Advantages of Soybean Peroxidase over Horseradish Peroxidase as the Enzyme Label in Chemiluminescent Enzyme-Linked Immunosorbent Assay of Sulfamethoxypyridazine

2010 ◽  
Vol 58 (6) ◽  
pp. 3284-3289 ◽  
Author(s):  
Ivan Yu. Sakharov ◽  
Anna N. Berlina ◽  
Anatoly V. Zherdev ◽  
Boris B. Dzantiev
Keyword(s):  
Horseradish Peroxidase ◽  
Immunosorbent Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
Soybean Peroxidase ◽  
Enzyme Label

Competitive Direct Enzyme-Linked Immunosorbent Assay for Detection of Sulfamethazine Residues in Swine Urine and Muscle Tissue

1988 ◽  
Vol 71 (6) ◽  
pp. 1137-1140 ◽  
Author(s):  
Deborah E Dixon-Holland ◽  
Stanley E Katz
Keyword(s):  
Horseradish Peroxidase ◽  
Muscle Tissue ◽  
Immunosorbent Assay ◽  
Microtiter Plate ◽  
Test System ◽  
Enzyme Linked Immunosorbent Assay ◽  
Sensitive Assay ◽  
Saline Extract ◽  
Swine Urine ◽  
Phosphate Buffered Saline

Abstract A sensitive assay for the detection of sulfamethazine in swine urine and muscle tissue using a direct competitive enzyme-linked immunosorbent assay (ELISA) has been developed. Undiluted urine or a phosphate-buffered saline extract of pork muscle tissue is mixed with an enzyme-labeled conjugate of sulfamethazine and horseradish peroxidase. The mixture is added to wells of a microtiter plate coated with antibody to sulfamethazine. After the test system is incubated, washed, and re-incubated with substrate and the reaction is stopped, the absorbance is measured at 405 nm. Levels of sulfamethazine as low as 20 ng sulfamethazine/g muscle tissue and 10 ng sulfamethazine/ mL swine urine were detected and estimated

Sandwich ELISA for Detection of Pig Meat in Raw Beef Using Antisera to Muscle Soluble Proteins

1988 ◽  
Vol 51 (10) ◽  
pp. 790-798 ◽  
Author(s):  
ROSARIO MARTÍN ◽  
JUAN I. AZCONA ◽  
CARMEN CASAS ◽  
PABLO E. HERNÁNDEZ ◽  
BERNABÉ SANZ
Keyword(s):  
Horseradish Peroxidase ◽  
Immunosorbent Assay ◽  
Peroxidase Activity ◽  
Sandwich Elisa ◽  
Soluble Proteins ◽  
Enzyme Linked Immunosorbent Assay ◽  
Specific Antibodies ◽  
Double Antibody ◽  
Colored Product ◽  
Pig Meat

A double-antibody sandwich ELISA (enzyme-linked immunosorbent assay) has been successfully developed for the detection of defined amounts of pig meat (1–50%) in raw beef. Antibodies against pig sarcoplasmic extracts were produced in rabbits. Pig-specific antibodies were affinity purified by removing antibodies which crossreacted with horse, chicken or beef extracts followed by immunoadsorption and elution from a pig-extract column. The ELISA involved capturing antigens in sarcoplasmic extracts with pig specific antibodies immobilized on 96-well plates, detecting bound antigen with pig specific, horseradish peroxidase-labeling antibody, and measuring peroxidase activity by the conversion of a clear substrate to a colored product.

Effect of Spacer and the Enzyme-Linked Immunosorbent Assay

2016 ◽  
Vol 66 (5) ◽  
pp. 471
Author(s):  
Manisha Sathe ◽  
Shruti Srivastava ◽  
Sumit Agrawal ◽  
Ramrao Ghorpade
Keyword(s):  
Immunosorbent Assay ◽  
Inhibitory Concentration ◽  
Methylphosphonic Acid ◽  
Enzyme Linked Immunosorbent Assay ◽  
Direct Immunoassay ◽  
Ic50 Value ◽  
Lower Detection ◽  
Enzyme Conjugate ◽  
Enzyme Label ◽  
Free Antigen

The effect of spacers and the enzyme-linked immunosorbent assay (ELISA) formats on the functional parameters of assays such as lower detection limit, inhibitory concentration at 50 per cent (IC50), and specificity were studied. Enzyme conjugates having hydrophobic and hydrophilic spacers were prepared using O-isopropyl methylphosphonic acid (IMPA) and horseradish peroxidase (HRP) as an enzyme label. Comparison was made with reference to enzyme conjugate without any spacer. The present investigation revealed that the presence of a hydrophilic spacer in the enzyme conjugate significantly improves the sensitivity of assays. An enhanced IC50 value achieved was 0.01 μg mL−1 for free antigen detection by direct immunoassay using hydrophilic spacers and precoating of ELISA plates by secondary antibody. The use of a hydrophilic spacer might have helped in projecting the hapten in the aqueous phase, leading to enhanced antibody binding signal and improved sensitivity of the assay.

Use of Soybean Peroxidase in Chemiluminescent Enzyme-Linked Immunosorbent Assay

2006 ◽  
Vol 54 (5) ◽  
pp. 1584-1587 ◽  
Author(s):  
Ivan Yu. Sakharov ◽  
Inna S. Alpeeva ◽  
Evgeny E. Efremov
Keyword(s):  
Immunosorbent Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
Soybean Peroxidase

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.

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.

Enzyme-Linked Immunosorbent Assay of Ochratoxin A in Wheat

1984 ◽  
Vol 67 (1) ◽  
pp. 45-49 ◽  
Author(s):  
Sungsoo C Lee ◽  
Fun S Chu
Keyword(s):  
Horseradish Peroxidase ◽  
Ochratoxin A ◽  
Immunosorbent Assay ◽  
Sodium Phosphate ◽  
Methanolic Extract ◽  
Tween 20 ◽  
Enzyme Linked Immunosorbent Assay ◽  
Final Solution ◽  
Reverse Phase ◽  
Methanol Content

Abstract An enzyme-linked immunosorbent assay (ELISA) was developed for the quantitation of ochratoxin A amended to wheat. Ochratoxin A conjugated to horseradish peroxidase (HRP) was used as an enzyme marker in the assay. At toxin levels below 30 ppb, a cleanup treatment was necessary for ELISA. Among 3 cleanup methods tested (solvent partition, Sep-Pak cartridge treatment, solvent partition and cartridge treatment), reverse phase cartridge treatment was the most simple and effective. In the analysis, ochratoxin A was extracted from wheat with methanol. The methanolic extract was diluted with water to a final 10–15% methanol content, and then passed through a cartridge. Ochratoxin A was eluted from the cartridge with 85% methanol which was then concentrated. The final solution, in 0.1M, pH 7.5 sodium phosphate–Tween 20 buffer and 5% methanol, was then subjected to ELISA. ELISA allowed minimal detection of the toxin in wheat at the 1–2 ppb level after cleanup. Recoveries of toxin added to wheat samples in the 1.0–30 ppb range were 85–90% with standard deviations of 10–15%.

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