The purpose of this study was to evaluate a commercial enzyme-linked immunosorbent assay (ELISA) for human von Willebrand factor antigen (vWF:Ag) with respect to its potential value in quantitating the protein in canine plasma. The assay was a sandwich technique using F(ab')2 fragments specific for von Willebrand factor (vWF) and a peroxidase conjugated rabbit anti-vWF second antibody, with a microplate as the support surface. Canine plasmas were assayed by ELISA, and by Laurell electroimmunoassay (EIA), our reference methodology. The ELISA had a within-day variation of 1.21-4.44% and a between-day variation of 0.85-4.88% depending on the level of vWF:AG. The sensitivity of the assay was less than 0.1% vWF:AG. The range of vWF:AG concentrations in plasmas from 24 clinically normal dogs compared favorably with the range for the same plasmas when assayed by EIA (ELISA = 60-152% of normal; EIA = 50-142% of normal). In 121 canine plasmas with vWF:AG concentrations (as assessed by EIA) ranging from undetectable levels (< 6% of normal) to 142% of normal, there was good correlation with measurements made by ELISA (correlation coefficient = 0.835). It was concluded that this commercial ELISA technique could be used to provide reliable, same-day measurements of canine plasma vWF:AG. Since it requires no special equipment other than a microplate reader and washer it is particularly suitable for laboratories lacking the electrophoretic expertise or equipment required for EIA.

A reliable bovine viral diarrhea (BVD) viral antigen was prepared from BVD virus grown on Madin Darby bovine kidney (MDBK) cells by solubilizing the virus with detergent MEGA-10 (decanoyl-N-methylglucamide) followed by removal of hydrophobic proteins with Triton X-100 treatment. By these treatments, problems of high background associated with BVD viral antigen in the enzyme-linked immunosorbent assay (ELISA) were eliminated. With this new antigen, an ELISA was adapted to detect bovine serum antibody against BVD virus. The diagnostic specificity of the assay in 403 bovine sera collected from a BVD virus-free herd was 100%; in 296 bovine sera with serum neutralizing antibody titers of greater than or equal to 1:2, 289 sera were ELISA positive (relative sensitivity of 97.6%), two sera gave false negative reactions (0.7%) and five sera gave suspicious reactions (1.7%). These interpretations were based on positive/negative (P/N) ratio readings, i.e. a P/N ratio of < 1.50, 1.50-1.99 and greater than or equal to 2.00 were interpreted as negative, suspicious and positive reactions, respectively. The ELISA results gave excellent agreement with serum neutralization in detecting both seropositive and seronegative animals (Kappa = 0.994). The ELISA assay was considered to be technically superior to the serum neutralization test for the routine detection of BVD viral antibody in bovine sera.

Three ejaculates from each of 3 stallions were used to evaluate a computerized system (Hamilton-Thorn motility analyzer; HTMA) for measuring equine spermatozoal motility. Variance components (ejaculate-within-stallion, chamber-within-ejaculate, and microscopic field-within-chamber) were determined for each stallion after diluting ejaculates to 25 X 10(6) spermatozoa/ml with a skim milk-glucose seminal extender. The HTMA was compared with frame-by-frame playback videomicrography (VIDEO) for determining: percentage of spermatozoal motility and spermatozoal number in microscopic fields; curvilinear velocity and straight-line velocity of individual spermatozoa for 5 track types; and repeatability of those velocity measurements. The effect of spermatozoal number per microscopic field on incidence of intersecting spermatozoa and the outcome of intersecting spermatozoa also were evaluated. Greatest variability in motility measures was generally attributed to the microscopic field-within-chamber component. The HTMA was highly correlated with VIDEO for estimation of spermatozoal numbers per microscopic field (r = 0.99; P <0.001) and motility (r = 0.97; P <0.001); however over the entire range of spermatozoal numbers, the HTMA yielded higher spermatozoal numbers per microscopic field (P <0.05) and higher motility (P <0.05) than did VIDEO. The HTMA- and VIDEO-derived measurements of curvilinear and straight-line velocities were highly correlated for all spermatozoal track types, but both measures were higher (P <0.05) by use of the HTMA than by use of VIDEO for most track types. For 3 of 5 track types, measurements of curvilinear and straight-line velocities were less variable (P <0.05), using the HTMA, rather than VIDEO. Using the HTMA, the number of intersecting spermatozoa was highly correlated with spermatozoal numbers per microscopic field (r = 0.97; P <0.001). The percentage of erroneous track interpretations involving intersecting spermatozoa was high (85.3 +/- 2.7%). The HTMA was a reliable system for determining percentage of spermatozoal motility and velocity measures in video recordings of equine semen diluted to spermatozoal concentration of 25 X 10(6)/ml prior to evaluation.