Saturation Dynamics Measures Absolute Cross Section and Generates Contrast within a Neuron

Kumar, Suraj and Singh, Aditya and Singh, Vijay R and George, Jude B and Balaji, J (2016) Saturation Dynamics Measures Absolute Cross Section and Generates Contrast within a Neuron. In: BIOPHYSICAL JOURNAL, 111 (6). pp. 1328-1336.

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Abstract

The intensity required to optically saturate a chromophore is a molecular property that is determined by its absorption cross section (sigma) and the excited state lifetime. We present an analytical description of such a system and show that fluorescence around the onset of saturation is characterized by product of absorption cross section and lifetime. Using this approach we formulate a generalized method for measuring the multiphoton cross section of fluorophores and use it to obtain the absolute three-photon cross-section spectra of tryptophan. We find that the tryptophan three-photon cross section ranges from 0.28 S.I. units (m(6)s(2)photon(-2)) at 870 nm to 20 S.I. units at 740 nm. Further, we show that the product of molecular rate of excitation and de-excitation, denoted as beta, serves as a vital contrasting agent for imaging local environment. Our contrast parameter, 0, is related to fraction of the population present in the excited state and is independent of the fluorophore concentration. We show that beta-imaging can be carried out in a regular two-photon microscope setup through a series of intensity scans. Using enhanced green fluorescent protein (EGFP) fluorescence from the brain slices of Thy-1 EGFP transgenic mice, we show that there is an inherent, concentration independent, variation in contrast across the soma and the dendrite.

Item Type:	Journal Article
Publication:	BIOPHYSICAL JOURNAL
Additional Information:	Copy right for this article belongs to the CELL PRESS, 600 TECHNOLOGY SQUARE, 5TH FLOOR, CAMBRIDGE, MA 02139 USA
Department/Centre:	Division of Biological Sciences > Centre for Neuroscience Division of Interdisciplinary Sciences > Centre for Nano Science and Engineering
Date Deposited:	28 Oct 2016 07:13
Last Modified:	28 Oct 2016 07:13
URI:	http://eprints.iisc.ac.in/id/eprint/55142

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