Identification of hepatitis a virus in cell cultures by solid phase immune electron microscopy

1986 ◽  
Vol 44 ◽  
pp. 238-239
Author(s):  
C.D. Humphrey ◽  
T.L. Cromeans ◽  
E.H. Cook ◽  
D.W. Bradley
Keyword(s):  
Electron Microscopy ◽  
Liquid Phase ◽  
Cell Cultures ◽  
Rapid Detection ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Solid Phase ◽  
Immune Electron Microscopy ◽  
Detection And Identification

There is a variety of methods available for the rapid detection and identification of viruses by electron microscopy as described in several reviews. The predominant techniques are classified as direct electron microscopy (DEM), immune electron microscopy (IEM), liquid phase immune electron microscopy (LPIEM) and solid phase immune electron microscopy (SPIEM). Each technique has inherent strengths and weaknesses. However, in recent years, the most progress for identifying viruses has been realized by the utilization of SPIEM.

Application of solid-phase immune electron microscopy to study physical and stability characteristics of enterically transmitted non-a, non-b hepatitis virus

1988 ◽  
Vol 46 ◽  
pp. 80-81
Author(s):  
Charles D. Humphrey ◽  
E. H. Cook ◽  
Karen A. McCaustland ◽  
Daniel W. Bradley
Keyword(s):  
Electron Microscopy ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Solid Phase ◽  
Etiologic Agent ◽  
World Health ◽  
Health Concern ◽  
Morphological Identification ◽  
Immune Electron Microscopy

Enterically transmitted non-A, non-B hepatitis (ET-NANBH) is a type of hepatitis which is increasingly becoming a significant world health concern. As with hepatitis A virus (HAV), spread is by the fecal-oral mode of transmission. Until recently, the etiologic agent had not been isolated and identified. We have succeeded in the isolation and preliminary characterization of this virus and demonstrating that this agent can cause hepatic disease and seroconversion in experimental primates. Our characterization of this virus was facilitated by immune (IEM) and solid phase immune electron microscopic (SPIEM) methodologies.Many immune electron microscopy methodologies have been used for morphological identification and characterization of viruses. We have previously reported a highly effective solid phase immune electron microscopy procedure which facilitated identification of hepatitis A virus (HAV) in crude cell culture extracts. More recently we have reported utilization of the method for identification of an etiologic agent responsible for (ET-NANBH).

scholarly journals Enzyme-Linked Immunosorbent Assay for Detection of Hepatitis A Antigen in Stool and Antibody to Hepatitis A Antigen in Sera: Comparison with Solid-Phase Radioimmunoassay, Immune Electron Microscopy, and Immune Adherence Hemagglutination Assay

1978 ◽  
Vol 7 (2) ◽  
pp. 184-193
Author(s):  
Lars R. Mathiesen ◽  
Stephen M. Feinstone ◽  
Doris C. Wong ◽  
Peter Skinhoej ◽  
Robert H. Purcell
Keyword(s):  
Electron Microscopy ◽  
Hepatitis A ◽  
Solid Phase ◽  
Enzyme Linked Immunosorbent Assay ◽  
Immune Electron Microscopy ◽  
Specific Detection ◽  
Immune Adherence ◽  
Ag Particles ◽  
Solid Phase Radioimmunoassay ◽  
Human Stool

Previously described techniques for detection of hepatitis A antigen (HA Ag) and antibody (anti-HA) have required purified HA Ag and expensive equipment. Herein is described an enzyme-linked immunosorbent assay (ELISA) for specific detection of HA Ag in human stool filtrates and of anti-HA in sera by using selected HA Ag-containing human stool filtrates as the antigen source. Because human stools often react nonspecifically in serological tests for HA Ag, blocking with preexposure and hyperimmune anti-HA sera from a chimpanzee inoculated with hepatitis A virus was used to confirm specific detection of HA Ag. The sensitivity of ELISA was found to be comparable to that of solid-phase radioimmunoassay (SPRIA) and immune electron microscopy (IEM). Of 37 acute-phase stools collected from nine patients, 16 were positive for HA Ag by ELISA. In 13 of these, HA Ag particles were found by IEM, and an additional 3 stools negative by ELISA contained HA Ag particles by IEM. Eight control stools were negative by both ELISA and IEM. Anti-HA was measured in sera by demonstrating its ability to block binding of the enzyme conjugate to HA Ag in a stool without detectable nonspecificity. This test (blocking ELISA) was as sensitive and specific as blocking SPIRA, IEM, and immune adherence hemagglutination and, like SPRIA and IEM, detected early-developing antibody. The ELISA is simple to perform and requires only a minimum of equipment. It is useful for screening stools for HA Ag and for monitoring HA Ag during purification, as well as for detecting early and late anti-HA in sera.

scholarly journals Solid-phase enzyme-linked immunosorbent assay for detection of hepatitis A virus

1978 ◽  
Vol 8 (3) ◽  
pp. 277-282 ◽  
Author(s):  
S A Locarnini ◽  
S M Garland ◽  
N I Lehmann ◽  
R C Pringle ◽  
I D Gust
Keyword(s):  
Immunosorbent Assay ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Solid Phase ◽  
Field Work ◽  
Enzyme Linked Immunosorbent Assay ◽  
Technical Equipment ◽  
Immune Electron Microscopy ◽  
Naked Eye ◽  
Household Contacts

An enzyme-linked immunosorbent assay (ELISA) was developed for the detection of hepatitis A virus in human fecal specimens. Investigations with 88 fecal specimens from 77 patients with suspected viral hepatitis and 8 of their household contacts showed that ELISA was as specific and sensitive as radioimmunoassay and almost as sensitive as immune electron microscopy. The ELISA is quick and simple to perform, does not require sophisticated technical equipment, and can be read with the naked eye, making it suitable for field work and rapid diagnosis.

