Simple liquid-chromatographic method for Nile Red quantification in cell culture in spite of photobleaching

J Chromatogr B Analyt Technol Biomed Life Sci. 2003 Apr 25;787(2):415-9. doi: 10.1016/s1570-0232(02)00962-5.


Nile Red fluorescent marker is widely-used for different purposes, such as staining cell structures and for the visualization and localization of colloidal drug carriers. However, when fluorescence-dependent imaging or quantification is performed, the risk of inexact results is increased due to photobleaching. The proposed, simple quantification method of using an HPLC-UV-Vis system allows the determination of Nile Red even in photobleached samples. The intra- and inter-assay accuracies for all analytes were found to be within 94.9 and 100.8%, respectively, of target values. When samples underwent photobleaching by laser, UV-Vis detection varied at around 99+/-5%, whereas fluorescence decreased down to 86%. Such results show this method to be interesting for approaches where quantification should be performed after analysis such as fluorescent imaging.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Calibration
  • Cell Culture Techniques
  • Chromatography, High Pressure Liquid / methods*
  • Oxazines / analysis*
  • Photochemistry
  • Spectrophotometry, Ultraviolet


  • Oxazines
  • nile red