scholarly journals Production of antiserum to the E. coli-expressed coat protein of Apple stem grooving virus in citrus and its application for the diagnosis by enzyme-linked immunosorbent assay.

2004 ◽  
Vol 70 (2) ◽  
pp. 123-127 ◽  
Author(s):  
Y. IDE ◽  
K. KUROKI ◽  
N. TASHIRO ◽  
H. MAGOME ◽  
N. YOSHIKAWA ◽  
...  
Keyword(s):  
Coat Protein ◽  
Immunosorbent Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
E Coli ◽  
Apple Stem Grooving Virus ◽  
Apple Stem

Use of a Shiga Toxin (Stx)-Enzyme-Linked Immunosorbent Assay and Immunoblot for Detection and Isolation of Stx-Producing Escherichia coli from Naturally Contaminated Beef

2000 ◽  
Vol 63 (9) ◽  
pp. 1167-1172 ◽  
Author(s):  
HEBA NASHED ATALLA ◽  
ROGER JOHNSON ◽  
SCOTT MCEWEN ◽  
R. W. USBORNE ◽  
C. L. GYLES
Keyword(s):  
Escherichia Coli ◽  
Vero Cell ◽  
Immunosorbent Assay ◽  
Shiga Toxin ◽  
Total Coliform ◽  
Complete Agreement ◽  
Enzyme Linked Immunosorbent Assay ◽  
Spearman Correlation ◽  
Southern Ontario ◽  
E Coli

The purpose of this study was to evaluate an enzyme-linked immunosorbent assay (ELISA) and an immunoblot procedure for detection and isolation of Shiga toxin-producing Escherichia coli (STEC) from beef, and to correlate the presence of STEC in beef with E. coli and total coliform counts. A total of 120 samples of boneless beef supplied to a meat processor in southern Ontario were tested for the presence of STEC, E. coli, and total coliforms. Following enrichment in modified tryptic soy broth, samples were screened for Shiga toxin (Stx) by a Stx-ELISA and a Vero cell assay (VCA). Samples that were positive in the Stx-ELISA were subjected to the Stx-immunoblot for STEC isolation. Overall, 33.3% of samples were positive in the VCA, and 34.2% were positive in the Stx-ELISA. There was almost complete agreement between the Stx-ELISA and the VCA results (kappa = 0.98). The sensitivity and specificity of the Stx-ELISA with respect to the VCA were 100% and 98.75%, respectively. STEC were isolated by the Stx-immunoblot from 87.8% of the samples that were positive in the Stx-ELISA. The STEC isolates belonged to 19 serotypes, with serotype O113:H21 accounting for 10 of 41 isolates. No STEC of serotype O157:H7 were isolated. There was a significant correlation between E. coli counts and total coliform counts (Spearman correlation coefficient = 0.68, P < 0.01). The E. coli count was positively correlated with detection of STEC by both the Stx-ELISA and the VCA (P < 0.01).

Evaluation of Three Commercially Available Enzyme-Linked Immunosorbent Assay Kits for Detection of Shiga Toxin

2009 ◽  
Vol 72 (4) ◽  
pp. 741-747 ◽  
Author(s):  
JOHN WILLFORD ◽  
KENNETH MILLS ◽  
LAWRENCE D. GOODRIDGE
Keyword(s):  
Escherichia Coli ◽  
Immunosorbent Assay ◽  
Shiga Toxin ◽  
Screening Method ◽  
Enzyme Linked Immunosorbent Assay ◽  
Microplate Assay ◽  
E Coli ◽  
Elisa Kit ◽  
Pure Cultures ◽  
Order Of Magnitude

Three commercially available Shiga toxin (Stx) enzyme-linked immunosorbent assay (ELISA) kits were evaluated for their ability to detect Stx in pure cultures of Stx-producing Escherichia coli (specificity). The detection limits (sensitivity) of each ELISA kit were also evaluated. Seventy-eight Stx-producing E. coli (STEC) isolates that produced Stx1, Stx2, or Stx1 and Stx2 variants were examined in this study. The specificities of the tests were comparable, and the sensitivities of two of the tests (Premier EHEC and rBiopharm Ridascreen Verotoxin Enzyme Immunoassay) were within the same order of magnitude. The ProSpecT Shiga Toxin E. coli Microplate Assay was approximately 10-fold less sensitive. The inability of all three tests to detect the Stx2d and Stx2e variants indicated that some STEC strains may not be detected by Stx ELISA. The ability of the Premier EHEC ELISA to detect toxin in artificially inoculated bovine fecal samples (following enrichment) indicated that this kit may be used to screen cattle for the presence of Stx as an indicator of the presence of STEC. In particular, such a screening method could be useful during the summer, when the number of STEC-positive animals and the number of STEC that they shed increase.

scholarly journals Enzyme-linked immunosorbent assay for detection of Escherichia coli heat-labile enterotoxin

