ISU VDL researchers have developed methods to adapt the IDEXX HerdCheck X3® Elisa for PRRS to the oral fluid matrix. The PRRS X3 ORAL FLUID ELISA is interpreted exactly as the PRRS serum ELISA: S:P ratios ≥ 0.4 are considered positive. The test was estimated to be 94.7% sensitive and close to 100% specific at the ≥ 0.4 cutoff (Kittawornrat, Zimmerman et al., 2012). As shown in the example below, the PRRS X3 ORAL FLUID ELISA detects maternal antibody, as well as antibody produced in response to PRRSV infection.
Dr. David Baum
ISU VDL Serology Section Head
- Oral fluid samples can be readily collected from adult and growing pigs
- Submit oral fluid samples in 5 ml Falcon™ snap-cap tubes
- Keep samples cold or frozen (ship in brucellosis boxes on ice-packs)
- PRRS X3 Oral Fluid Elisa Assay is a 2-day test, as methods require an overnight incubation
References from AAVLD Proceedings September 2011:
Kittawornrat A, Prickett J, Wang C, Olsen C, Irwin C, Panyasing Y, Ballagi A, Rice A, Main R, Rademacher C, Hoogland M, Zimmerman J. 2011. Adaptation of a commercial PRRS serum antibody ELISA to oral fluid specimens. AAVLD Proceedings Sept 2011, p99. [View pdf]
Kittawornrat A, Prickett J, Wang C, Olsen C, Irwin C, Panyasing Y, Ballagi A, Rice A, Rowland R, Zimmerman J. 2011. Diagnostic performance of a commercial PRRS serum antibody ELISA adapted to oral fluid specimens: longitudinal response in experimentally-inoculated populations. AAVLD Proceedings Sept 2011, p100. [View pdf]
Kittawornrat A, Prickett J, Wang C, Olsen C, Irwin C, Panyasing Y, Ballagi A, Rice A, Main R, Rademacher C, Hoogland M, Zimmerman J. 2011. Diagnostic performance of a commercial PRRS serum antibody ELISA adapted to oral fluid specimens: field samples. AAVLD Proceedings Sept 2011, p201. [View pdf]
ISU VDL will begin using HerdChek* PRRS X3 ELISA as the routine PRRS ELISA test on Monday April 26, 2010.
Our collaborative efforts with IDEXX have demonstrated that the HerdChek X3™ PRRS Elisa offers a significant improvement in specificity (i.e., reducing false positives by 75 – 90%) while retaining excellent sensitivity. This new indirect enzyme-linked immunosorbent assay (ELISA) detects PRRS antibodies to either the North American or European strains of the virus in either plasma or serum with 99.9% specificity and 98.8% sensitivity. Probably the most exciting feature of the test is its reduction of false positive singleton reactors by 90%. The test while not approved for use in the United States at this time is approved and is in wide use across Europe.
The ISU VDL, the University of Minnesota VDL and the diagnostic facility of Boehringer Ingelheim Vetmedica were involved in a field trial of the IDEXX X3 kit last September. All three labs tested a sample set that was developed by IDEXX consisting of known status samples. All three labs were also asked to generate an internal panel of known (positive and negative) status samples from field samples that had been previously tested. ISU VDL was also given the latitude to assemble a panel of suspected false positive samples. We selected 26 singleton reactors as tested on the 2XR kit from known PRRS negative flows. We found 23 of 26 of these singleton reactors tested negative on the new PRRS X3 kits or an 88.46% reduction in false positive results. The field trial data generated at the 3 independent diagnostic facilities mirrored the data generated at IDEXX’s own X3 validation study. This information is now in the hands of the Federal Regulatory Agency and IDEXX is awaiting approval to market the kit for use in the United States. The ISU VDL has obtained a permit allowing us to import the new X3 kit to the United States from Europe. Presently we are using the PRRS X3 kits on selected boar stud submissions and on selected genetic multipliers that are PRRS negative. The ISU VDL also uses the test as an alternative test to be offered on suspected false positive PRRS ELISA results generated on the present 2XR kits. The VDL has taken a look at the data from one PRRS-negative genetic source to compare 2XR results versus X3 results on a like number of samples tested.
|ISU-VDL Evaluation of Field Samples from PRRSV Negative Herds|
|PRRS2XR Elisa||HerdChek* X3|
|# of false positives||13||3|
|Total # of samples||740||740|
|% of false positives||1.8%||.04%|
|ISU VDL field samples suggest ~76% reduction in false positives|
In a January to February test period the VDL tested 740 samples for this negative PRRS flow on the 2XR kit finding 13 of the 740 samples submitted to be ELISA positive. In a similar time period from March to present using the new PRRS X3 kit the VDL has tested an additional 740 samples from these same sites. We found using the PRRS X3 ELISA that 3 of 740 samples tested suspect (.2 - .399) or positive (>.4). For this particular site, this translates into a 1.76% false positive rate using the old 2XR kit versus a .405% false positive rate using the new X3 kit.
We fully appreciate the amount of anxiety, additional costs incurred, the disruption of pig movements, labor of retesting, and the valuable time loss that occurs when a false positive PRRS ELISA result is released from the lab. These negative economic impacts, coupled with additional time factors and the client anxiety level associated with the false positive results generated presently with the 2XR PRRS kit coupled with the confidence we have developed in the X3 assay are the reasons we are making the switch now to PRRS X3 as our routine PRRS ELISA.
IDEXX Validation Data Summary
Studies were performed to evaluate the diagnostic performance of the new HerdChek® X3 ELISA for PRRS. Multiple populations were used to evaluate the sensitivity and specificity. The X3 assay resulted in 98.8% diagnostic sensitivity when tested on >1000 characterized PRRS positive sera samples. Seroconversion was detected as early as eight days post infection (dpi) and positive status detection was maintained as late as 175 dpi in experimentally infected populations. The diagnostic specificity of PRRS X3 was determined to be 99.9% using populations of 1445 negative sera. Also a direct comparison was made to the PRRS 2XR ELISA using the singleton rector population of PRRS 2XR ELISA false positive sera. Results from the X3 ELISA indicate a 90% reduction in the singleton reactors. Therefore, the new PRRS X3 kit for Ab detection in swine sera exhibits improved diagnostic specificity with reduced risk of false positives, as well as equivalent or improved detection of seroconversion as compared to PRRS 2XR.
Note: The PRRS 2XR Elisa will continue to be available for use upon special request.