Protocol of living cell separation using the microfluidic dielectrophoresis integrated chip.

This protocol demonstrates the separation of living cells with the microfluidic dielectrophoresis chip, using the Jurkat cell as a model. The successful living cell separation lies in familiarity with the detailed tips, which are aided by this stepwise protocol. The knowledge of correct chip installation, sample and buffer filling, flow rate and cell concentration adjustments, and contamination sources increases the efficiency of target viable cell collection. Such instructions, although trivial, are critical for achieving cell separation. For complete details on the use and execution of this protocol, please refer to Oshiro et al. (2022).

Fabrication of a new all-in-one microfluidic dielectrophoresis integrated chip and living cell separation.

Microfluidic dielectrophoresis (DEP) technology has been applied to many devices to perform label-free target cell separation. Cells separated by these devices are used in laboratories, mainly for medical research. The present study designed a microfluidic DEP device to fabricate a rapid and semiautomated cell separation system in conjunction with microscopy to enumerate the separated cells. With this device, we efficiently segregated bacterial cells from liquid products and enriched one cell type from two mixed eukaryotic cell types. The device eliminated sample pretreatment and established cell separation by all-in-one operation in a lab-on-chip, requiring only a small sample volume (0.5-1 mL) to enumerate the target cells and completing the entire separation process within 30 min. Such a rapid cell separation technique is in high demand by many researchers to promptly characterize the target cells.

AFI Corp. PixeeMo™ for Enumeration of Aerobic Bacteria in Drinking Water: AOAC Performance Tested MethodSM 012002.

BACKGROUND: PixeeMo™ is a compact instrument that enables bacterial cell counting using microfluidic chips instead of counting of colonies on culture media. Chips containing electrodes, based on fluid, electric filtering and sorting technology (FES), allow the selection of bacterial cells from other components in the sample. In the United States (US), surface water or ground water affected by surface water must be treated to reduce the total microbial load to less than 500 CFU/mL. In Japan, drinking water regulations limit the total bacterial load to 100 CFU/mL. OBJECTIVE: To validate the PixeeMoTM aerobic bacteria method based on the Japanese regulation in the range of 30-300 CFU/mL in drinking water. METHOD: PixeeMoTM aerobic bacteria method was compared to the Standard Method for the Examination of Water and Wastewater (SMEWW) 9215B (2017) using naturally contaminated drinking water. RESULTS: The maximum repeatability standard deviation of the PixeeMoTM method was 14.8%. The difference of mean log10 values between the PixeeMoTM and SMEWW 9215B methods ranged from -0.015 to 0.258. Similar results were obtained in the independent laboratory study. CONCLUSIONS: The PixeeMoTM method is equivalent to that of the SMEWW 9215B methods. The product consistency and stability study demonstrated no significant difference within the expiration date. The robustness study confirmed that there was no effect within the expected range. The instrument variation study also demonstrated no significant difference among the data of three PixeeMoTM instruments. HIGHLIGHTS: Total counts of bacteria in drinking water can be determined accurately within 1 h with PixeeMoTM.

Separation of viable and nonviable animal cell using dielectrophoretic filter.

Selective separation of cells using dielectrophoresis (DEP) has recently been studied and methods have been proposed. However, these methods are not applicable to large-scale separation because they cannot be performed efficiently. In DEP separation, the DEP force is effective only when it is applied close to the electrodes. Utilizing a DEP filter is a solution for large-scale separation. In this article, the separation efficiency for viable and nonviable cells in a DEP filter was examined. The effects of an applied AC electric field frequency and the gradient of the squared electric field intensity on a DEP velocity for the viable and nonviable animal cells (3-2H3 cell) were discussed. The frequency response of the DEP velocity differed between the viable and the nonviable cells. We deducted an empirical equation that can be used as guiding principle for the DEP separation. The results indicate that the viable and the nonviable cells were separated using the DEP filter, and the best operating conditions such as the applied voltage and the flow rate were discussed.

Separation characteristics of animal cells using a dielectrophoretic filter.

The separation characteristics of a wire-wire type dielectrophoretic (DEP) filter were evaluated using animal cells. The separation of cells with different activities was examined using a DEP filter. The specific growth rate of the cells in retention liquid was larger than that in permeation liquid. From the culture results of the separated cells, it becomes clear that the specific growth rate of the cells of the retention liquid was higher than that of the cells of the permeation liquid. Furthermore, as a result of separating cells two kinds of cell suspensions using the DEP filter, the difference between the retention ratios of the two groups of obtained cells was about 20% at maximum.