Improvements to the immunoassay for detection of Rathayibacter toxicus in hay

2011 ◽  
Vol 62 (6) ◽  
pp. 523 ◽  
Author(s):  
A. M. Masters ◽  
B. Samarasinghe ◽  
M. J. Kalkhoven ◽  
G. L. den Hollander ◽  
D. G. Palmer
Keyword(s):  
Monoclonal Antibody ◽  
Horseradish Peroxidase ◽  
Immunosorbent Assay ◽  
Cost Effective ◽  
Enzyme Linked Immunosorbent Assay ◽  
Export Industry ◽  
Large Numbers ◽  
Assay Protocol ◽  
Very High ◽  
Peroxidase Conjugate

An improved protocol for the previously described enzyme-linked immunosorbent assay for Rathayibacter toxicus in hay is described. The improvements were driven mainly by the export hay industry requirement of same-day turnaround for testing of hay extracts. The preparation of hay extracts was shortened by 8 h. The time for adding samples to the enzyme-linked immunosorbent assay plates was shortened by the use of sample tubes with penetrable stoppers combined with specially designed racks. The monoclonal antibody used in the original protocol was purified and conjugated to horseradish peroxidase. This eliminated the need for a secondary step with an anti-mouse horseradish peroxidase conjugate and thereby shortened the assay by over 1 h. Results with the improved assay protocol showed a very high correlation with results obtained with the original protocol (r = 0.98). The assay is still sensitive enough to detect antigen equivalent to less than 1 average gall per kg of hay. These cost-effective changes have streamlined the testing of large numbers of samples for the presence of R. toxicus, in support of the hay export industry.

An enzyme-linked immunosorbent assay for the detection of Rathayibacter toxicus, the bacterium involved in annual ryegrass toxicity, in hay

2006 ◽  
Vol 57 (7) ◽  
pp. 731 ◽  
Author(s):  
A. M. Masters ◽  
A. R. Gregory ◽  
R. J. Evans ◽  
J. E. Speijers ◽  
S. S. Sutherland
Keyword(s):  
Monoclonal Antibody ◽  
Immunosorbent Assay ◽  
Specific Antigen ◽  
Cost Effective ◽  
Enzyme Linked Immunosorbent Assay ◽  
Annual Ryegrass ◽  
Capture Assay ◽  
Large Numbers ◽  
Soluble Polysaccharide ◽  
Annual Ryegrass Toxicity

An enzyme-linked immunosorbent assay (ELISA) for Rathayibacter toxicus is described. The development of a monoclonal antibody for a specific antigen from R. toxicus and a polyclonal antibody raised against the same R. toxicus preparation enabled a capture assay format. The assay is specific for a soluble polysaccharide produced by the bacterium and was found to be sensitive enough to detect antigen equivalent to less than one gall per kilogram of hay. The applicability of the assay to samples of pasture or hay is demonstrated. Cost-effective testing of large numbers of samples for the presence of R. toxicus is possible with the ELISA. This will assist stockowners, hay producers, and hay exporters in the management of the risk of annual ryegrass toxicity.

scholarly journals Preparation of Monoclonal Antibody and Development of Indirect Competitive Enzyme-Linked Immunosorbent Assay and Fluorescence-Linked Immunosorbent Assay for Detecting 3-Amino-5-methylmorpholino-2-oxazolidinone (AMOZ) in Edible Animal Tissue

Molecules ◽  
2021 ◽  
Vol 26 (14) ◽  
pp. 4243
Author(s):  
Yong Xie ◽  
Yarong Wang ◽  
Xueling Yan ◽  
Lu Gan ◽  
Tao Le
Keyword(s):  
Monoclonal Antibody ◽  
Immunosorbent Assay ◽  
Correlation Coefficients ◽  
Cost Effective ◽  
Enzyme Linked Immunosorbent Assay ◽  
Monitoring Methods ◽  
Animal Tissues ◽  
Coefficients Of Variation ◽  
Highly Sensitive ◽  
Ic50 Values

To monitor the illegal used of furaltadone, a highly sensitive indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and fluorescence-linked immunosorbent assay (FLISA) based on a monoclonal antibody (mAb) were developed for the detection of 3-amino-5-methylmorpholino-2-oxazolidinone (AMOZ), the major metabolite of furaltadone in animal tissues. The highly specific mAb, which was very sensitive to a nitrophenyl derivative of AMOZ (2-NP-AMOZ) with IC50 values of 0.11 and 0.09 ng/mL for ic-ELISA and FLISA, respectively, was selected for the development of immunoassays. For both the ic-ELISA and FLISA for AMOZ-spiked experiments, acceptable recovery rates of 81.1–105.3% and coefficients of variation of 4.7–9.8% were obtained. In addition, results from both ic-ELISA and FLISA methods for spiked samples’ data showed excellent correlation coefficients ranging from 0.9652 to 0.9927. Meanwhile, the proposed ic-ELISA and FLISA for thirty spiked samples were confirmed by standard LC-MS/MS with high correlation coefficients of 0.9911 and 0.9921, respectively. These results suggest that the developed ic-ELISA and FLISA are valid and cost-effective tools for high-throughput monitoring methods for AMOZ residues in animal tissues.

