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Published on: June 4, 2020
Staining of cellular mitochondria with LDS-751
1Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 903 S. 4th St. Hamilton, MT 59840, USA. ssnyder@NIAID.NIH.gov
Journal of Immunological Methods
|November 1, 2001
Summary
The dye LDS-751 selectively binds to mitochondria, not the nucleus, in viable cells. Its fluorescence indicates mitochondrial polarization, not nuclear status, which is crucial for accurate cell analysis.
Area of Science:
- Cell Biology
- Biochemistry
- Microscopy
Background:
- The dye LDS-751 is often used to assess nuclear status in cells.
- Accurate cell analysis relies on specific and reliable staining methods.
Purpose of the Study:
- To investigate the precise cellular localization and binding characteristics of the dye LDS-751.
- To determine if LDS-751 can be reliably used as a nuclear stain.
Main Methods:
- Incubation of viable, nucleated cells (fibroblasts and monocytes) with LDS-751.
- Confocal microscopy was used to examine colocalization with nuclear stain acridine orange.
- Comparison of LDS-751 staining patterns with rhodamine 123, a known mitochondrial stain.
- Assessment of LDS-751 and rhodamine 123 fluorescence after mitochondrial depolarization using phenyl arsine oxide and carbonyl cyanide m-chlorophenylhydrazone (CCCP).
Main Results:
- LDS-751 exclusively bound to mitochondria, showing minimal colocalization with the nucleus.
- The staining pattern of LDS-751 closely matched that of rhodamine 123.
- Mitochondrial depolarization significantly reduced both LDS-751 and rhodamine 123 fluorescence.
- Consistent staining was observed across a range of LDS-751 concentrations (0.02-20 microg/ml) and cell types.
Conclusions:
- LDS-751 is excluded from the nucleus and binds to the polarized membranes of mitochondria.
- Using LDS-751 fluorescence to determine nuclear status is unwarranted and can lead to errors, especially if mitochondria are depolarized.
