing molecules, purified green fluorescent protein (GFP) was applied to the dissociated HBcAg for encapsidation. The HBcAg particles harbouring the GFP molecules were purified using sucrose density gradient ultracentrifugation and analysed using native agarose gel electrophoresis and TEM. A method for the encapsidation of GFP in HBcAg particles which has the potential to capture drugs or nucleic acids was established.

Matched MeSH terms: Hepatitis B Core Antigens/genetics*; Hepatitis B Core Antigens/isolation & purification; Hepatitis B Core Antigens/ultrastructure
Fulltext Delivery of chimeric hepatitis B core particles into liver cells

Lee KW, Tey BT, Ho KL, Tan WS

J Appl Microbiol, 2012 Jan;112(1):119-31.
PMID: 21992228 DOI: 10.1111/j.1365-2672.2011.05176.x

To display a liver-specific ligand on the hepatitis B virus core particles for cell-targeting delivery.

Matched MeSH terms: Hepatitis B Core Antigens/genetics*; Hepatitis B Core Antigens/metabolism*
Interactions of hepatitis B core antigen and peptide inhibitors

Tang KF, Abdullah MP, Yusoff K, Tan WS

J Med Chem, 2007 Nov 15;50(23):5620-6.
PMID: 17918821

The core protein (HBcAg) of hepatitis B virus (HBV) has been shown to interact with the large surface antigen during HBV morphogenesis, and these interactions can be blocked by small peptides selected from either linear or constrained phage display peptide libraries. The association of HBcAg with peptide inhibitors was quantitatively evaluated by isothermal titration calorimetry. The thermodynamic data show that the interaction between HBcAg and peptide MHRSLLGRMKGA is enthalpy-driven and occurs at a 3:1 stoichiometry and dissociation constant (Kd) value of 79.4 muM. However, peptide WSFFSNI displays a higher binding affinity for HBcAg with a Kd value of 18.5 muM when compared to peptide MHRSLLGRMKGA. A combinatorial approach using chemical cross-linking and surface-enhanced laser desorption/ionization-time-of-flight-mass spectrometry shows that the Lys of peptide MHRSLLGRMKGA interacted either with D64, E77, or D78 of HBcAg.

Matched MeSH terms: Hepatitis B Core Antigens/isolation & purification; Hepatitis B Core Antigens/chemistry*
Effect of different operating modes and biomass concentrations on the recovery of recombinant hepatitis B core antigen from thermal-treated unclarified Escherichia coli feedstock

Ng MY, Tan WS, Abdullah N, Ling TC, Tey BT

J Biotechnol, 2008 Nov 25;138(3-4):74-9.
PMID: 18786579 DOI: 10.1016/j.jbiotec.2008.08.004

Expanded bed adsorption chromatography (EBAC) is a single pass operation that has been used as primary capture step in various protein purifications. The most common problem in EBAC is often associated with successful formation of a stable fluidized bed during the absorption stage, which is critically dependent on parameters such as liquid velocity, bed height, particle (adsorbent) size and density as well as design of column and type of flow distributor. In this study, residence time distribution (RTD) test using acetone as non-binding tracer acetone was performed to evaluate liquid dispersion characteristics of the EBAC system. A high B(o) number was obtained indicating the liquid dispersion in the system employed is very minimal and the liquid flow within the bed was close to plug flow, which mimics a packed bed chromatography system. Evaluation on the effect of flow velocities and bed height on the performance of Streamline DEAE using feedstock containing heat-treated crude Escherichia coli homogenate of different biomass concentrations was carried out in this study. The advantages and disadvantages as well as the problems encountered during recovery of HBcAg with aforementioned parameters are also discussed in this paper.

