Our group has developed a microfluidic cell-trapping device and characterization protocol that is able to overcome conventional limitations on microenvironment control, time sensitivity and single cell analysis in vitro 10, 11, 12, 13, 14, 15. Devices with multitudinous single cell traps, however, offer a powerful alternative to traditional cell culture and analysis 9. Microfluidic cell trapping devices are often cited as a solution to this problem, but those designed with only one cell trap are constrained by low throughput 7, 8.
Flow cytometry can measure the fluorescence of one cell at one moment in time and multi-well plate fluorometry can measure the fluorescence of a large population of cells over time, but neither can adequately perform both tasks simultaneously.
Multi-well culture plates containing large populations of cells can be observed over time but, for non-adherent cells, altering the composition of the extracellular media requires cumbersome centrifugation and resuspension that may induce unintended changes in gene expression 6. Flow cytometry can measure single cell fluorescence, internal complexity and volume 5, but it cannot measure time-dependent, transient cell responses to stimuli. Therefore, a reliable method for time-dependent analysis of patients' single cancer cells in vitro may enhance early cancer detection, refine neoplastic cell characterization and enable chemotherapeutic treatment customization 4. From initial transformation to drug resistance, the progression of malignancy depends upon the survival and proliferation of individual cells with unique genotypes expressing environmentally dependent phenotypes 1, 2, 3. Hematologic cancer is a disease of single cells. The transporter data indicate that Jurkat T cells exposed to indomethacin continue to accumulate fluorescent calcein for over 60 minutes after calcein-AM is removed from the extracellular space. We show how the MTNP platform can be used for hematologic cancer cell characterization by measuring single T cell levels of CRAC channel modulation, non-translational motility and ABC-transporter inhibition via a calcein-AM efflux assay. Here we demonstrate the multitrap nanophysiometer (MTNP), a low-volume microfluidic platform housing an array of cell traps, as an effective tool that can be used to study individual unattached cells over time with precise control over the intercellular microenvironment.
Unfortunately, many cellular processes, including signaling, motility and molecular transport, occur transiently over relatively short periods of time and at different magnitudes between cells. Cytometric studies utilizing flow cytometry or multi-well culture plate fluorometry are often limited by a deficit in temporal resolution and a lack of single cell consideration.