Neutrophil CD64 by Flow Cytometry Has Excellent Negative Predictive Value for Excluding Sepsis

Abstract

Sepsis remains a major health concern in acute care hospitals, and missed diagnoses can have significant consequences. Fast and accurate identification and treatment is paramount. However, up to half of antibiotic usage in U.S. acute care hospitals is unnecessary, in part because final negative microbiological culture results can take 2 to 5 days. Unnecessary antibiotic use has been implicated in microbial resistance. Neutrophil expression of CD64 (nCD64) is increased when neutrophils are exposed to infectious stimuli, and can be rapidly detected by flow cytometry. Neutrophil CD64 expression has shown promise in recognizing early sepsis. We enrolled 100 consecutive patients seen in the emergency department with a clinical suspicion of sepsis, where both complete blood count (CBC) and blood cultures were ordered. Fifty uL of whole blood in EDTA from the CBC tube was lysed and washed to produce a red-cell-free cell suspension. This suspension was incubated with CD64, CD45, and monocyte differentiation antibodies, and then analyzed under a granulocyte template on the BD FACS Canto II flow cytometer. Patient ages varied from newborn to 98 years. Ninety-one cultures showed no growth at 5 days, five cultures were positive but judged to be contaminants, four cultures met criteria for laboratory-confirmed bloodstream infection. Receiver operator characteristic (ROC) curves for nCD64 mean fluorescence (MF) and confirmed bloodstream infection were plotted and an optimum nCD64 MF cut point was determined by Youden J statistic (3488 MF: 75% sensitivity, 82% specificity, 99% negative predictive value). A cut point at 560 MF showed 100% sensitivity (28% specificity, 100% negative predictive value). The lowest calculated negative predictive value at any cut point was 97%. Neutrophil CD64 expression as detected by this method accurately predicts negative bloodstream infection. This may improve antibiotic utilization and stewardship, and decrease healthcare costs.