Matched MeSH terms: Hepatitis B Core Antigens/isolation & purification*; Hepatitis B Core Antigens/metabolism
Native agarose gel electrophoresis and electroelution: A fast and cost-effective method to separate the small and large hepatitis B capsids

Yoon KY, Tan WS, Tey BT, Lee KW, Ho KL

Electrophoresis, 2013 Jan;34(2):244-53.
PMID: 23161478 DOI: 10.1002/elps.201200257

Hepatitis B core antigen (HBcAg) expressed in Escherichia coli is able to self-assemble into large and small capsids comprising 240 (triangulation number T = 4) and 180 (triangulation number T = 3) subunits, respectively. Conventionally, sucrose density gradient ultracentrifugation and SEC have been used to separate these capsids. However, good separation of the large and small particles with these methods is never achieved. In the present study, we employed a simple, fast, and cost-effective method to separate the T = 3 and T = 4 HBcAg capsids by using native agarose gel electrophoresis followed by an electroelution method (NAGE-EE). This is a direct, fast, and economic method for isolating the large and small HBcAg particles homogenously based on the hydrodynamic radius of the spherical particles. Dynamic light scattering analysis demonstrated that the T = 3 and T = 4 HBcAg capsids prepared using the NAGE-EE method are monodisperse with polydispersity values of ∼15% and ∼13%, respectively. ELISA proved that the antigenicity of the capsids was not affected in the purification process. Overall, NAGE-EE produced T = 3 and T = 4 capsids with a purity above 90%, and the recovery was 34% and 50%, respectively (total recovery of HBcAg is ∼84%), and the operation time is 15 and 4 times lesser than that of the sucrose density gradient ultracentrifugation and SEC, respectively.

Matched MeSH terms: Hepatitis B Core Antigens/biosynthesis; Hepatitis B Core Antigens/genetics; Hepatitis B Core Antigens/isolation & purification*; Hepatitis B Core Antigens/chemistry
Direct purification of recombinant hepatitis B core antigen from two different pre-conditioned unclarified Escherichia coli feedstocks via expanded bed adsorption chromatography

Ng MY, Tan WS, Abdullah N, Ling TC, Tey BT

J Chromatogr A, 2007 Nov 16;1172(1):47-56.
PMID: 17945242

Direct recovery of hepatitis B core antigen (HBcAg) from unclarified Escherichia coli homogenates via expanded bed adsorption chromatography (EBA) has been explored in this study. Streamline DEAE was selected as the anion exchanger to recover HBcAg from heat-treated and non-heat-treated unclarified feedstocks. The use of anion-exchanger for direct extraction of proteins from unclarified feedstock is not preferred due to lack of specificity of its ligand. In this study, thermal treatment of the unclarified feedstock at 60 degrees C has resulted in 1.2- and 1.8-fold increases in yield and purity of HBcAg, respectively, compared with that purified from non-heat-treated feedstock. Heating the crude feedstock has resulted in denaturation and precipitation of contaminants in the feedstock, hence reducing non-specific interactions between the cell debris and adsorbent. The selectivity of the anion-exchanger has also been increased as shown in the breakthrough curve obtained. Enzyme-linked immunosorbent assay showed that the antigenicity of the HBcAg from heat-treated unclarified feedstock is still preserved.

Matched MeSH terms: Hepatitis B Core Antigens/isolation & purification*; Hepatitis B Core Antigens/chemistry
Purification of His-tagged hepatitis B core antigen from unclarified bacterial homogenate using immobilized metal affinity-expanded bed adsorption chromatography

Yap WB, Tey BT, Alitheen NB, Tan WS

J Chromatogr A, 2010 May 21;1217(21):3473-80.
PMID: 20388569 DOI: 10.1016/j.chroma.2010.03.012

Hepatitis B core antigen (HBcAg) is used as a diagnostic reagent for the detection of hepatitis B virus infection. In this study, immobilized metal affinity-expanded bed adsorption chromatography (IMA-EBAC) was employed to purify N-terminally His-tagged HBcAg from unclarified bacterial homogenate. Streamline Chelating was used as the adsorbent and the batch adsorption experiment showed that the optimal binding pH of His-tagged HBcAg was 8.0 with a binding capacity of 1.8 mg per ml of adsorbent. The optimal elution condition for the elution of His-tagged HBcAg from the adsorbent was at pH 7 in the presence of 500 mM imidazole and 1.5 M NaCl. The IMA-EBAC has successfully recovered 56% of His-tagged HBcAg from the unclarified E. coli homogenate with a purification factor of 3.64. Enzyme-linked immunosorbent assay (ELISA) showed that the antigenicity of the recovered His-tagged HBcAg was not affected throughout the IMA-EBAC purification process and electron microscopy revealed that the protein assembled into virus-like particles (VLP).