Isolation of Single Stranded DNA from Purified Hepatitis A-Virus

1978 ◽  
Vol 33 (7-8) ◽  
pp. 594-597 ◽  
Author(s):  
Bertram Flehmig ◽  
Hans Dieter Royer ◽  
Hans-Joachim Gerth
Keyword(s):  
Electron Microscopy ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Class I ◽  
Immune Electron Microscopy ◽  
Single Stranded Dna ◽  
Virus Particles ◽  
Size Classes ◽  
Length Measurements

Abstract Hepatitis A-Yirus-Single-Sranded D N A , Electron Microscopy, Length Measurements, Parvovirus Hepatitis A-virus was purified from human stools by three purification steps. Virus was identified by radioimmuno-assay and purity monitored with immune electron microscopy. Virus particles, serologically and morphologically identical, banded in CsCl in two density ranges at 1.31 — 1.34 g/cm3 and at 1.41 — 1.43 g/cm3. Virions of density 1.31 — 1.34 g/cm3 were shown to contain single-stranded D N A of different size classes. Class I 1.33 kb, class I I 4.61 kb in addition a small amount of molecules was de­ tected with lengths up to 15 kb.

scholarly journals Antibodies to hepatitis A virus: patterns by two procedures

1977 ◽  
Vol 5 (1) ◽  
pp. 110-111
Author(s):  
J Rakela ◽  
D Stevenson ◽  
V M Edwards ◽  
I Gordon ◽  
J W Mosley
Keyword(s):  
Electron Microscopy ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Immune Electron Microscopy ◽  
Immune Adherence ◽  
Low Levels

Antibody to hepatitis A virus demonstrable by immune electron microscopy appeared early but remained at low levels for several weeks. Antibody detectable by immune adherence hemagglutination was delayed.

Immune Electron Microscopy (IEM) of a Canine Adeno-Associated Virus

1974 ◽  
Vol 32 ◽  
pp. 256-257
Author(s):  
Gunter F. Thomas ◽  
M. David Hoggan
Keyword(s):  
Electron Microscopy ◽  
Cell Cultures ◽  
Complement Fixation ◽  
Hepatitis Virus ◽  
Wide Distribution ◽  
Immune Electron Microscopy ◽  
Adeno Associated Virus ◽  
Virus Like Particle ◽  
Dog Kidney ◽  
Green Monkeys

In 1968, Sugimura and Yanagawa described a small 25 nm virus like particle in association with the Matsuda strain of infectious canine hepatitis virus (ICHV). Domoto and Yanagawa showed that this particle was dependent on ICHV for its replication in primary dog kidney cell cultures (PDK) and was resistant to heating at 70°C for 10 min, and concluded that it was a canine adeno-associated virus (CAAV). Later studies by Onuma and Yanagawa compared CAAV with the known human serotypes (AAV 1, 2, 3) and AAV-4, known to be associated with African Green Monkeys. Using the complement fixation (CF) test, they found that CAAV was serologically related to AAV-3 and had wide distribution in the dog population of Japan.

Detection of Hepatitis a Virus and other Enteroviruses in Wastewater and Surface Water Samples by Gene Probe Assay

1991 ◽  
Vol 24 (2) ◽  
pp. 267-272 ◽  
Author(s):  
S. Dubrou ◽  
H. Kopecka ◽  
J. M. Lopez Pila ◽  
J. Maréchal ◽  
J. Prévot
Keyword(s):  
Surface Water ◽  
Cell Cultures ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Limit Of Detection ◽  
Blot Hybridization ◽  
Noncoding Region ◽  
Gene Probe ◽  
Dot Blot ◽  
Lower Efficiency

Enteroviruses were specifically detected by dot blot hybridization when using poliovirus type 1 (PV1) derived subgenomic radiolabeled cRNA probes (riboprobes) in environmental water specimens and in the cell cultures in which the viruses were amplificated. The riboprobe corresponding to the 5' noncoding sequence detected the majority of enteroviruses. Hepatitis A virus (HAV) was specifically detected by an HAV cRNA probe corresponding to the 5' noncoding region of its genome. By this test, the limit of detection of coxsackievirus B5 and echovirus 7 seeded in mineral water was 103 to 104 PFU/spot. In cell cultures, positive signals were observed in the lysates of cells infected by one PFU. Higher positive signals were obtained with a short PV1 probe (nt 221-670) corresponding to the 5' noncoding region, which is a well preserved sequence among the enteroviruses, than with PV1 genomic probe. Hybridization allowed a good detection of enteroviral RNAs in wastewater specimens, but with a lower efficiency in surface water. In this case, amplification of viruses in the cell cultures gave significant hybridization results.

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