Dataset Information

A New ELISA Using the ANANAS Technology Showing High Sensitivity to diagnose the Bovine Rhinotracheitis from Individual Sera to Pooled Milk.

ABSTRACT: Diagnostic tests for veterinary surveillance programs should be efficient, easy to use and, possibly, economical. In this context, classic Enzyme linked ImmunoSorbent Assay (ELISA) remains the most common analytical platform employed for serological analyses. The analysis of pooled samples instead of individual ones is a common procedure that permits to certify, with one single test, entire herds as "disease-free". However, diagnostic tests for pooled samples need to be particularly sensitive, especially when the levels of disease markers are low, as in the case of anti-BoHV1 antibodies in milk as markers of Infectious Bovine Rhinotracheitis (IBR) disease. The avidin-nucleic-acid-nanoassembly (ANANAS) is a novel kind of signal amplification platform for immunodiagnostics based on colloidal poly-avidin nanoparticles that, using model analytes, was shown to strongly increase ELISA test performance as compared to monomeric avidin. Here, for the first time, we applied the ANANAS reagent integration in a real diagnostic context. The monoclonal 1G10 anti-bovine IgG1 antibody was biotinylated and integrated with the ANANAS reagents for indirect IBR diagnosis from pooled milk mimicking tank samples from herds with IBR prevalence between 1 to 8%. The sensitivity and specificity of the ANANAS integrated method was compared to that of a classic test based on the same 1G10 antibody directly linked to horseradish peroxidase, and a commercial IDEXX kit recently introduced in the market. ANANAS integration increased by 5-fold the sensitivity of the 1G10 mAb-based conventional ELISA without loosing specificity. When compared to the commercial kit, the 1G10-ANANAS integrated method was capable to detect the presence of anti-BHV1 antibodies from bulk milk of gE antibody positive animals with 2-fold higher sensitivity and similar specificity. The results demonstrate the potentials of this new amplification technology, which permits improving current classic ELISA sensitivity limits without the need for new hardware investments.

SUBMITTER: Casarin E

PROVIDER: S-EPMC4712047 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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A New ELISA Using the ANANAS Technology Showing High Sensitivity to diagnose the Bovine Rhinotracheitis from Individual Sera to Pooled Milk.

Casarin Elisabetta E Lucchese Laura L Grazioli Santina S Facchin Sonia S Realdon Nicola N Brocchi Emiliana E Morpurgo Margherita M Nardelli Stefano S

PloS one 20160113 1

Diagnostic tests for veterinary surveillance programs should be efficient, easy to use and, possibly, economical. In this context, classic Enzyme linked ImmunoSorbent Assay (ELISA) remains the most common analytical platform employed for serological analyses. The analysis of pooled samples instead of individual ones is a common procedure that permits to certify, with one single test, entire herds as "disease-free". However, diagnostic tests for pooled samples need to be particularly sensitive, e ...[more]

PMID: 26761815

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Project description:BACKGROUND:The detection of antibodies against capripoxvirus has become easier with a commercially available ELISA validated for serum and plasma. In order to explore its suitability for immunological investigations on alternative samples, this study targeted milk as sample matrix available through non-invasive sampling. METHODS:Samples for this study were collected from dairy cows vaccinated against LSD in an area without reported LSD virus circulation. Paired serum and milk (individual and bulk) samples were tested by ELISA without and with modifications of the sample incubation time for the milk samples. For the evaluation of the test specificity, 352 milk samples from a milk repository in Germany were used as negative control. Receiver operating characteristic analysis was performed for determination of the Youden index and determination of the most suitable cut-off value for maximum specificity. RESULTS:From 154 analyzed serum samples from Serbia, 75 were detected as positive in the ELISA. Sensitivity and specificity of the ELISA test for milk samples reached values of 88 to 91% using Youden criteria. A cut-off of 10 was determined aiming for maximum specificity. This cut-off value was used for further analysis. Using the protocol for serum, out of 154 milk samples, 38 were detected as positive, number of positive detected milk samples increase up to 48 with modified protocol. Milk samples from Germany reacted negative, except two samples that had borderline results using modified protocol. Significant statistical difference (p?<?0.05) was observed between two incubation protocols. The detection of LSD-specific antibodies from bulk milk samples (pools of 2-10 individuals) came along with a reduced sensitivity over the sample of individual animals. CONCLUSIONS:Results show that the detection of capripoxvirus specific antibodies in milk samples using the commercially available ELISA from IDvet is feasible and can represent a helpful tool for LSDV monitoring programs.

Dataset Information

A New ELISA Using the ANANAS Technology Showing High Sensitivity to diagnose the Bovine Rhinotracheitis from Individual Sera to Pooled Milk.

Publications

A New ELISA Using the ANANAS Technology Showing High Sensitivity to diagnose the Bovine Rhinotracheitis from Individual Sera to Pooled Milk.

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