Bessel-Beam Hyperspectral CARS Microscopy with Sparse Sampling: Enabling High-Content High-Throughput Label-Free Quantitative Chemical Imaging

Anal Chem. 2018 Mar 20;90(6):3775-3785. doi: 10.1021/acs.analchem.7b04039. Epub 2018 Mar 9.

Abstract

Microscopy-based high-content and high-throughput analysis of cellular systems plays a central role in drug discovery. However, for contrast and specificity, the majority of assays require a fluorescent readout which always comes with the risk of alteration of the true biological conditions. In this work, we demonstrate a label-free imaging platform which combines chemically specific hyperspectral coherent anti-Stokes Raman scattering microscopy with sparse sampling and Bessel beam illumination. This enabled us to screen multiwell plates at high speed, while retaining the high-content chemical analysis of hyperspectral imaging. To demonstrate the practical applicability of the method we addressed a critical side effect in drug screens, namely, drug-induced lipid storage within hepatic tissue. We screened 15 combinations of drugs and neutral lipids added to human HepG2 liver cells and developed a high-content quantitative data analysis pipeline which extracted the spectra and spatial distributions of lipid and protein components. We then used their combination to train a support vector machine discriminative algorithm. Classification of the drug responses in terms of phospholipidosis versus steatosis was achieved in a completely label-free assay.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Drug Evaluation, Preclinical / instrumentation*
  • Drug Evaluation, Preclinical / methods
  • Equipment Design
  • Hep G2 Cells
  • High-Throughput Screening Assays / instrumentation*
  • High-Throughput Screening Assays / methods
  • Humans
  • Lipids / analysis
  • Microscopy / instrumentation*
  • Microscopy / methods
  • Proteins / analysis
  • Spectrum Analysis, Raman / instrumentation*
  • Spectrum Analysis, Raman / methods

Substances

  • Lipids
  • Proteins