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A rapid micro method for counting cells "in situ" using a fluorogenic alkaline phosphatase enzyme assay.
L I Huschtscha1, F C Lucibello, W F Bodmer
1Imperial Cancer Research Fund, London, United Kingdom.
Summary
A novel alkaline phosphatase assay offers a rapid and efficient method for estimating cell numbers in situ. This fluorogenic enzyme assay accurately correlates enzyme activity with cell count across various cell types for growth studies.
Area of Science:
- Biotechnology
- Cell Biology
- Enzyme Assays
Background:
- Accurate cell counting is crucial for biological research.
- Existing methods can be time-consuming or require cell lysis.
Purpose of the Study:
- To develop and validate a new in situ cell counting method.
- To assess the utility of alkaline phosphatase activity for cell enumeration.
Main Methods:
- Development of a fluorogenic enzyme assay measuring alkaline phosphatase activity.
- Correlation of enzyme activity with cell number across diverse cell lines (Hep-2, SW1222, SW837, LS174T, HT29, MRC5, NIH-3T3).
- Validation of assay performance over time in culture.
Main Results:
- A strong positive correlation was observed between cell number and alkaline phosphatase activity.
- The assay accurately estimated relative cell numbers from 10^4 to 5 x 10^5.
- This relationship remained consistent over time in culture for tested cell types.
Conclusions:
- The alkaline phosphatase assay (ALP assay) provides a rapid, efficient, and reliable method for in situ cell counting.
- This technique is suitable for various cell types and applicable to growth assay studies.