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Methods Mol Biol. 2005;289:251-62.

Fluorimetric DNA assay of cell number.

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1
Histopathology Unit, London Rsearch Intitute, Cancer Research UK, London, UK.

Abstract

This fluorimetric assay has utility for the accurate assessment of cultured epidermal cell numbers by virtue of their deoxyribonucleic acid content, which is the most significant correlate available. The improvement in fluorochromes in the recent past makes PicoGreen the dye of choice for this, with its greatly increased sensitivity (+/- 50 cells) over the Hoechst and DAPI stains and which remains linear over several orders of magnitude with a single dye concentration. The assay involves minimal liquid handling to achieve cell disruption by sodium dodecyl sulfate in saline sodium citrate buffer, and PicoGreen staining is rapidly assayed by a multiwell plate reading fluorimeter, which can be automated for robotic high throughput use. Highly fibrous cells like epidermal keratinocytes can be disrupted using 8 M urea and assayed after dilution. The assay is also compatible with tritiated thymidine incorporation.

PMID:
15502190
[Indexed for MEDLINE]
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