Analytical Performance of an Ultrasonic Particle Focusing Flow Cytometer

Abstract

Creation of inexpensive small-flow cytometers is important for applications ranging from disease diagnosis in resource-poor areas to use in distributed sensor networks. In conventional-flow cytometers, hydrodynamics focus particles to the center of a flow stream for analysis, which requires sheath fluid that increases consumable use and waste while dramatically reducing instrument portability. Here we have evaluated, using quantitative measurements of fluorescent microspheres and cells, the performance of a flow cytometer that uses acoustic energy to focus particles to the center of a flow stream. This evaluation demonstrated measurement precision for fluorescence and side scatter CVs for alignment microspheres of 2.54% and 7.7%, respectively. Particles bearing 7 x 10(3) fluorophores were well resolved in a background of 50 nM free fluorophore. The lower limit of detection was determined to be about 650 fluorescein molecules. Analysis of Chinese hamster cells on the system demonstrated that acoustic focusing had no effect on cellular viability. These results indicate that the ultrasonic flow cytometer has the necessary performance for most flow cytometry applications. Furthermore, through robust engineering approaches and the combination of acoustic focusing with low-cost light sources, detectors, and data acquisition systems, it will be possible to achieve a low-cost, truly portable flow cytometer.

A microflow cytometer exploited for the immunological differentiation of leukocytes

“…Alternatively, acoustic focusing (15) can be used to analyze particles and cells (16). Dron et al demonstrated acoustic focusing of particles in a planar microchannel using a standing acoustic wave (41).…”

“…When analyzing micro particles, focusing of the sample stream is usually applied to restrict their trajectories inside the flow channel (1,3,5,6,(8)(9)(10)(11)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26). This positioning serves to increase signal stability when measuring particle properties and to avoid surface fouling in microfluidic channels.…”

Flow Cytometry: A Multipurpose Technology for a Wide Spectrum of Global Biosecurity Applications

“…60 Combining these approaches enables development of a simpler device, with fewer options for the operator, and potentially more sustainable operation. A lost-cost, portable flow cytometer is being developed currently at Los Alamos National Laboratory using innovative approaches such as acoustic energy, rather than large volumes of sheath fluid, to focus particles to the center of a flow stream 21 ; low-cost light sources such as laser pointer modules 24 ; and open, reconfigurable digital data acquisition systems. 46 The availability of a ''minimalist'' flow cytometer 29 would immediately improve the quality of medical care that is available to diagnose and treat infectious diseases in poor regions of the world, which could in turn have a positive impact on international security.…”

A novel microfluidic flow focusing method

“…For instance, Shi et al 17,18 used a standing surface acoustic waves (SSAWs) method to realize on-chip micro-particle focusing. Goddard et al 19 evaluated the performance of a flow cytometer that used acoustic energy to focus particles/cells to the center of a flow stream. Suthanthiraraj et al 20 presented the one-dimensional acoustic standing waves for focusing of flowing cells and particles in a rectangular flow channel.…”