Microfluidics delivery of DARPP-32 into HeLa cells maintains viability for in-cell NMR spectroscopy

Abstract

High-resolution structural studies of proteins and protein complexes in a native eukaryotic environment present a challenge to structural biology. In-cell NMR can characterize atomic resolution structures but requires high concentrations of labeled proteins in intact cells. Most exogenous delivery techniques are limited to specific cell types or are too destructive to preserve cellular physiology. The feasibility of microfluidics transfection or volume exchange for convective transfer, VECT, as a means to deliver labeled target proteins to HeLa cells for in-cell NMR experiments is demonstrated. VECT delivery does not require optimization or impede cell viability; cells are immediately available for long-term eukaryotic in-cell NMR experiments. In-cell NMR-based drug screening using VECT was demonstrated by collecting spectra of the sensor molecule DARPP32, in response to exogenous administration of Forskolin.