1977 ◽  
Vol 6 (5) ◽  
pp. 439-444 ◽  
Author(s):  
R H Yolken ◽  
H B Greenberg ◽  
M H Merson ◽  
R B Sack ◽  
A Z Kapikian
Keyword(s):  
Escherichia Coli ◽  
Immunosorbent Assay ◽  
Radioactive Isotope ◽  
Enzyme Linked Immunosorbent Assay ◽  
Adrenal Cell ◽  
Elisa System ◽  
Culture Techniques ◽  
E Coli ◽  
Heat Labile ◽  
Tissue Culture Techniques

The development of an enzyme-linked immunosorbent assay (ELISA) for the detection of heat-labile Escherichia coli enterotoxin is described. The assay, which is based on the immunological similarity between Vibrio cholerae toxin and heat-labile E. coli enterotoxin, is similar in design to a radioimmunoassay but utilizes enzyme-labeled rather than radioactive isotope-labeled reagents. The ELISA system is as sensitive as both radioimmunoassay and the y-1 adrenal cell assay for the detection of heat-labile E. coli enterotoxin but requires neither radioactive reagents nor tissue culture techniques. The ELISA is easy to perform and is adaptable for use in small laboratories.

scholarly journals Sequence analysis of the capsid protein gene of an isolate of Apple stem grooving virus, and its survey in Southern Brazil

2001 ◽  
Vol 26 (3) ◽  
pp. 655-659 ◽  
Author(s):  
OSMAR NICKEL ◽  
THOR V.M. FAJARDO ◽  
WILHELM JELKMANN ◽  
GILMAR B. KUHN
Keyword(s):  
Amino Acid ◽  
Coat Protein ◽  
Coat Protein Gene ◽  
Large Scale ◽  
Protein Gene ◽  
Amino Acid Sequences ◽  
Close Relative ◽  
Apple Stem Grooving Virus ◽  
Apple Stem ◽  
High Amino Acid

Apple stem grooving virus (ASGV) is one of the most important viruses infecting fruit trees. This study aimed at the molecular characterization of ASGV infecting apple (Malus domestica) plants in Santa Catarina (SC). RNA extracted from plants infected with isolate UV01 was used as a template for RT-PCR using specific primers. An amplified DNA fragment of 755 bp was sequenced. The coat protein gene of ASGV isolate UV01 contains 714 nucleotides, coding for a protein of 237 amino acids with a predicted Mr of approximately 27 kDa. The nucleotide and the deduced amino acid sequences of the coat protein gene showed identities of 90.9% and 97.9%, respectively, with a Japanese isolate of ASGV. Very high amino acid homologies (98.7%) were also found with Citrus tatter leaf capillovirus (CTLV), a very close relative of ASGV. These results indicate low coat protein gene variability among Capillovirus isolates from distinct regions. In a restricted survey, mother stocks in orchards and plants introduced into the country for large scale fruit production were indexed and shown to be infected by ASGV (20%), usually in a complex with other (latent) apple viruses (80%).

scholarly journals Occurrence of Apple stem grooving virus in commercial apple seedlings and analysis of its coat protein sequence

2016 ◽  
Vol 43 (1) ◽  
pp. 21-27 ◽  
Author(s):  
Jae-Yeong Han ◽  
Chan-Hwan Park ◽  
Eun-Yeong Seo ◽  
Jung-Kyu Kim ◽  
John Hammond ◽  
...  
Keyword(s):  
Coat Protein ◽  
Protein Sequence ◽  
Coat Protein Sequence ◽  
Apple Stem Grooving Virus ◽  
Apple Stem

scholarly journals Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis

2004 ◽  
Vol 29 (5) ◽  
pp. 558-562 ◽  
Author(s):  
Osmar Nickel ◽  
Maria L.P.N. Targon ◽  
Thor V.M. Fajardo ◽  
Marcos A. Machado ◽  
Ana P. Trivilin
Keyword(s):  
Escherichia Coli ◽  
Coat Protein ◽  
Fusion Protein ◽  
Polyclonal Antibodies ◽  
Molecular Techniques ◽  
Bacterial Cells ◽  
Additional Tool ◽  
Apple Stem Grooving Virus ◽  
Apple Stem ◽  
Specificity And Sensitivity

The coat protein gene of Apple stem grooving virus (ASGV) was amplified by RT-PCR, cloned, sequenced and subcloned in the expression vector pMal-c2. This plasmid was used to transform Escherichia coli BL21c+ competent cells. The ASGV coat protein (cp) was expressed as a fusion protein containing a fragment of E. coli maltose binding protein (MBP). Bacterial cells were disrupted by sonication and the ASGVcp/MBP fusion protein was purified by amylose resin affinity chromatography. Polyclonal antibodies from rabbits immunized with the fusion protein gave specific reactions to ASGV from infected apple (Malus domestica) cv. Fuji Irradiada and Chenopodium quinoa at dilutions of up to 1:1,000 and 1:2,000, respectively, in plate trapped ELISA. The ASGVcp/MBP fusion protein reacted to a commercial antiserum against ASGV in immunoblotting assay. The IgG against ASGVcp/MBP performed favorably in specificity and sensitivity to the virus. This method represents an additional tool for the efficient ASGV-indexing of apple propagative and mother stock materials, and for use in support of biological and molecular techniques.

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