Intra-microfluidic pinocytic loading of human T cells

Pinocytic loading (PL) exposes cells sequentially to hypertonic (0.5 M sucrose and 10% PEG 1000) and hypotonic (60% v/v media in water) solutions to transfer cell-impermeant macromolecules into the cytosol. Here we leverage the advantages of microfluidics to achieve high-efficiency pinocytic loading of Jurkat T cells with fluorescein dextran. To our knowledge, we report the first observation of the time course of PL in single cells. In contrast to conventional PL, cells in the microfluidic device can be rapidly cycled through the various solutions. While the conventional protocol provides only 40-60% efficiency, we have achieved over 90% loading efficiency in the microfluidic device. Mean cell fluorescence from conventional PL, is approximately half that of microfluidic PL with three 10-minute hypertonic cycles (49.7plusmn21.8 and 21.7plusmn17.8 a.u. respectively, P=4ldr10^-17). While the mean and median cell fluorescence increases with each cycle in the microfluidic device, fluorescence begins to plateau after three 10-minute hypertonic cycles; subsequent cycles see dramatic increases in cell death from 10% to over 30%. Hypotonic media does not significantly affect loading efficiency for intra-inicrofluidic PL, but causes slightly higher cell death. We believe that intra-microfluidic PL of living cells is a valuable tool for BioMEMS-based systems biology.