Imaging mitochondria in living corneal endothelial cells using autofluorescence microscopy

Nicholas A. Ramey, Choul Yong Park, Peter L. Gehlbach, Roy S. Chuck

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

We report noninvasive autofluorescence mitochondrial imaging in cultured human corneal endothelial cells (HCECs). HCECs harvested from eye bank corneas were cultured in thin glass-bottom plates. Mitochondria were imaged with an autofluorescence microscope using a DAPI filter set (excitation: G365, emission: band pass 445/50) and then, after fixation with 4% paraformaldehyde, cells were stained with MitoTracker™ Green FM (MTG). Both images were aligned using a linear conformal algorithm for image mapping based on manually selected corresponding feature points, and then mathematically compared using two-dimensional spatial image correlation coefficients. Autofluorescence imaging provided highly resolved mitochondrial signals from living HCECs, comparable to those taken with MTG. Both techniques yielded very similar images at high magnification and high resolution, demonstrating the tubular morphology and cytoplasmic distribution that are characteristic of mitochondria. Image registration using a linear conformal mapping technique and cross-correlations showed high correlation of overlapping autofluorescence and MTG images. This study validates the novel use of autofluorescence vital imaging as a noninvasive, inexpensive and functional alternative to the mitochondria-specific dyes in cultured HCEC. This noninvasive mitochondrial imaging technique can be useful in future applications studying mitochondrial biology of ocular cells.

Original languageEnglish (US)
Pages (from-to)1325-1329
Number of pages5
JournalPhotochemistry and Photobiology
Volume83
Issue number6
DOIs
StatePublished - Nov 2007
Externally publishedYes

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Mitochondria
mitochondria
Endothelial cells
Microscopy
Optical Imaging
Microscopic examination
Endothelial Cells
microscopy
Imaging techniques
frequency modulation
Eye Banks
Conformal mapping
Image registration
Cornea
Glass
Cell Biology
Microscopes
Coloring Agents
cornea
conformal mapping

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Biophysics

Cite this

Imaging mitochondria in living corneal endothelial cells using autofluorescence microscopy. / Ramey, Nicholas A.; Park, Choul Yong; Gehlbach, Peter L.; Chuck, Roy S.

In: Photochemistry and Photobiology, Vol. 83, No. 6, 11.2007, p. 1325-1329.

Research output: Contribution to journalArticle

Ramey, Nicholas A. ; Park, Choul Yong ; Gehlbach, Peter L. ; Chuck, Roy S. / Imaging mitochondria in living corneal endothelial cells using autofluorescence microscopy. In: Photochemistry and Photobiology. 2007 ; Vol. 83, No. 6. pp. 1325-1329.
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