TY - GEN
T1 - Effects of pH on FAD autofluorescence lifetimes
AU - Schmitz, Rebecca
AU - Walsh, Christine
AU - Walsh, Alex J.
AU - Skala, Melissa C.
N1 - Funding Information:
Research reported in this publication was supported by the National Cancer Institute of the National Institutes of Health under Award Numbers R01 CA205101, R01 CA211082, R01 CA185747, and the WISCIENCE Entering Research Summer Program. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Publisher Copyright:
© COPYRIGHT SPIE. Downloading of the abstract is permitted for personal use only.
PY - 2019
Y1 - 2019
N2 - 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.
AB - 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.
KW - FAD
KW - Fluorescence lifetime imaging
KW - Multiphoton microscopy
KW - pH
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U2 - 10.1117/12.2513482
DO - 10.1117/12.2513482
M3 - Conference contribution
AN - SCOPUS:85067810845
T3 - Progress in Biomedical Optics and Imaging - Proceedings of SPIE
BT - Multiphoton Microscopy in the Biomedical Sciences XIX
A2 - Periasamy, Ammasi
A2 - So, Peter T. C.
A2 - Konig, Karsten
PB - SPIE
T2 - Multiphoton Microscopy in the Biomedical Sciences XIX 2019
Y2 - 3 February 2019 through 6 February 2019
ER -