Frequency of enumeration errors of total nucleated cell counts on canine bal fluid using the Sysmex XT-2000iV

Although some concerns exist on the practical utility of total nucleated cell counts (TNCCs) on bronchoalveolar lavage (BAL) fluids, the TNCCs is considered a useful tool to estimate the quality of the sample, based on the assumption that only when samples have a sufficient number of cells cytology may provide reliable information. No information about the possible presence of artifacts in automated counts on BAL fluids from dogs is available. The aim of this study was to assess the frequency of enumeration errors on BAL fluids using the laser counter Sysmex XT2000iV. A total of 60 BAL fluids collected from dogs with respiratory disorders using standard endoscopic techniques were analyzed. Counts generated in DIFF and BASO channels and DWBC were recorded. The DIFF channel counts the cells stained with a fluorescent dye that specifically binds to nucleic acids; the BASO channel counts the cellular residues produced after contact with an acidic reagent that collapses the nucleated cells. DWBC is the ratio between DIFF and BASO counts, which should be close to 1.00 without artifacts. In our caseload, only 4 samples (6.7%) had a DWBC of 1.00 _ 0.05, probably not affected by artifacts. Conversely, 56 samples (93.3%) had an abnormal DWBC. Specifically, 42 samples (70%) had a DWBC < 1.00 (mean _ SD=0.57 _ 0.24; median=0.57; min-max=0.12-0.94), likely due to the presence of non-cellular particles (eg, fragments of mucus clots, cellular debris), resulting in higher BASO compared with DIFF counts. On the contrary, 13 samples (21.7%) had a DWBC higher than 1.00 (2,15 _ 1,74; 1,32; 1,06-6,17), due to mucous clots in BASO reagent entrapping the cells, resulting in falsely low BASO counts, or DNA positive non-cellular particles (eg bacterial aggregates), falsely elevating the DIFF count. In conclusion, this study demonstrated that the automated TNCCs on canine BAL fluids may suffer from pre-analytical errors resulting in inaccurate results. Cytologic examination of BAL fluids is required to verify automated counts and potential artifacts.