High-Throughput Single-Cell Manipulation in Brain Tissue
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Steinmeyer-2012-High-throughput sing.pdf
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1.8 MB
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Author(s)
Steinmeyer, Joseph Daly
Yanik, Mehmet Fatih
Date Issued
April 2012
Journal
PLoS ONE
Publisher
Public Library of Science
Citation
Steinmeyer, Joseph D., and Mehmet Fatih Yanik. “High-Throughput Single-Cell Manipulation in Brain Tissue.” Ed. Michele Giugliano. PLoS ONE 7.4 (2012): e35603.
Version
Final published version
Abstract
The complexity of neurons and neuronal circuits in brain tissue requires the genetic manipulation, labeling, and tracking of single cells. However, current methods for manipulating cells in brain tissue are limited to either bulk techniques, lacking single-cell accuracy, or manual methods that provide single-cell accuracy but at significantly lower throughputs and repeatability. Here, we demonstrate high-throughput, efficient, reliable, and combinatorial delivery of multiple genetic vectors and reagents into targeted cells within the same tissue sample with single-cell accuracy. Our system automatically loads nanoliter-scale volumes of reagents into a micropipette from multiwell plates, targets and transfects single cells in brain tissues using a robust electroporation technique, and finally preps the micropipette by automated cleaning for repeating the transfection cycle. We demonstrate multi-colored labeling of adjacent cells, both in organotypic and acute slices, and transfection of plasmids encoding different protein isoforms into neurons within the same brain tissue for analysis of their effects on linear dendritic spine density. Our platform could also be used to rapidly deliver, both ex vivo and in vivo, a variety of genetic vectors, including optogenetic and cell-type specific agents, as well as fast-acting reagents such as labeling dyes, calcium sensors, and voltage sensors to manipulate and track neuronal circuit activity at single-cell resolution.
MIT Department
Massachusetts Institute of Technology. Department of Electrical Engineering and Computer Science
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Creative Commons Attribution
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DOI of Published Version
https://doi.org/10.1371/journal.pone.0035603
