Effects of pH on FAD autofluorescence lifetimes

Rebecca Schmitz, Christine Walsh, Alex J. Walsh, Melissa C. Skala

Research output: Chapter in Book/Report/Conference proceedingConference contribution

Abstract

Fluorescence lifetime imaging (FLIM) data has consistently revealed a significant difference in mean FAD lifetime between in vivo and in vitro models. We hypothesized that the observed difference in mean FAD lifetime could be a result of environmental differences, such as differing glucose levels, oxygen levels, or pH, between the two models. We investigated the effects of environmental pH on the autofluorescence lifetime of FAD. We adjusted the pH of HEPEScontaining media using sodium hydroxide and hydrochloric acid. We then replaced the normal media of plated BT474 cells with the pH-adjusted media, allowed 20 minutes for cellular changes to occur, and then imaged the cells using time correlated single photon counting FLIM. We found that the mean lifetime of FAD increased with increased pH, resulting in a significant increase between pH 3.9, 6.2, 7.4, 9.1, and 9.5. The mean lifetime of NAD(P)H decreased at pH 3.9, 9.1, and 9.5 relative to a control pH of 7.3, and the optical redox ratio showed no significant changes except at pH 3.9 relative to a control pH of 7.3. These results suggest that the difference in mean FAD lifetime between in vivo and cell culture models could result from pH changes in the cellular environment.

Original languageEnglish (US)
Title of host publicationMultiphoton Microscopy in the Biomedical Sciences XIX
EditorsAmmasi Periasamy, Peter T. C. So, Karsten Konig
PublisherSPIE
ISBN (Electronic)9781510624061
DOIs
StatePublished - 2019
EventMultiphoton Microscopy in the Biomedical Sciences XIX 2019 - San Francisco, United States
Duration: Feb 3 2019Feb 6 2019

Publication series

NameProgress in Biomedical Optics and Imaging - Proceedings of SPIE
Volume10882
ISSN (Print)1605-7422

Conference

ConferenceMultiphoton Microscopy in the Biomedical Sciences XIX 2019
CountryUnited States
CitySan Francisco
Period2/3/192/6/19

Keywords

  • FAD
  • Fluorescence lifetime imaging
  • Multiphoton microscopy
  • pH

ASJC Scopus subject areas

  • Electronic, Optical and Magnetic Materials
  • Biomaterials
  • Atomic and Molecular Physics, and Optics
  • Radiology Nuclear Medicine and imaging

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