A miniaturized device for bioluminescence analysis of caspase-3/7 activity in a single apoptotic cell

Eva Adamova, Marcela Lišková, Eva Matalová, Karel Klepárník

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Caspases are key enzymes activated during the apoptotic machinery. Apoptosis as a way of programmed cell death becomes deregulated in some pathologies including cancer transformations, neurodegenerative, or autoimmune diseases. Most of the methods available for the detection of apoptosis and caspases provide qualitative information only or quantification data as an average from cell populations or cell lysates. Several reports point to the importance of more accurate single-cell analyses in biomedical studies due to heterogeneity at tissue as well as cell level. To meet these requirements, we developed a miniaturized device enabling detection and quantification of active caspase-3/7 in individual cells at a femtogram level (10-15 g). The active caspase-3/7 detection protocol is based on the bioluminescence chemistry commercially available as a Caspase-Glo™ 3/7 reagent developed by Promega. As a model, we used human stem cells treated by camptothecin to induce apoptosis. Individual apoptotic cells were captured from a culture medium under a microscope and transferred by a micromanipulation system into a detection capillary containing 2 μl of the reagent. Cells without activation by camptothecin served as negative controls. The detection limit of active caspase-3/7 achieved in the miniaturized system was determined as 0.20 and limit of quantification as 0.65 of the amount found in a single apoptotic human stem cell. Such a sensitive method could have a wide application potential in laboratory medicine and related clinically oriented research.

Original languageEnglish (US)
Pages (from-to)5389-5394
Number of pages6
JournalAnalytical and Bioanalytical Chemistry
Volume406
Issue number22
DOIs
StatePublished - Jan 1 2014

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Bioluminescence
Caspase 7
Caspases
Caspase 3
Camptothecin
Cells
Apoptosis
Stem cells
Equipment and Supplies
Bioelectric potentials
Pathology
Cell death
Medicine
Machinery
Culture Media
Microscopes
Stem Cells
Chemical activation
Micromanipulation
Tissue

All Science Journal Classification (ASJC) codes

  • Analytical Chemistry
  • Biochemistry
  • Medicine(all)

Cite this

A miniaturized device for bioluminescence analysis of caspase-3/7 activity in a single apoptotic cell. / Adamova, Eva; Lišková, Marcela; Matalová, Eva; Klepárník, Karel.

In: Analytical and Bioanalytical Chemistry, Vol. 406, No. 22, 01.01.2014, p. 5389-5394.

Research output: Contribution to journalArticle

Adamova, Eva ; Lišková, Marcela ; Matalová, Eva ; Klepárník, Karel. / A miniaturized device for bioluminescence analysis of caspase-3/7 activity in a single apoptotic cell. In: Analytical and Bioanalytical Chemistry. 2014 ; Vol. 406, No. 22. pp. 5389-5394.
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