Computerized microassay of keratinocyte cell-plastic attachment and proliferation for assessing net stimulatory, inhibitory and toxic effects of compounds on nonimmortalized cell lines

Sergei A. Grando, Raul Cabrera, Bruce S. Hostager, Paul L. Bigliardi, John S. Blake, Michael J. Herron, Mark V. Dahl, Robert D. Nelson

Research output: Contribution to journalArticle

11 Scopus citations

Abstract

Testing of pharmacological agents that affect growth of epidermal keratinocytes (EK) requires a standardized assay. We have developed an assay measuring net effects of stimulatory (e.g. growth factors), inhibitory (e.g. methotrexate) or toxic (e.g. Triton X-100) compounds. The amount of crystal violet staining viable EK attached to the wells of standard 96-well microplates is measured in situ using an ELISA plate reader. Optical density readings are directly converted into cell counts by computer software. Counts obtained by this method strongly correlate with the results obtained using the [3H]thy-midine uptake assay and direct cell counts. The assay standardizes measurements of nonimmortalized EK lines with different innate proliferative properties and allows accurate quantitation of EK numbers in the range of 2,500-500.000 EK/well.

Original languageEnglish (US)
Pages (from-to)135-147
Number of pages13
JournalSkin Pharmacology and Physiology
Volume6
Issue number2
DOIs
StatePublished - Jan 1 1993

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Keywords

  • Attachment
  • Growth inhibition
  • Growth stimulation
  • In vitro assay
  • Keratinocytes
  • Proliferation
  • Toxicity

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