Matched MeSH terms: Hepatitis B Core Antigens/genetics; Hepatitis B Core Antigens/isolation & purification*; Hepatitis B Core Antigens/metabolism
A phage-displayed single chain variable fragment that interacts with hepatitis B core antigen: library construction, selection and diagnosis

Tan GH, Yusoff K, Seow HF, Tan WS

J Clin Virol, 2007 Jan;38(1):49-56.
PMID: 17074533

Phage display is an alternative method for constructing and selecting antibodies with desired specificity towards an antigen.

Matched MeSH terms: Hepatitis B Core Antigens/analysis; Hepatitis B Core Antigens/genetics*; Hepatitis B Core Antigens/immunology
A Dual Bioconjugated Virus-Like Nanoparticle as a Drug Delivery System and Comparison with a pH-Responsive Delivery System

Biabanikhankahdani R, Ho KL, Alitheen NB, Tan WS

Nanomaterials (Basel), 2018 Apr 13;8(4).
PMID: 29652827 DOI: 10.3390/nano8040236

Modifications of virus-like nanoparticles (VLNPs) using chemical conjugation techniques have brought the field of virology closer to nanotechnology. The huge surface area to volume ratio of VLNPs permits multiple copies of a targeting ligand and drugs to be attached per nanoparticle. By exploring the chemistry of truncated hepatitis B core antigen (tHBcAg) VLNPs, doxorubicin (DOX) was coupled covalently to the external surface of these nanoparticles via carboxylate groups. About 1600 DOX molecules were conjugated on each tHBcAg VLNP. Then, folic acid (FA) was conjugated to lysine residues of tHBcAg VLNPs to target the nanoparticles to cancer cells over-expressing folic acid receptor (FR). The result demonstrated that the dual bioconjugated tHBcAg VLNPs increased the accumulation and uptake of DOX in the human cervical and colorectal cancer cell lines compared with free DOX, resulting in enhanced cytotoxicity of DOX towards these cells. The fabrication of these dual bioconjugated nanoparticles is simple, and drugs can be easily conjugated with a high coupling efficacy to the VLNPs without any limitation with respect to the cargo's size or charge, as compared with the pH-responsive system based on tHBcAg VLNPs. These dual bioconjugated nanoparticles also have the potential to be modified for other combinatorial drug deliveries.

Matched MeSH terms: Hepatitis B Core Antigens
Detection of hepatitis B virus core antigen by phage display mediated TaqMan real-time immuno-PCR

Monjezi R, Tan SW, Tey BT, Sieo CC, Tan WS

J Virol Methods, 2013 Jan;187(1):121-6.
PMID: 23022731 DOI: 10.1016/j.jviromet.2012.09.017

The core antigen (HBcAg) of hepatitis B virus (HBV) is one of the markers for the identification of the viral infection. The main purpose of this study was to develop a TaqMan real-time detection assay based on the concept of phage display mediated immuno-PCR (PD-IPCR) for the detection of HBcAg. PD-IPCR combines the advantages of immuno-PCR (IPCR) and phage display technology. IPCR integrates the versatility of enzyme-linked immunosorbent assay (ELISA) with the sensitivity and signal generation power of PCR. Whereas, phage display technology exploits the physical association between the displayed peptide and the encoding DNA within the same phage particle. In this study, a constrained peptide displayed on the surface of an M13 recombinant bacteriophage that interacts tightly with HBcAg was applied as a diagnostic reagent in IPCR. The phage displayed peptide and its encoding DNA can be used to replace monoclonal antibody (mAb) and chemically bound DNA, respectively. This method is able to detect as low as 10ng of HBcAg with 10(8)pfu/ml of the recombinant phage which is about 10,000 times more sensitive than the phage-ELISA. The PD-IPCR provides an alternative means for the detection of HBcAg in human serum samples.

