Estimating weak ratiometric signals in imaging data I. Dual-channel data

Josef Broder, Anirban Majumder, Erika Porter, Ganesh Srinivasamoorthy, Charles Keith, James Lauderdale, Andrew Sornborger

Research output: Contribution to journalArticlepeer-review

17 Scopus citations


Ratiometric fluorescent indicators are becoming increasingly prevalent in many areas of biology. They are used for making quantitative measurements of intracellular free calcium both in vitro and in vivo, as well as measuring membrane potentials, pH, and other important physiological variables of interest to researchers in many subfields. Often, functional changes in the fluorescent yield of ratiometric indicators are small, and the signal-to-noise ratio (SNR) is of order unity or less. In particular, variability in the denominator of the ratio can lead to very poor ratio estimates. We present a statistical optimization method for objectively detecting and estimating ratiometric signals in dual-wavelength measurements of fluorescent, ratiometric indicators that improves on standard methods. With the use of an appropriate statistical model for ratiometric signals and by taking the pixel-pixel covariance of an imaging dataset into account, we are able to extract user-independent spatiotemporal information that retains high resolution in both space and time.

Original languageEnglish
Pages (from-to)2921-2931
Number of pages11
JournalJournal of the Optical Society of America A: Optics and Image Science, and Vision
Issue number9
StatePublished - Sep 2007
Externally publishedYes


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