High-Content Reporter Assays

Methods Mol Biol. 2018:1755:179-195. doi: 10.1007/978-1-4939-7724-6_13.

Abstract

While luminescent reporter gene assays allow for a rapid and relatively interference free assessment of the activation state of a luminescent reporter, fluorescent reporters do not. They suffer from artifacts such as compound fluorescence and cellular debris which makes the assessment of whole well fluorescence signals difficult. However, the use of high-content screening allows for the isolation of individual cells, segmentation and thus enables the screener to utilize fluorescent reporters to assess the activation state of such a high-content reporter on a cell by cell level, thus minimizing artifacts. Here we discuss the use of such a high-content reporter that enables screening for compounds useful for HIV reactivation on Jurkat cells with high-content screening.

Keywords: 1536-well plates; Drug discovery; HCS; HIV; HTS; High-content screening; High-throughput screening; Latency; Lentivirus; Reactivation; Reporter gene assay.

MeSH terms

  • Alkaline Phosphatase / chemistry
  • Alkaline Phosphatase / genetics
  • Biological Assay / instrumentation
  • Biological Assay / methods*
  • Enzyme Assays / instrumentation
  • Enzyme Assays / methods
  • Genes, Reporter / genetics*
  • Green Fluorescent Proteins / chemistry
  • Green Fluorescent Proteins / genetics
  • HIV / physiology
  • High-Throughput Screening Assays / instrumentation
  • High-Throughput Screening Assays / methods*
  • Humans
  • Intravital Microscopy / instrumentation
  • Intravital Microscopy / methods
  • Jurkat Cells
  • Luciferases / chemistry
  • Luciferases / genetics
  • Microscopy, Fluorescence / instrumentation
  • Microscopy, Fluorescence / methods
  • Transfection / instrumentation
  • Transfection / methods*
  • Virus Activation / genetics
  • beta-Galactosidase / chemistry
  • beta-Galactosidase / genetics

Substances

  • Green Fluorescent Proteins
  • Luciferases
  • Alkaline Phosphatase
  • beta-Galactosidase