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Comparison of sample types and location of skin biopsies for BVDV detection using antigen capture ELISA Testing. Dr. Brian Vander Ley BVD Symposium January 26, 2009 Phoenix, Arizona. Introduction. We are looking for ways to screen for PI animals more effectively Sample choices

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Comparison of sample types and location of skin biopsies for BVDV detection using antigen capture ELISA Testing

Dr. Brian Vander Ley

BVD Symposium January 26, 2009

Phoenix, Arizona


Introduction
Introduction BVDV detection using antigen capture ELISA Testing

  • We are looking for ways to screen for PI animals more effectively

    • Sample choices

    • Assay evaluation

    • Assay development


Overview
Overview BVDV detection using antigen capture ELISA Testing

  • 2 Projects

    • Sample project

      • 40 PIs

      • Several different samples evaluated

    • Hide project

      • 3 PIs

      • Involved systematic evaluation of whole hides for BVDV antigen


Sample project
Sample Project BVDV detection using antigen capture ELISA Testing

  • 40 PI calves in 2 stocker operations in Oklahoma

    • Identified by ACE

    • Segregated from non-PI herdmates


Skin samples
Skin Samples BVDV detection using antigen capture ELISA Testing

  • 3 ear notches

    • IHC

    • ACE

    • One was archived for future use

  • 1 tail fold biopsy

Above: Ear notcher with ear notch. Below: tail fold

Photo from: www.drugs.com/vet/images/1136000a.png


Swab samples
Swab Samples BVDV detection using antigen capture ELISA Testing

  • Nasal

  • Oral

  • Conjuctival

  • Vaginal/Preputial

  • Rectal

Collecting an oral swab

Photo from: www.puritanmedproducts.com


Sample processing
Sample Processing BVDV detection using antigen capture ELISA Testing

  • Skin Samples

    • Placed in dry tubes and frozen until testing

    • Incubated in 1 mL PBS for 24 hours at 4°C prior to using ACE

  • Swab Samples

    • Collected and place immediately in 1 mL PBS then frozen

    • Thawed and allowed to incubate for 24 hours 4°C prior to ACE


Results
Results BVDV detection using antigen capture ELISA Testing

  • Accuracy reported as a percentage: # positive of sample type divided by # positive IHC.

  • Accuracies

    • Ear notch-98%

    • Tail biopsy-98%

    • Nasal swab-98%

    • Vaginal/Preputial swab-90%

    • Conjunctival swab-63%

    • Rectal swab-10%

    • Oral swab-8%


Sample conclusions
Sample Conclusions BVDV detection using antigen capture ELISA Testing

  • Nasal swabs could be a good sample for detecting PI cattle.

    • Simple

    • Non-invasive

  • Other samples could also be good, but would pose some challenges

  • More work would need to be done to validate any of these samples


Hide project
Hide Project BVDV detection using antigen capture ELISA Testing

  • 3 PIs selected based on previous diagnosis as PI by ACE

  • Each animal was euthanized and the skin was removed in its entirety

A complete hide


Hide project1
Hide Project BVDV detection using antigen capture ELISA Testing

  • Each hide was quartered in the field and packed on ice

  • The hides were frozen until processing

Quarter division of hides


Hide processing
Hide Processing BVDV detection using antigen capture ELISA Testing

  • Each quarter was thawed and divided into 10 cm by 10 cm sections

  • A 4 mm punch biopsy was taken from each section and placed in a dry microcentrifuge tube.

Hide section schematic


Sample evaluation
Sample Evaluation BVDV detection using antigen capture ELISA Testing

  • All samples were incubated in 1 mL PBS at 4°C for the same amount of time.

  • All steps were performed the same way on each plate and in sequential order from plate 1 to plate 7.

  • Multiple controls were used in each plate

    • Kit positive control

    • Kit negative control

    • Titered BVDV infected cell culture

    • Titered BVDV infected cell culture diluted 1:10 in PBS


Results1
Results BVDV detection using antigen capture ELISA Testing

  • All sections were positive by ACE

  • No significant differences in S/P ratios were found

    • Between one particular quarter in all three hides

    • Between quarters on an individual

    • Between animals


Conclusions
Conclusions BVDV detection using antigen capture ELISA Testing

  • Large amounts of sample material are available for evaluation/development of diagnostic assays aimed at detecting PI cattle.

  • The whole hide can be utilized as a source of samples for running the ACE.


Overall conclusions
Overall Conclusions BVDV detection using antigen capture ELISA Testing

  • BVDV Antigen can be recovered from a variety of places.

    • Many swabs

    • Essentially any haired skin sample

  • There are large amounts of material that can be harvested from an individual animal for evaluation or development of diagnostic assays, especially those that use skin.

  • More research needs to be done to validate any of the samples.


References
References BVDV detection using antigen capture ELISA Testing

  • Idexx Herdchek® Bovine Virus Diarrhea Antigen Test Kit. U.S. Vet. License No. 313. 2007.


Questions? BVDV detection using antigen capture ELISA Testing

Thank You


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