Automated 96-well format high throughput colony formation assay for siRNA library screen

STAR Protoc. 2022 May 1;3(2):101355. doi: 10.1016/j.xpro.2022.101355. eCollection 2022 Jun 17.

Abstract

The colony formation assay is the gold-standard technique to assess cell viability after treatment with cytotoxic reagents, ionizing radiation, and cytotoxic combinatorial treatments. This protocol describes a high-throughput automated and high-content imaging approach to screen siRNA molecular libraries in HeLa cervical cancer cells in 96-well format. We detail reverse transfection of cells with siRNAs, followed by ionizing radiation, fixing, and staining of the plates for automated colony counting. This protocol can be used across a broad range of cell types. For complete details on the use and execution of this protocol, please refer to Tiwana et al. (2015).

Keywords: Cancer; Cell Biology; Cell culture; Cell-based Assays; High Throughput Screening; Molecular Biology; Single Cell.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Gene Library
  • High-Throughput Screening Assays* / methods
  • RNA, Small Interfering / genetics
  • Radiation, Ionizing*
  • Transfection

Substances

  • RNA, Small Interfering