NDLI: Microfluidic cell sorter for use in developing red fluorescent proteins with improved photostability

Content Provider	PubMed Central
Author	Davis, Lloyd M. Lubbeck, Jennifer L. Dean, Kevin M. Palmer, Amy E. Jimenez, Ralph
Abstract	This paper presents a novel microfluidic cytometer for mammalian cells that rapidly measures the irreversible photobleaching of red fluorescent proteins expressed within each cell and achieves high purity (>99%) selection of individual cells based on these measurements. The selection is achieved by using sub-millisecond timed control of a piezo-tilt mirror to steer a focused 1064-nm laser spot for optical gradient force switching following analysis of the fluorescence signals from passage of the cell through a series of 532-nm laser beams. In transit through each beam, the fluorescent proteins within the cell undergo conversion to dark states, but the microfluidic chip enables the cell to pass sufficiently slowly that recovery from reversible dark states occurs between beams, thereby enabling irreversible photobleaching to be quantified separately from the reversible dark-state conversion. The microfluidic platform achieves sorting of samples down to sub-millilitre volumes with minimal loss, wherein collected cells remain alive and can subsequently proliferate. The instrument provides a unique first tool for rapid selection of individual mammalian cells on the merits of photostability and is likely to form the basis of subsequent lab-on-a-chip platforms that combine photobleaching with other spectroscopic measurements for on-going research to develop advanced red fluorescent proteins by screening of genetic libraries.
Related Links	http://dx.doi.org/10.1039/c3lc50191d
Ending Page	2327
Page Count	8
Starting Page	2320
File Format	PDF
ISSN	14730197
e-ISSN	14730189
Journal	Lab on a chip
Issue Number	12
Volume Number	13
Language	English
Publisher Date	2013-01-01
Access Restriction	Open
Subject Keyword	Biochemistry Bioengineering Chemistry(all) Biomedical Engineering Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Chemistry Nanoscience and Nanotechnology Biochemistry Bioengineering Biomedical Engineering

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1	Ministry of Education (GoI), Department of Higher Education	Sanctioning Authority	https://www.education.gov.in/ict-initiatives
2	Indian Institute of Technology Kharagpur	Host Institute of the Project: The host institute of the project is responsible for providing infrastructure support and hosting the project	https://www.iitkgp.ac.in
3	National Digital Library of India Office, Indian Institute of Technology Kharagpur	The administrative and infrastructural headquarters of the project	Dr. B. Sutradhar bsutra@ndl.gov.in
4	Project PI / Joint PI	Principal Investigator and Joint Principal Investigators of the project	Dr. B. Sutradhar bsutra@ndl.gov.in Prof. Saswat Chakrabarti will be added soon
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