Quantitative pH Imaging in Cells Using Confocal Fluorescence Lifetime Imaging Microscopy

The pH-sensitive probe carboxy SNAFL-1 can be used for imaging using ratiometric and fluorescence lifetime techniques. The former method suffers from the drawback that quantitative pH imaging in cells requires a time-consuming and cumbersome calibration procedure. In contrast, straightforward calibr...

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Bibliographic Details
Published inAnalytical biochemistry Vol. 227; no. 2; pp. 302 - 308
Main Authors Sanders, R., Draaijer, A., Gerritsen, H.C., Houpt, P.M., Levine, Y.K.
Format Journal Article
LanguageEnglish
Published United States Elsevier Inc 20.05.1995
Subjects
Animals
Buffers
Calibration
Cells - chemistry
CHO Cells
Cricetinae
Hydrogen-Ion Concentration
Microscopy, Confocal
Microscopy, Fluorescence - methods
Proteins - chemistry
Sensitivity and Specificity
Solubility
Tumor Cells, Cultured
Water - chemistry
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Summary:The pH-sensitive probe carboxy SNAFL-1 can be used for imaging using ratiometric and fluorescence lifetime techniques. The former method suffers from the drawback that quantitative pH imaging in cells requires a time-consuming and cumbersome calibration procedure. In contrast, straightforward calibrations in buffer suffice for fluorescence lifetime imaging. This is illustrated here by a comparative study of the two techniques under different controlled conditions. The effect of probe concentration, protein concentration, and hydrophobicity, the contents of damaged cells and living cells on the emission ratio, and the fluorescence lifetime of carboxy SNAFL-1 were studied. The results clearly demonstrate that the fluorescence lifetime imaging technique is more convenient than the ratiometric method for pH determination.
Bibliography:ObjectType-Article-1
SourceType-Scholarly Journals-1
ObjectType-Feature-2
content type line 23
ISSN:0003-2697
1096-0309
DOI:10.1006/abio.1995.1285