Intratumoral heterogeneity is a substantial cause of drug resistance development during chemotherapy or other drug treatments for cancer. Therefore, monitoring and understanding cell exposure and response to drugs at a single-cell level are crucial. Previous research suggested that single-cell growth rate can be used to investigate drug-cell interactions. However, currently established methods for quantifying single-cell growth are limited to isolated or monolayer cells. Here, we introduce a technique that accurately measures both 2D and 3D cell growth rates using label-free ratiometric stimulated Raman scattering (SRS) microscopy. We use deuterated amino acids, leucine, isoleucine, and valine, as a tracer and measure the C-D SRS signal from the deuterium-labeled proteins and the C-H SRS signal from unlabeled proteins simultaneously to determine the cell protein synthesis rate at a single-cell level. The technique provides us a way to perform single-cell level drug testing with a shorter turnaround time (&lt;12 hours) than most traditional assays. The submicron resolution of the imaging technique allows us to examine the effects of chemotherapeutic drugs, including kinase inhibitors, mitotic inhibitors, and topoisomerase II inhibitors, on both cell growth rate and morphology. The capability of quantifying 3D cell growth rates provides insight into a deeper understanding of the drug-cell interaction in the actual tumor environment.

Quantification of cell growth rates by ratiometric stimulated Raman scattering microscopy

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