Comparison of Radioimmunoassay and Enzyme-linked Immunosorbent Assay for Determining Aflatoxin M1 in Milk

1981 ◽  
Vol 64 (2) ◽  
pp. 294-301
Author(s):  
James J Pestka ◽  
Yaguan Li ◽  
William O Harder ◽  
Fun S Chu
Keyword(s):  
Horseradish Peroxidase ◽  
Sensitivity And Specificity ◽  
Immunosorbent Assay ◽  
Specific Antibody ◽  
Cross Reactivity ◽  
Lower Degree ◽  
Aflatoxin M1 ◽  
Enzyme Linked Immunosorbent Assay ◽  
Whole Milk ◽  
Peroxidase Conjugate

Abstract Using a highly specific antibody against aflatoxin Mi, a radioimmunoassay (RIA) and an enzyme-linked immunosorbent assay (ELISA) were developed for the quantitation of Mi in milk. RIA was sensitive in the range of 5-50 ng per assay but was subject to interference by whole milk. Extraction and cleanup were therefore necessary for the detection of M1 in milk at 0.5 ng/mL. An ELISA procedure was developed by using an aflatoxin M1-carboxymethyl-horseradish peroxidase conjugate as the ligand. Competitive assays revealed that this system was relatively more sensitive for M1 than for B1, and had a much lower degree of cross-reactivity for aflatoxins B2, G1, G2, B2a, and aflatoxicol. As low as 0.25 ng M1/mL in artificially contaminated milk (raw, whole, skim) could be detected by ELISA in 3 h without extraction or cleanup. Because of its simplicity, sensitivity, and specificity, ELISA is the preferred method for monitoring aflatoxin M1 in milk.

Integration of Liquid Chromatography with Immunoassay: An Approach Combining the Strengths of Both Methods

1994 ◽  
Vol 77 (5) ◽  
pp. 1275-1287 ◽  
Author(s):  
Petra M Krämer ◽  
Qing X Li ◽  
Bruce D Hammock
Keyword(s):  
Liquid Chromatography ◽  
Sensitivity And Specificity ◽  
Immunosorbent Assay ◽  
Uv Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Multiresidue Analysis ◽  
Selective Method ◽  
Separation Power ◽  
Immunochemical Detection ◽  
Very High

Abstract The integration of liquid chromatography (LC) with immunochemical detection combines the superior separation power of LC and the sensitivity and specificity of immunoassays. This approach is shown with 3 LC systems (Perkin-Elmer, C18 RP, 4.6 mm; Varian, C18 RP, 1 mm microbore; Michrom, C18 RP, 1 mm microbore) Integrated with an enzyme-linked immunosorbent assay (ELISA) selective for five 4-nitrophenols. The nitrophenols were separated with the 3 LC systems with isocratic runs of 15 to 20 min. Microbore LC separation showed a 10-20 times reduction in solvent amount compared to conventional separation. LC–immunoassay was about 8- to 10-fold more sensitive compared with LC with UV detection. Integrated LC–immunoassay proved to be a very selective method when 2-methylphenol was injected with an equimolar mixture of 2-amino-4-nitrophenol and 3-methyl-4-nrtrophenol; 2-methy I phenol does not crossreact with the serum used. Only 2 peaks could be seen in the detection, even when 2-methylphenol was present in very high amounts (3000 pmol). Further, the EUSA-LC detection proved to be selective and sensitive for complex matrixes. 2-Amlno-4-nitrophenol was clearly identified in spiked extracts of soil and plant, even when a very small amount (2.4 ng) was injected. Although LC–immunoassay is more labor intensive than LC with UV detection, it offers great advantages in multiresidue analysis and is generally applicable for peak confirmation.

scholarly journals Unreliable Measurement of Basophil Maximum Leukotriene Release with the Bühlmann CAST 2000 Enzyme-Linked Immunosorbent Assay Kit

2006 ◽  
Vol 13 (3) ◽  
pp. 420-422 ◽  
Author(s):  
S. E. Burastero ◽  
C. Paolucci ◽  
D. Breda ◽  
G. Monasterolo ◽  
R. E. Rossi ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Immunosorbent Assay ◽  
False Positive ◽  
Positive Control ◽  
Enzyme Linked Immunosorbent Assay ◽  
Positive Results

ABSTRACT The Bühlmann CAST 2000 enzyme-linked immunosorbent assay is a potentially useful assay for measuring sulfidoleukotrienes released in vitro by allergen-challenged basophils. However, we observed that the positive-control reagent yielded positive signals in cell-free systems. These false-positive results depended on using a mouse anti-FcεRI monoclonal antibody and were prevented by degranulation-inducing reagents other than mouse monoclonal antibodies.

Development of a monoclonal antibody-based enzyme-linked immunosorbent assay for the analysis of the new fungicide 2-allylphenol in strawberry fruits

2010 ◽  
Vol 120 (4) ◽  
pp. 1178-1184 ◽  
Author(s):  
Yuanyuan Xia ◽  
Qing X. Li ◽  
Shuangjun Gong ◽  
Yong Li ◽  
Yongsong Cao ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Immunosorbent Assay ◽  
Enzyme Linked Immunosorbent Assay
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