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A Plug-Based Microfluidic System for Dispensing Lipidic Cubic Phase (LCP) Material Validated by Crystallizing Membrane Proteins in Lipidic Mesophases

Li, Liang and Fu, Qiang and Kors, Christopher A. and Stewart, Lance and Nollert, Peter and Laible, Philip D. and Ismagilov, Rustem F. (2010) A Plug-Based Microfluidic System for Dispensing Lipidic Cubic Phase (LCP) Material Validated by Crystallizing Membrane Proteins in Lipidic Mesophases. Microfluidics and Nanofluidics, 8 (6). pp. 789-798. ISSN 1613-4982. PMCID PMC2868346.

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This article presents a plug-based microfluidic system to dispense nanoliter-volume plugs of lipidic cubic phase (LCP) material and subsequently merge the LCP plugs with aqueous plugs. This system was validated by crystallizing membrane proteins in lipidic mesophases, including LCP. This system allows for accurate dispensing of LCP material in nanoliter volumes, prevents inadvertent phase transitions that may occur due to dehydration by enclosing LCP in plugs, and is compatible with the traditional method of forming LCP material using a membrane protein sample, as shown by the successful crystallization of bacteriorhodopsin from Halobacterium salinarum. Conditions for the formation of LCP plugs were characterized and presented in a phase diagram. This system was also implemented using two different methods of introducing the membrane protein: (1) the traditional method of generating the LCP material using a membrane protein sample and (2) post LCP-formation incorporation (PLI), which involves making LCP material without protein, adding the membrane protein sample externally to the LCP material, and allowing the protein to diffuse into the LCP material or into other lipidic mesophases that may result from phase transitions. Crystals of bacterial photosynthetic reaction centers from Rhodobacter sphaeroides and Blastochloris viridis were obtained using PLI. The plug-based, LCP-assisted microfluidic system, combined with the PLI method for introducing membrane protein into LCP, should be useful for minimizing consumption of samples and broadening the screening of parameter space in membrane protein crystallization.

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URLURL TypeDescription ReadCube access CentralArticle
Ismagilov, Rustem F.0000-0002-3680-4399
Additional Information:© Springer 2010. Received: 5 August 2009. Accepted: 22 September 2009. Published online: 14 October 2009. This work was supported through Accelerated Technologies Center for Gene to 3D Structure (ATCG3D) funded by the National Institute of General Medical Sciences (NIGMS), National Center for Research Resources under the PSI-2 Specialized Center program (U54 GM074961); the National Institutes of Health Roadmap for Medical Research (R01 GM075827 and P01 GM75913), and University of Chicago/Argonne National Laboratory (ANL) Collaborative Seed Funding. We thank Nina Ponomarenko and James R. Norris at the University of Chicago for samples of Reaction Center from B. viridis. We thank Ray C. Stevens and Peter Kuhn for helpful discussion and Elizabeth B. Haney for contributions in writing and editing this manuscript. Use of the ANL General Medicine and Cancer Institute Collaborative Access Team (GM/CA CAT) beamlines at the Advanced Photon Source was supported by the U.S. Department of Energy, Basic Energy Sciences, Office of Science, under Contract No. DE-AC02-06CH11357. GM/CA CAT has been funded in whole or in part with Federal funds from the National Cancer Institute (Y1-CO-1020) and the NIGMS (Y1-GM-1104).
Funding AgencyGrant Number
National Institute of General Medical SciencesUNSPECIFIED
NIHU54 GM074961
NIHR01 GM075827
NIHP01 GM75913
University of Chicago/Argonne National Laboratory (ANL) Collaborative Seed FundingUNSPECIFIED
Department of Energy (DOE)DE-AC02-06CH11357
National Cancer InstituteUNSPECIFIED
Subject Keywords:Droplet; Plugs; Lipidic cubic phase; Membrane protein; Protein crystallization
Issue or Number:6
PubMed Central ID:PMC2868346
Record Number:CaltechAUTHORS:20130821-160723624
Persistent URL:
Official Citation:Li, L., Fu, Q., Kors, C.A. et al. A plug-based microfluidic system for dispensing lipidic cubic phase (LCP) material validated by crystallizing membrane proteins in lipidic mesophases. Microfluid Nanofluid 8, 789–798 (2010).
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:40822
Deposited By: Whitney Barlow
Deposited On:23 Aug 2013 23:45
Last Modified:05 Mar 2020 15:44

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