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Three-Dimensional Spot Detection in Ratiometric Fluorescence Imaging For Measurement of Subcellular Organelles

Thursday, November 07, 2013 — Poster Session II

12:00 p.m. – 2:00 p.m.

FAES Academic Center (Upper-Level Terrace)




  • W.W. Lau
  • C Johnson
  • R Kuo
  • S Lioi
  • J.A. Mindell


Lysosomes are subcellular organelles playing a vital role in the endocytosis process of the cell. Lysosomal acidity is an important factor in assuring proper functioning of the enzymes within the organelle, and can be assessed by labeling the lysosomes with pH-sensitive fluorescence probes. To enhance our understanding of the acidification mechanisms, the goal of this work is to develop a method that can accurately detect and characterize the acidity of each lysosome captured in ratiometric fluorescence images. We present an algorithm that utilizes h-dome transformation and reconciles spots detected independently from two wavelength channels. We evaluated our algorithm using simulated images for which the exact locations and intensity ratios of the spots were known. The h-dome algorithm achieved an f-score as high as 0.952. Furthermore, the reconciliation procedure was able to improve the performance for images with low signal-to-noise ratios. We also computed the fluorescence ratios from lysosomes in live HeLa cell images, and found the resulting emission ratio curve to be in agreement with those found in the literature. Empirical comparison with a conventional method also indicates that the proposed method is superior in terms of consistency.

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