Automated counting of nucleated cells in equine synovial fluid without and with hyaluronidase pretreatment.

Abstract

BACKGROUND: Microscopy is usually used to obtain manual total and differential cell counts in equine synovial fluid. A faster, more precise method is desirable. OBJECTIVES: The objectives were to compare an automated impedance method with a manual method for obtaining total and differential cell counts in equine synovial fluid and to evaluate the effect of pretreatment with hyaluronidase on automated results. METHODS: Synovial fluid samples (n=48) were collected into EDTA and analyzed within 48 hours. Automated total and differential cell counts were evaluated using a Medonic CA620-VET hematology analyzer before and after pretreatment for 5-30 minutes with hyaluronidase (final concentration 0.01 mg/mL). A hemacytometer count and microscopic evaluation of a direct smear were used as the reference method. Intra-assay coefficients of variation (CV) were determined. RESULTS: Thirty-one of 46 untreated samples and 0/46 hyaluronidase-treated samples were error-flagged by the analyzer. Correlation between automated (ANCC) and manual (MNCC) nucleated cell counts in untreated samples (n=15; R(2)=0.93) and pretreated samples (n=46; R(2)=0.94) was high, and pseudomedian difference was low. Intra-assay CVs for samples with medium and high cellularity were significantly lower for ANCC (1.5-2.7%) compared with MNCC (6.1-15.7%) (P<.01). Valid automated differential cell counts were not obtained. CONCLUSIONS: Automated total cell counts obtained on the Medonic analyzer correlate well with manual counts in equine synovial fluid; however, pretreatment with hyaluronidase is required to minimize error flags. Automated differential counts are not accurate for synovial fluid.