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Multiplexed and reproducible high content screening of live and fixed cells using Dye Drop

Author

Listed:
  • Caitlin E. Mills

    (Harvard Medical School)

  • Kartik Subramanian

    (Harvard Medical School
    Bristol Myers Squibb)

  • Marc Hafner

    (Harvard Medical School
    Genentech, Inc.)

  • Mario Niepel

    (Harvard Medical School
    Ribon Therapeutics, Inc.)

  • Luca Gerosa

    (Harvard Medical School
    Genentech, Inc.)

  • Mirra Chung

    (Harvard Medical School)

  • Chiara Victor

    (Harvard Medical School)

  • Benjamin Gaudio

    (Harvard Medical School)

  • Clarence Yapp

    (Harvard Medical School)

  • Ajit J. Nirmal

    (Harvard Medical School
    Dana-Farber Cancer Institute)

  • Nicholas Clark

    (Harvard Medical School)

  • Peter K. Sorger

    (Harvard Medical School)

Abstract

High-throughput measurement of cells perturbed using libraries of small molecules, gene knockouts, or different microenvironmental factors is a key step in functional genomics and pre-clinical drug discovery. However, it remains difficult to perform accurate single-cell assays in 384-well plates, limiting many studies to well-average measurements (e.g., CellTiter-Glo®). Here we describe a public domain Dye Drop method that uses sequential density displacement and microscopy to perform multi-step assays on living cells. We use Dye Drop cell viability and DNA replication assays followed by immunofluorescence imaging to collect single-cell dose-response data for 67 investigational and clinical-grade small molecules in 58 breast cancer cell lines. By separating the cytostatic and cytotoxic effects of drugs computationally, we uncover unexpected relationships between the two. Dye Drop is rapid, reproducible, customizable, and compatible with manual or automated laboratory equipment. Dye Drop improves the tradeoff between data content and cost, enabling the collection of information-rich perturbagen-response datasets.

Suggested Citation

  • Caitlin E. Mills & Kartik Subramanian & Marc Hafner & Mario Niepel & Luca Gerosa & Mirra Chung & Chiara Victor & Benjamin Gaudio & Clarence Yapp & Ajit J. Nirmal & Nicholas Clark & Peter K. Sorger, 2022. "Multiplexed and reproducible high content screening of live and fixed cells using Dye Drop," Nature Communications, Nature, vol. 13(1), pages 1-18, December.
  • Handle: RePEc:nat:natcom:v:13:y:2022:i:1:d:10.1038_s41467-022-34536-7
    DOI: 10.1038/s41467-022-34536-7
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