scholarly journals Significance of Hepatitis B Virus Diagnosis by Real-time Polymerase Chain Reaction over Serological Markers in Hepatitis B Virus Patients

2020 ◽  
Vol 4 (1) ◽  
pp. 52-56
Author(s):  
Amer A. Khaleel ◽  
Salah T. Jalal ◽  
Saeed G. Hussain
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Quantitative Estimation ◽  
Hbv Dna ◽  
Hbv Infection ◽  
Chain Reaction ◽  
Hbs Antigen ◽  
B Virus ◽  
Nucleic Acid Assay ◽  
Polymerase Chain

Hepatitis B virus (HBV) is the leading cause of viral hepatitis, as currently over 2 billion people have HBV infection worldwide. Nucleic acid assay and quantitative hepatitis B surface antigen (HBsAg) have been developed for diagnostic and therapeutic monitoring of patients with HBV infection. These tests might also show correlation between HBV DNA and HBs serostatus. The study aimed to find and analyze the frequency and impact of HBsAg seropositivity among patients revealed HBV DNA negative level through quantitative estimation of both seromarkers. Real-time polymerase chain reaction (RT-PCR) and Elecsys assays were used for quantitative estimation of HBV DNA and HBs antigen, respectively. A total of 256 blood samples were used from patients referred for either diagnostic purpose and/or HBV viral load monitoring after antiviral therapy. Blood profile analysis showed 12.26% HBs antigen seropositivity among patients revealed negative for nucleic acid assay for HBV DNA. Positive HBs antigen titers ranged from 1000–50,000 COI, with seronegative anti-HBs antibody test for all samples tested positive for HBs antigen. This study delineated that negative or undetectable quantitation of HBV DNA level does not exclude HBV infection; as the level might fluctuate in different phases of HBV replication. This gives an impression and raising a question about significance of replacing test for HBsAg with quantitation of HBV DNA PCR assay. Thus, the study refers to a special HBV profile outside the classical pattern.

Molecular Analysis of Hepatitis B Virus DNA and Mutation Status in Patients Under Lamivudine [(-) 2' 3'-dideoxy-3'-thiacytidine] Therapy

1970 ◽  
Vol 4 (2) ◽  
Author(s):  
Mamun Ahmed ◽  
Shaila Nazneen ◽  
Anwarul Azim Akhand
Keyword(s):  
Polymerase Chain Reaction ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Hbv Dna ◽  
Chain Reaction ◽  
Lamivudine Resistance ◽  
Lamivudine Therapy ◽  
Sequence Detection ◽  
B Virus ◽  
Polymerase Chain

Quantitative analysis of HBV-DNA is extensively used worldwide for monitoring of lamivudine therapy of Hepatitis B virus (HBV) infection. We have analyzed the quantity of HBV-DNA during lamivudine therapy and investigated the relationship of lamivudine resistance to mutation type. Ninety-one hepatitis B patients were enrolled in the study where Real Time Polymerase Chain Reaction (PCR) did estimation of HBV DNA and mutation was analyzed by sequence detection via PCR. HBV-DNA was detected in the serum of 96.7% (88/91) patients with mean viral load ranging from 1 ´ 105 to 1 ´ 109. More than 80% patients responded to and 17.3% patients showed resistance to lamivudine therapy. All lamivudine resistant patients had HBV YMDD mutation of either rtL180M/M204V or rtL180M/ M204I type. PCR based analysis of HBV DNA and sequence based mutation detection can be a practically feasible approach in Bangladesh to monitor hepatitis B patients under lamivudine therapy. Key words: Lamivudine, Hepatitis B Virus (HBV), Polymerase Chain Reaction (PCR), Mutation Dhaka Univ. J. Pharm. Sci. Vol.4(2) 2005 The full text is of this article is available at the Dhaka Univ. J. Pharm. Sci. website

scholarly journals Detection of hepatitis B virus in plasma using flow cytometric analyses of polymerase chain reaction-amplified DNA incorporating digoxigenin-11- dUTP

Blood ◽  
1993 ◽  
Vol 81 (4) ◽  
pp. 1083-1088 ◽  
Author(s):  
G Yang ◽  
PP Ulrich ◽  
RA Aiyer ◽  
BD Rawal ◽  
GN Vyas
Keyword(s):  
Polymerase Chain Reaction ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Hbv Dna ◽  
Viral Envelope ◽  
Chain Reaction ◽  
Flow Cytometric ◽  
B Virus ◽  
Pcr Products ◽  
Polymerase Chain

