SrasV1, Main, Exploration, bibRecord, 005387

Development of a Western Blot Assay for Detection of Antibodies against Coronavirus Causing Severe Acute Respiratory Syndrome

Identifieur interne : 005387 ( Main/Exploration ); précédent : 005386; suivant : 005388

Development of a Western Blot Assay for Detection of Antibodies against Coronavirus Causing Severe Acute Respiratory Syndrome

Auteurs : Qigai He ; Kooi Hoong Chong ; Hiok Hee Chng ; Bernard Leung ; Ai Ee Ling ; Ting Wei ; Shzu-Wei Chan ; Eng Eong Ooi ; Jimmy Kwang

Source :

Clinical and Diagnostic Laboratory Immunology [ 1071-412X ] ; 2004.

RBID : PMC:371214

Descripteurs français

KwdFr :
- Animaux, Anticorps antiviraux (analyse), Chiens, Clonage moléculaire, Humains, Poulets, Protéines recombinantes (génétique), Protéines recombinantes (immunologie), Suidae, Syndrome respiratoire aigu sévère (diagnostic), Syndrome respiratoire aigu sévère (immunologie), Technique de Western (), Virus du SRAS (génétique), Virus du SRAS (immunologie), Virus du SRAS (isolement et purification), Épitopes.
MESH :
- analyse : Anticorps antiviraux.
- diagnostic : Syndrome respiratoire aigu sévère.
- génétique : Protéines recombinantes, Virus du SRAS.
- immunologie : Protéines recombinantes, Syndrome respiratoire aigu sévère, Virus du SRAS.
- isolement et purification : Virus du SRAS.
- Animaux, Chiens, Clonage moléculaire, Humains, Poulets, Suidae, Technique de Western, Épitopes.

English descriptors

KwdEn :
- Animals, Antibodies, Viral (analysis), Blotting, Western (methods), Chickens, Cloning, Molecular, Dogs, Epitopes, Humans, Recombinant Proteins (genetics), Recombinant Proteins (immunology), SARS Virus (genetics), SARS Virus (immunology), SARS Virus (isolation & purification), Severe Acute Respiratory Syndrome (diagnosis), Severe Acute Respiratory Syndrome (immunology), Swine.
MESH :
- chemical , analysis : Antibodies, Viral.
- chemical , genetics : Recombinant Proteins.
- chemical , immunology : Recombinant Proteins.
- diagnosis : Severe Acute Respiratory Syndrome.
- genetics : SARS Virus.
- immunology : SARS Virus, Severe Acute Respiratory Syndrome.
- isolation & purification : SARS Virus.
- methods : Blotting, Western.
- Animals, Chickens, Cloning, Molecular, Dogs, Epitopes, Humans, Swine.

Abstract

To identify a major antigenic determinant for use in the development of a rapid serological diagnostic test for severe acute respiratory syndrome (SARS) coronavirus infection and to study the immune response during SARS coronavirus infection in humans, we cloned the full length and six truncated fragments of the nucleocapsid gene, expressed them, and purified them as glutathione S-transferase-tagged recombinant proteins. The reactivities of the recombinant proteins to a panel of antibodies containing 33 SARS coronavirus-positive sera and 66 negative sera and to antibodies against other animal coronaviruses were screened. A truncated 195-amino-acid fragment from the C terminus of the nucleocapsid protein (N195) was identified that had a strong ability to detect antibodies against SARS coronavirus. No cross-reaction was found between the N195 protein and antibodies against chicken, pig, and canine coronaviruses. The N195 protein was used to develop a Western blot assay to detect antibodies against SARS coronavirus in 274 clinically blinded samples. The specificity and sensitivity of this test were 98.3 and 90.9%, respectively. The correlation between our Western blotting assay and an immunofluorescence assay (IFA) was also analyzed. The results of our Western blot assay and IFA for the detection of SARS coronavirus-positive sera were the same. Thus, the N195 protein was identified as a suitable protein to be used as an antigen in Western blot and other possible assays for the detection of SARS coronavirus infection.

Url:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC371214

DOI: 10.1128/CDLI.11.2.417-422.2004
PubMed: 15013997
PubMed Central: 371214

Affiliations:

Le document en format XML

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<term>Cloning, Molecular</term>
<term>Dogs</term>
<term>Epitopes</term>
<term>Humans</term>
<term>Recombinant Proteins (genetics)</term>
<term>Recombinant Proteins (immunology)</term>
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<term>SARS Virus (immunology)</term>
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<term>Poulets</term>
<term>Protéines recombinantes (génétique)</term>
<term>Protéines recombinantes (immunologie)</term>
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<term>Syndrome respiratoire aigu sévère (diagnostic)</term>
<term>Syndrome respiratoire aigu sévère (immunologie)</term>
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<term>Virus du SRAS (génétique)</term>
<term>Virus du SRAS (immunologie)</term>
<term>Virus du SRAS (isolement et purification)</term>
<term>Épitopes</term>
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</keywords>
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<front><div type="abstract" xml:lang="en"><p>To identify a major antigenic determinant for use in the development of a rapid serological diagnostic test for severe acute respiratory syndrome (SARS) coronavirus infection and to study the immune response during SARS coronavirus infection in humans, we cloned the full length and six truncated fragments of the nucleocapsid gene, expressed them, and purified them as glutathione <italic>S</italic>
-transferase-tagged recombinant proteins. The reactivities of the recombinant proteins to a panel of antibodies containing 33 SARS coronavirus-positive sera and 66 negative sera and to antibodies against other animal coronaviruses were screened. A truncated 195-amino-acid fragment from the C terminus of the nucleocapsid protein (N195) was identified that had a strong ability to detect antibodies against SARS coronavirus. No cross-reaction was found between the N195 protein and antibodies against chicken, pig, and canine coronaviruses. The N195 protein was used to develop a Western blot assay to detect antibodies against SARS coronavirus in 274 clinically blinded samples. The specificity and sensitivity of this test were 98.3 and 90.9%, respectively. The correlation between our Western blotting assay and an immunofluorescence assay (IFA) was also analyzed. The results of our Western blot assay and IFA for the detection of SARS coronavirus-positive sera were the same. Thus, the N195 protein was identified as a suitable protein to be used as an antigen in Western blot and other possible assays for the detection of SARS coronavirus infection.</p>
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<tree><noCountry><name sortKey="Chan, Shzu Wei" sort="Chan, Shzu Wei" uniqKey="Chan S" first="Shzu-Wei" last="Chan">Shzu-Wei Chan</name>
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<name sortKey="He, Qigai" sort="He, Qigai" uniqKey="He Q" first="Qigai" last="He">Qigai He</name>
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Development of a Western Blot Assay for Detection of Antibodies against Coronavirus Causing Severe Acute Respiratory Syndrome

Development of a Western Blot Assay for Detection of Antibodies against Coronavirus Causing Severe Acute Respiratory Syndrome

Source :

Descripteurs français

English descriptors

Abstract

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