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@article{1227815,
author = {Lu, Z.H. and Barnard, D. and Shaikh, Tanvir and Meng, X. and Mannella, C.A. and Yassin, A.S. and Agrawal, R.K. and Wagenknecht, T. and Lu, T.M.},
article_location = {BRISTOL},
article_number = {11},
doi = {http://dx.doi.org/10.1088/0960-1317/24/11/115001},
keywords = {microfluidics; micronozzle; microspray; microdroplet; monolithic device; cryo-EM; time-resolved TEM},
language = {eng},
issn = {0960-1317},
journal = {JOURNAL OF MICROMECHANICS AND MICROENGINEERING},
title = {Gas-assisted annular microsprayer for sample preparation for time-resolved cryo-electron microscopy},
url = {http://iopscience.iop.org/0960-1317/24/11/115001/pdf/0960-1317_24_11_115001.pdf},
volume = {24},
year = {2014}
}
TY - JOUR
ID - 1227815
AU - Lu, Z.H. - Barnard, D. - Shaikh, Tanvir - Meng, X. - Mannella, C.A. - Yassin, A.S. - Agrawal, R.K. - Wagenknecht, T. - Lu, T.M.
PY - 2014
TI - Gas-assisted annular microsprayer for sample preparation for time-resolved cryo-electron microscopy
JF - JOURNAL OF MICROMECHANICS AND MICROENGINEERING
VL - 24
IS - 11
SP - "nestránkováno"
EP - "nestránkováno"
PB - IOP PUBLISHING LTD
SN - 09601317
KW - microfluidics
KW - micronozzle
KW - microspray
KW - microdroplet
KW - monolithic device
KW - cryo-EM
KW - time-resolved TEM
UR - http://iopscience.iop.org/0960-1317/24/11/115001/pdf/0960-1317_24_11_115001.pdf
L2 - http://iopscience.iop.org/0960-1317/24/11/115001/pdf/0960-1317_24_11_115001.pdf
N2 - Time-resolved cryo-electron microscopy (TRCEM) has emerged as a powerful technique for transient structural characterization of isolated biomacromolecular complexes in their native state within the time scale of seconds to milliseconds. For TRCEM sample preparation, a microfluidic device has been demonstrated to be a promising approach to facilitate TRCEM biological sample preparation. It is capable of achieving rapidly aqueous sample mixing, controlled reaction incubation, and sample deposition on electron microscopy (EM) grids for rapid freezing. One of the critical challenges is to transfer samples to cryo-EM grids from the microfluidic device. By using a microspraying method, the generated droplet size needs to be controlled to facilitate thin ice film formation on the grid surface for efficient data collection, whilst not being so thin that it dries out before freezing, i.e. an optimized mean droplet size needs to be achieved. In this work, we developed a novel monolithic three dimensional (3D) annular gas-assisted microfluidic sprayer using 3D MEMS (MicroElectroMechanical System) fabrication techniques. The microsprayer demonstrated dense and consistent microsprays with average droplet size between 6 and 9 mu m, which fulfilled the droplet size requirement for TRCEM sample preparation. With droplet density of around 12-18 per grid window (window size 58 x 58 mu m), and a data collectible thin ice region of >50% total wetted area, we collected similar to 800-1000 high quality CCD micrographs in a 6-8 h period of continuous effort. This level of output is comparable to what were routinely achieving using cryo-grids prepared by conventional blotting and manual data collection. In this case, weeks of data collection with the previous device has been shortened to a day or two. And hundreds of microliters of valuable sample consumption can be reduced to only a small fraction.
ER -
LU, Z.H., D. BARNARD, Tanvir SHAIKH, X. MENG, C.A. MANNELLA, A.S. YASSIN, R.K. AGRAWAL, T. WAGENKNECHT and T.M. LU. Gas-assisted annular microsprayer for sample preparation for time-resolved cryo-electron microscopy. \textit{JOURNAL OF MICROMECHANICS AND MICROENGINEERING}. BRISTOL: IOP PUBLISHING LTD, vol.~24, No~11, p.~''nestránkováno'', 9 pp. ISSN~0960-1317. doi:10.1088/0960-1317/24/11/115001. 2014.
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