Matched MeSH terms: Hepatitis B Core Antigens/blood*; Hepatitis B Core Antigens/genetics; Hepatitis B Core Antigens/immunology
The direct recovery of recombinant hepatitis B core antigen from disruptate derived from continuous-flow bead milling

Ho CW, Tan WS, Kamaruddin S, Ling TC, Tey BT

Biotechnol Appl Biochem, 2008 May;50(Pt 1):49-59.
PMID: 17760564

HBcAg (hepatitis B core antigen) is a nanoplex bioproduct that has a great potential in the development of therapeutic drugs and vaccines. In the present study, a continuous-flow bead milling for the disruption of Escherichia coli was optimized and a direct recovery protocol to isolate the recombinant HBcAg from the unclarified E. coli disruptate was developed. The optimal condition for continuous-flow bead milling for the release of HBcAg from E. coli was achieved at a feed flow rate of 15 litres/h, biomass concentration of 10% [ww/v (wet weight/vol.)] and impeller tip speed of 14 m/s. The sucrose-density-gradient analysis showed that the particulate form of the HBcAg released by this optimal condition is still preserved. In the direct purification of HBcAg from the unclarified disruptate, the AE-EBAC (anion-exchange expanded-bed adsorption chromatography) technique was employed. A 54% adsorption and 50.7% recovery of HBcAg were achieved in this direct recovery process. The purity of HBcAg recovered was 49.8%, which corresponds to a purification factor of 2.0. ELISA showed that the HBcAg recovered is functionally active.

Matched MeSH terms: Hepatitis B Core Antigens/isolation & purification*
The frequency of pre-core gene mutations in chronic hepatitis B infection: a study of Malaysian subjects

Yap SF, Wong PW, Chen YC, Rosmawati M

Southeast Asian J. Trop. Med. Public Health, 2002 Mar;33(1):102-9.
PMID: 12118437

A retrospective study was carried out to determine the frequency of the pre-core stop codon mutant virus in a group of chronic hepatitis B carriers: 81 cases were considered [33 hepatits B e antigen (HBe) positive and 48 HBe negative]. All of the HBe positive cases had detectable viral DNA by hybridization analysis; in the case of the HBe negative cases, one third had detectable viral DNA by hybridization analysis and two thirds had HBV DNA detectable by polymerase chain reaction (PCR) amplification. Pre-core stop codon mutant detection was carried out on all specimens using allele-specific oligonucleotide hybridization following PCR amplification of the target sequence. The pre-core mutant was detected in 13/33 (39.4%) of HBe positive cases and in 32/48 (66.7%) of HBe negative cases. Sequence analysis was carried out on 8 of the 16 HBe negative specimens that did not carry the pre-core mutant virus to determine the molecular basis for the HBe minus phenotype in these cases: the 1762/1764 TA paired mutation in the second AT rich region of the core promoter was detected in five cases; a start codon mutation was detected in one case. The predominant mutation resulting in the HBe minus phenotype in our isolates was the 1896A pre-core ("pre-core stop codon") mutation; other mutations responsible for the phenotype included the core promoter paired mutation and pre-core start codon mutation. In view of the high frequency of the pre-core mutant virus, sequence analysis was performed to determine the virus genotype on the basis of the nucleotide sequence of codon 15. The sequences of 21 wild type virus (14 HBe positive and 7 HBe negative cases) were examined: 15 were found to be codon 15 CCT variants (71.4%); the frequency in the HBe positive group was 12/14 (85.7%), while that in the HBe negative group was 3/7 (42.9%). The high frequency of the codon 15 CCT variant in association with the frequent occurrence of the pre-core mutant in our isolates concurs with the results of other studies.

Matched MeSH terms: Hepatitis B Core Antigens/genetics*
Prevalence of anti-HBc in Malaysian male blood donors and its correlation with DNA polymerase activity

Ton SH, Lopez CG, Hasnah H

Southeast Asian J. Trop. Med. Public Health, 1979 Mar;10(1):1-6.
PMID: 483004

A study of Kuala Lumpur blood donors for HBsAG, anti-HBc and DNA polymeraes showed that 5.5% in the sample population was positive for HBsAG, 50.1% for anti-HBc and 10.1% for DNA polymerase activity. There was no significant difference of the HBsAG among the Malay, Chinese and Indian groups. However, a significant difference was observed for the anti-HBc and DNA polymerase activity between the Indian and the Malay/Chinese groups. Both analysis were significantly lower in the Indians but there was no significant difference between the Chinese and the Malays.

Matched MeSH terms: Hepatitis B Core Antigens/analysis*
Fulltext Shielding of Hepatitis B Virus-Like Nanoparticle with Poly(2-Ethyl-2-Oxazoline)

Fam SY, Chee CF, Yong CY, Ho KL, Mariatulqabtiah AR, Lau HY, et al.

