Interference and Artifacts in High-content Screening

Excerpt

Since the late 1990s, high-content screening (HCS) approaches have contributed greatly to cell and chemical biology, drug discovery, and toxicology research. Despite the numerous advantages of multi-parameter interrogation of cell models using HCS approaches, these assay formats are also susceptible to artifacts and interference. This chapter describes 1) autofluorescence interference contributed by media, cells, and tissues which may complicate or preclude HCS assay development, 2) interference by environmental or microorganism contaminants, and 3) compound-mediated autofluorescence, fluorescence quenching, and cellular injury/cytotoxicity which may obscure whether compounds modulate the desired target or cellular phenotype or may produce false-positives or -negatives. It provides best-practice recommendations for identifying and mitigating such interference through the application of experimental design strategies, statistics, orthogonal, and counter-screens, and the use of reference compounds. Adoption of these recommendations should enhance the quality of chemical matter identified by HCS assays.