A microfluidic device enabling drug resistance analysis of leukemia cells via coupled dielectrophoretic detection and impedimetric counting

Abstract

We report the development of a lab-on-a-chip system, that facilitates coupled dielectrophoretic detection (DEP-D) and impedimetric counting (IM-C), for investigating drug resistance in K562 and CCRF-CEM leukemia cells without (immuno) labeling. Two IM-C units were placed upstream and downstream of the DEP-D unit for enumeration, respectively, before and after the cells were treated in DEP-D unit, where the difference in cell count gave the total number of trapped cells based on their DEP characteristics. Conductivity of the running buffer was matched the conductivity of cytoplasm of wild type K562 and CCRF-CEM cells. Results showed that DEP responses of drug resistant and wild type K562 cells were statistically discriminative (at p = 0.05 level) at 200 mS/m buffer conductivity and at 8.6 MHz working frequency of DEP-D unit. For CCRF-CEM cells, conductivity and frequency values were 160 mS/m and 6.2 MHz, respectively. Our approach enabled discrimination of resistant cells in a group by setting up a threshold provided by the conductivity of running buffer. Subsequent selection of drug resistant cells can be applied to investigate variations in gene expressions and occurrence of mutations related to drug resistance.

Figure 1

Overall schematic (a) and working principle (b) of the LOC system, comprising DEP-D and IM-C units.

Figure 2

Fabricated LOC system emphasizing electrodes in DEP-D and IM-C units and flow distributor.

Figure 3

Single shell cell modeling to obtain Re(f_CM) characteristics of cells having cytoplasmic conductivities of 210, 215, referring K562/wt cells, and 374 mS/m, referring K562/imaR cells, in LCB (conductivity, σ = 2.5 mS/m) (a) and relatively HCB (σ = 200 mS/m) (b). (c) Selection of the working frequency (8.6 MHz) for K562/wt and imaR cells in relatively HCB (σ = 200 mS/m). (d) Selection of the working frequency (6.2 MHz) for CCRF-CEM/wt and doxR cells in buffers (at different conductivity values: 110, 125, and 160 mS/m).

Figure 4

Dielectrophoretic behavior of leukemia cells and their drug resistant progenies. (a) The trapping ratio of K562/wt and K562/imaR cells immersed in relatively HCB (σ = 200mS/m) at 8.6 MHz under 20V_pp and at a flow rate of 1 µL/min. (b) The trapping ratio of CCRF-CEM/wt and CCRF-CEM/doxR cells in different buffers, with 110, 125, and 160 mS/m conductivity, at 6.2 MHz under 20V_pp and at a flow rate of 1 µL/min. Results are presented as mean ± stdev. * indicates significantly different results at the p = 0.05 level (n = 3).