A Novel Staining Method For Detecting Bloodborne Bacteria After Their Dielectrophoretic Isolation

Current methods to diagnose bacteremia typically require 18 to 36 hours to detect positive samples and five days to identify negative ones. This delay stems in part from the incubation time necessary for bacteria to multiply and achieve detectable concentrations. In recent years, microfluidic chips have been developed that isolate and concentrate bacteria from the blood on a time scale of minutes to hours (1). This research seeks to stain and visualize these bacteria after their microfluidic processing thereby significantly shortening the time required to diagnose bacteremia. The traditional Gram stain is not sufficiently sensitive to detect bacteria in this setting (2). A novel staining procedure involving sample filtration was therefore developed and tested against the Gram stain with a focus on the relative performance of these methods in samples with low concentrations of bacteria. The experimental staining procedure was first optimized by testing different combinations of filters and counterstains. The resulting finalized procedure, when tested against the Gram stain, detected bacteria at concentrations roughly two orders of magnitude lower than those of the control method. The results indicate that this novel staining method may have utility when used in conjunction with a microfluidic condenser in certain applications. As for a broadly applicable method of diagnosing bacteremia more quickly, these results are promising but further improvements are required to increase the sensitivity of the test and decrease the time required to perform it