Abstract Blood donations are routinely screened by multiple serologic assays for antigens/antibodies associated with infection by blood-borne viruses, including hepatitis B virus (HBV), hepatitis C virus (HCV), human immunodeficiency viruses (HIV-1 and HIV-2), and human T-cell lymphotropic virus (HTLV-I and HTLV-II). A direct detection of these viruses would be more effective for the prevention of transfusion- transmitted infections than the indirect measurement of the variable host immune response to these agents. Because the polymerase chain reaction (PCR) for viral gene amplification offers the most sensitive and direct means of detecting viruses in blood, we have developed a nonisotopic PCR procedure for the detection of HBV, chosen as a prototype. The problems, common to previously described PCR methods, of nucleic acid extraction and inhibition of the PCR by plasma proteins were overcome by isolation of HBV from plasma by means of 450-microns polystyrene beads covalently coated with monoclonal antibody to the Pre- S1 region of the viral envelope protein. Detergent lysis and proteinase K digestion of the immunocaptured virions isolated from plasma released the HBV DNA. A modified PCR-amplification protocol, incorporating digoxigenin-labeled dUTP in the amplified gene products followed by hybridization with a specific biotinylated oligonucleotide probe bound to streptavidin-coated 2.8-microns magnetic beads, allowed flow cytometric analyses of HBV-specific PCR products by means of antibodies to digoxigenin labeled with fluorescein isothiocyanate. The endpoint serial dilutions of pedigreed human plasma samples containing chimpanzee infectious dose (CID50) of 10(7) for adw and CID50 of 10(7.5) for the ayw subtypes were compared in repeated testing of PCR products by our immunoreactive bead (PCR-IRB) assay. HBV DNA was consistently detected in a 5 x 10(-10) dilution of each sample. In testing 20 coded specimens of blood donors, with or without serologic markers of HBV infection, the PCR-IRB was specific and more sensitive than the PCR analyses by slot blot hybridization with radioactive probe. The PCR-IRB assay can be adapted for simultaneous detection of multiple blood-borne viruses by an automated flow cytometric analysis system.

scholarly journals Detection of hepatitis B virus in plasma using flow cytometric analyses of polymerase chain reaction-amplified DNA incorporating digoxigenin-11- dUTP

Blood ◽  
1993 ◽  
Vol 81 (4) ◽  
pp. 1083-1088
Author(s):  
G Yang ◽  
PP Ulrich ◽  
RA Aiyer ◽  
BD Rawal ◽  
GN Vyas
Keyword(s):  
Polymerase Chain Reaction ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Hbv Dna ◽  
Viral Envelope ◽  
Chain Reaction ◽  
Flow Cytometric ◽  
B Virus ◽  
Pcr Products ◽  
Polymerase Chain

Blood donations are routinely screened by multiple serologic assays for antigens/antibodies associated with infection by blood-borne viruses, including hepatitis B virus (HBV), hepatitis C virus (HCV), human immunodeficiency viruses (HIV-1 and HIV-2), and human T-cell lymphotropic virus (HTLV-I and HTLV-II). A direct detection of these viruses would be more effective for the prevention of transfusion- transmitted infections than the indirect measurement of the variable host immune response to these agents. Because the polymerase chain reaction (PCR) for viral gene amplification offers the most sensitive and direct means of detecting viruses in blood, we have developed a nonisotopic PCR procedure for the detection of HBV, chosen as a prototype. The problems, common to previously described PCR methods, of nucleic acid extraction and inhibition of the PCR by plasma proteins were overcome by isolation of HBV from plasma by means of 450-microns polystyrene beads covalently coated with monoclonal antibody to the Pre- S1 region of the viral envelope protein. Detergent lysis and proteinase K digestion of the immunocaptured virions isolated from plasma released the HBV DNA. A modified PCR-amplification protocol, incorporating digoxigenin-labeled dUTP in the amplified gene products followed by hybridization with a specific biotinylated oligonucleotide probe bound to streptavidin-coated 2.8-microns magnetic beads, allowed flow cytometric analyses of HBV-specific PCR products by means of antibodies to digoxigenin labeled with fluorescein isothiocyanate. The endpoint serial dilutions of pedigreed human plasma samples containing chimpanzee infectious dose (CID50) of 10(7) for adw and CID50 of 10(7.5) for the ayw subtypes were compared in repeated testing of PCR products by our immunoreactive bead (PCR-IRB) assay. HBV DNA was consistently detected in a 5 x 10(-10) dilution of each sample. In testing 20 coded specimens of blood donors, with or without serologic markers of HBV infection, the PCR-IRB was specific and more sensitive than the PCR analyses by slot blot hybridization with radioactive probe. The PCR-IRB assay can be adapted for simultaneous detection of multiple blood-borne viruses by an automated flow cytometric analysis system.

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