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A study of the frequency of infection of peripheral blood mononuclear cells of chronic hepatitis B virus carriers using the polymerase chain reaction and hybridization analysis

Abstract

Peripheral blood mononuclear cells (PBMCs) from 100 consecutive chronic carriers of the Hepatitis B virus (HBV) were analysed to determine the frequency of infection of the PBMCs. Cells were isolated using a ficoll gradient and DNA extracted by phenol following an overnight incubation with proteinase K and tween-20. Target nucleic acid were amplified using a set of primers spanning the S region of the viral genome between nucleotides 79 and 761. Following amplification, the samples were gel electrophoresed and the fragments visualized by ethidium bromide staining. The presence of a fragment of about 720 bp was taken as indicative of specific amplification of the HBV nucleic acid sequences. Specificity of amplification was confirmed by hybridization analysis using virus specific probes. Thirty-six out of 41 (87.8%) HBeAg seropositive cases and 15/54 (27.8%) anti-HBe positive cases had HBV DNA detectable by gel electrophoresis. Following hybridization all carriers were found to harbour the virus in their mononuclear cells. The sensitivity using ethidium bromide staining to visualize the amplified sequences was about 1 pg. With hybridization analysis, sensitivity was increased about 10{sup 5}-fold. (author). 12 refs, 1 fig., 1 tab.
Authors:
Yap, S F; Wong, P W; Goh, K L; Wong, N W [1] 
  1. Malaya Univ., Kuala Lumpur (Malaysia). Depts. of Pathology and Dept. of Medicine
Publication Date:
May 01, 1994
Product Type:
Technical Report
Report Number:
IAEA-TECDOC-748
Reference Number:
SCA: 553003; 550601; PA: AIX-26:032924; EDB-95:060064; ERA-20:013922; SN: 95001365262
Resource Relation:
Other Information: PBD: May 1994; Related Information: Is Part Of Radionuclides in molecular technology for diagnosis of communicable diseases; PB: 131 p.
Subject:
55 BIOLOGY AND MEDICINE, BASIC STUDIES; HEPATITIS; HYBRIDIZATION; POLYMERASE CHAIN REACTION; DNA; ELECTROPHORESIS; LEUKOCYTES; PATIENTS; VIRAL DISEASES
OSTI ID:
25269
Research Organizations:
International Atomic Energy Agency, Vienna (Austria)
Country of Origin:
IAEA
Language:
English
Other Identifying Numbers:
Journal ID: ISSN 1011-4289; Other: ON: DE95624121; TRN: XA9437534032924
Availability:
INIS; OSTI as DE95624121
Submitting Site:
INIS
Size:
pp. 93-96
Announcement Date:
Jan 15, 2004

Citation Formats

Yap, S F, Wong, P W, Goh, K L, and Wong, N W. A study of the frequency of infection of peripheral blood mononuclear cells of chronic hepatitis B virus carriers using the polymerase chain reaction and hybridization analysis. IAEA: N. p., 1994. Web.
Yap, S F, Wong, P W, Goh, K L, & Wong, N W. A study of the frequency of infection of peripheral blood mononuclear cells of chronic hepatitis B virus carriers using the polymerase chain reaction and hybridization analysis. IAEA.
Yap, S F, Wong, P W, Goh, K L, and Wong, N W. 1994. "A study of the frequency of infection of peripheral blood mononuclear cells of chronic hepatitis B virus carriers using the polymerase chain reaction and hybridization analysis." IAEA.
@misc{etde_25269,
title = {A study of the frequency of infection of peripheral blood mononuclear cells of chronic hepatitis B virus carriers using the polymerase chain reaction and hybridization analysis}
author = {Yap, S F, Wong, P W, Goh, K L, and Wong, N W}
abstractNote = {Peripheral blood mononuclear cells (PBMCs) from 100 consecutive chronic carriers of the Hepatitis B virus (HBV) were analysed to determine the frequency of infection of the PBMCs. Cells were isolated using a ficoll gradient and DNA extracted by phenol following an overnight incubation with proteinase K and tween-20. Target nucleic acid were amplified using a set of primers spanning the S region of the viral genome between nucleotides 79 and 761. Following amplification, the samples were gel electrophoresed and the fragments visualized by ethidium bromide staining. The presence of a fragment of about 720 bp was taken as indicative of specific amplification of the HBV nucleic acid sequences. Specificity of amplification was confirmed by hybridization analysis using virus specific probes. Thirty-six out of 41 (87.8%) HBeAg seropositive cases and 15/54 (27.8%) anti-HBe positive cases had HBV DNA detectable by gel electrophoresis. Following hybridization all carriers were found to harbour the virus in their mononuclear cells. The sensitivity using ethidium bromide staining to visualize the amplified sequences was about 1 pg. With hybridization analysis, sensitivity was increased about 10{sup 5}-fold. (author). 12 refs, 1 fig., 1 tab.}
place = {IAEA}
year = {1994}
month = {May}
}