Int J Mol Sci, 2019 Oct 03;20(19).
PMID: 31623310 DOI: 10.3390/ijms20194903

Virus-like nanoparticles (VLNPs) have been studied extensively as nanocarriers for targeted drug delivery to cancer cells. However, VLNPs have intrinsic drawbacks, in particular, potential antigenicity and immunogenicity, which hamper their clinical applications. Thus, they can be eliminated easily and rapidly by host immune systems, rendering these nanoparticles ineffective for drug delivery. The aim of this study was to reduce the antigenicity of hepatitis B core antigen (HBcAg) VLNPs by shielding them with a hydrophilic polymer, poly(2-ethyl-2-oxazoline) (PEtOx). In the present study, an amine-functionalized PEtOx (PEtOx-NH2) was synthesized using the living cationic ring-opening polymerization (CROP) technique and covalently conjugated to HBcAg VLNPs via carboxyl groups. The PEtOx-conjugated HBcAg (PEtOx-HBcAg) VLNPs were characterized with dynamic light scattering and UV-visible spectroscopy. The colloidal stability study indicated that both HBcAg and PEtOx-HBcAg VLNPs maintained their particle size in Tris-buffered saline (TBS) at human body temperature (37 °C) for at least five days. Enzyme-linked immunosorbent assays (ELISA) demonstrated that the antigenicity of PEtOx-HBcAg VLNPs reduced significantly as compared with unconjugated HBcAg VLNPs. This novel conjugation approach provides a general platform for resolving the antigenicity of VLNPs, enabling them to be developed into a variety of nanovehicles for targeted drug delivery.

Matched MeSH terms: Hepatitis B Core Antigens/chemistry
Hepatitis C virus core antigen as alternative diagnostic algorithm for active hepatitis C virus infection among haemodialysis population: Cost implications

Wong XZ, Amirah A, Gan CC, Fatiha S, Maznah D, Yahya R, et al.

Nephrology (Carlton), 2021 May;26(5):463-470.
PMID: 33580732 DOI: 10.1111/nep.13862

AIMS: In Malaysia, majority anti-HCV positive haemodialysis patients do not undergo hepatitis C confirmation due to the high cost of HCV RNA. HCV Core Antigen might be a cost-effective diagnostic test to identify HD patients who have active HCV infection eligible for Direct Acting Anti-viral therapy.
METHODS: A cross-sectional study was conducted to assess the correlation between HCV Ag and HCV RNA and the cost implications of different diagnostic algorithms to diagnose active HCV infection using Anti-HCV, HCV Ag, and HCV RNA. Pre-dialysis blood was tested for both HCV Ag and HCV RNA. HCV Ag was tested with Abbott ARCHITECT HCV Ag test.
RESULTS: Two-hundred twenty-seven haemodialysis patients were recruited from 20 centres with mean age of 57.68 ± 12.48 years, and male constitutes 56.8% (129) of the study population. HCV Ag correlated well with HCV RNA (Spearman test coefficient 0.943, p

Matched MeSH terms: Hepatitis B Core Antigens/blood*
Heat treatment of unclarified Escherichia coli homogenate improved the recovery efficiency of recombinant hepatitis B core antigen

Ng MY, Tan WS, Abdullah N, Ling TC, Tey BT

J Virol Methods, 2006 Oct;137(1):134-9.
PMID: 16860402

Heat precipitation procedure has been regularly incorporated as a selective purification step in various thermostable proteins expressed in different hosts. This method is efficient in precipitation of most of the host proteins and also deactivates various host proteases that can be harmful to the desired gene products. In this study, introduction of heat treatment procedure in the purification of hepatitis B core antigen (HBcAg) produced in Escherichia coli has been investigated. Thermal treatment of the cell homogenate at 60 degrees C for 30 min prior to subsequent clarification steps has resulted in 1.4 times and 18% higher in purity and recovery yield, respectively, compared to the non-heat-treated cell homogenate. In direct capture of HBcAg by using anion-exchangers from unclarified feedstock, pre-conditioning the feedstock by heat treatment at 60 degrees C for 45 min has increased the recovery yield of HBcAg by 2.9-fold and 42% in purity compared to that treated for 10 min. Enzyme-linked immunosorbent assay (ELISA) analysis showed that the antigenicity of the core particles was not affected by the heat treatment process.

Matched MeSH terms: Hepatitis B Core Antigens/isolation & purification*

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