High Content Image Analysis using a Laser Scanning Microplate Cytometer
Poster Feb 28, 2008
Paul Wylie, Ben Schenker, Sarah Payne and Andrew Goulter
Microscope-based high-content instruments offer high optical resolution, however, the limited field of view afforded by their objective lenses can mean lengthy read times for some assays, especially where multiple image capture per well is required. Wherever possible, only a small percentage of the total number of cells present in a test well are analysed to keep plate read times at a minimum, which may not always be ideal. Microplate laser- scanning cytometers, such as TTP LabTech’s Acumen® eX3, are an alternative platform. This technology offershigher throughput than imaging based systems processing 300,000 wells of data in 24 hours in 1536 plates. The Acumen eX3 is capable of generating cytometric data and TIFF images (8 or 16-bit) simultaneously. The images represent the whole well and correlate with those captured using a 20x microscope objective.The images produced can be subjected to the full range of algorithms contained in commercially available image analysis software. In most cases where image analysis and cytometric analysis have been validated, both methods were comparable where single cells were represent. For a limited number of applications, however, secondary information provided by image analysis may increase understanding. Such assays include angiogenesis, cell colony formation and tissue scanning.
Microplate cytometry offers whole well, high throughput capability with the increased flexibility of simultaneous cytometric and TIFF image outputs and presents a comprehensive solution for high content analysis studies.
When there is a need to quickly analyze samples using a number of different PCR assays, it is likely that optimal conditions for each assay will not be the same. First, different assays often will require different annealing temperatures for their primers. In addition, amplicons may be designed to be of different lengths and therefore require varying durations of the extension step.READ MORE