Fluorescence imaging plate reader

One very important device is the plate reader, which can be rate limiting in HTS. Most laboratories use multimodal readers that can detect various forms of fluorescence as well as luminescence and absorbance. The traditional readers are photomultiplier-based devices that usually read from one well to the next. This process can take considerable time for 384-well and higher-density plates. A more desirable HTS reader type images the entire plate with a charge-coupled device (CCD) camera. The latter device is usually a faster reader for 384-well and higher-density plates. Imagers can capture significant cross talk from one well to another, but with proper set up, they can produce data of equal quality. 

Fluorescent, voltage-sensitive dyes can be employed to investigate the activity of ion channels by reflecting the cellular membrane potential (Epps et al. 1994 Plasek and Sigler 1996). This approach is suitable for use in conjunction with different measurement systems including Fluorometric imaging plate readers (FLIPR), Voltage/Ion probe readers (VIPR) or conventional fluorescence readers. 

Some cellular assays also present the ability to measure kinase activation using recombinantly expressed intracellular sensor molecules that fluoresce in response to phosphorylation. These sensors are reviewed by Lawrence et al. (2007). The most interesting versions use yellow and blue fluorescent proteins to create a FRET through phosphorylation-induced folding with an intramolecular SH2 domain. SH2 domains are known to bind tightly to phosphotyrosine molecules and have been successfully shown to serve as phosphotyrosine detectors. In this type of assay, imaging is not necessary because the total fluorescence can be measured in a population of cells using a standard plate reader. 

The AequoScreen system developed by Euroscreen utilizes this biolumines-cent Ca -indicator in cell-based assays. This system offers an interesting alternative to the traditional, fluorescence-based assays for intracellular Ca measurements (see Section 10.3.1 10.3.1 Fluorimetric Imaging Plate Reader (FLIPR) ). 

Abbreviations FLIPR, fluorescence imaging plate reader VIPR, voltage ion probe reader. 

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