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Nondestructive nanostraw intracellular sampling for longitudinal cell monitoring

Cao, Yuhong ; Hjort, Martin LU orcid ; Chen, Haodong ; Birey, Fikri ; Leal-Ortiz, Sergio A. ; Han, Crystal M. ; Santiago, Juan G. ; Paşca, Sergiu P. ; Wu, Joseph C. and Melosh, Nicholas A. (2017) In Proceedings of the National Academy of Sciences of the United States of America 114(10). p.1866-1874
Abstract

Here, we report a method for time-resolved, longitudinal extraction and quantitative measurement of intracellular proteins and mRNA from a variety of cell types. Cytosolic contents were repeatedly sampled from the same cell or population of cells for more than 5 d through a cell-culture substrate, incorporating hollow 150-nm-diameter nanostraws (NS) within a defined sampling region. Once extracted, the cellular contents were analyzed with conventional methods, including fluorescence, enzymatic assays (ELISA), and quantitative real-time PCR. This process was nondestructive with >95% cell viability after sampling, enabling long-term analysis. It is important to note that the measured quantities from the cell extract were found to... (More)

Here, we report a method for time-resolved, longitudinal extraction and quantitative measurement of intracellular proteins and mRNA from a variety of cell types. Cytosolic contents were repeatedly sampled from the same cell or population of cells for more than 5 d through a cell-culture substrate, incorporating hollow 150-nm-diameter nanostraws (NS) within a defined sampling region. Once extracted, the cellular contents were analyzed with conventional methods, including fluorescence, enzymatic assays (ELISA), and quantitative real-time PCR. This process was nondestructive with >95% cell viability after sampling, enabling long-term analysis. It is important to note that the measured quantities from the cell extract were found to constitute a statistically significant representation of the actual contents within the cells. Of 48 mRNA sequences analyzed from a population of cardiomyocytes derived from human induced pluripotent stem cells (hiPSC-CMs), 41 were accurately quantified. The NS platform samples from a select subpopulation of cells within a larger culture, allowing native cell-to-cell contact and communication even during vigorous activity such as cardiomyocyte beating. This platform was applied both to cell lines and to primary cells, including CHO cells, hiPSC-CMs, and human astrocytes derived in 3D cortical spheroids. By tracking the same cell or group of cells over time, this method offers an avenue to understand dynamic cell behavior, including processes such as induced pluripotency and differentiation.

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author
; ; ; ; ; ; ; ; and
publishing date
type
Contribution to journal
publication status
published
keywords
Cellular biology, Molecular biology, Nanotechnology, Sampling
in
Proceedings of the National Academy of Sciences of the United States of America
volume
114
issue
10
pages
1866 - 1874
publisher
National Academy of Sciences
external identifiers
  • scopus:85014600106
  • pmid:28223521
ISSN
0027-8424
DOI
10.1073/pnas.1615375114
language
English
LU publication?
no
additional info
Publisher Copyright: © 2017, National Academy of Sciences. All rights reserved.
id
27491fd9-8680-47f5-860a-aa2424ea92cf
date added to LUP
2021-10-16 21:42:56
date last changed
2021-10-19 02:34:20
@article{27491fd9-8680-47f5-860a-aa2424ea92cf,
  abstract     = {<p>Here, we report a method for time-resolved, longitudinal extraction and quantitative measurement of intracellular proteins and mRNA from a variety of cell types. Cytosolic contents were repeatedly sampled from the same cell or population of cells for more than 5 d through a cell-culture substrate, incorporating hollow 150-nm-diameter nanostraws (NS) within a defined sampling region. Once extracted, the cellular contents were analyzed with conventional methods, including fluorescence, enzymatic assays (ELISA), and quantitative real-time PCR. This process was nondestructive with &gt;95% cell viability after sampling, enabling long-term analysis. It is important to note that the measured quantities from the cell extract were found to constitute a statistically significant representation of the actual contents within the cells. Of 48 mRNA sequences analyzed from a population of cardiomyocytes derived from human induced pluripotent stem cells (hiPSC-CMs), 41 were accurately quantified. The NS platform samples from a select subpopulation of cells within a larger culture, allowing native cell-to-cell contact and communication even during vigorous activity such as cardiomyocyte beating. This platform was applied both to cell lines and to primary cells, including CHO cells, hiPSC-CMs, and human astrocytes derived in 3D cortical spheroids. By tracking the same cell or group of cells over time, this method offers an avenue to understand dynamic cell behavior, including processes such as induced pluripotency and differentiation.</p>},
  author       = {Cao, Yuhong and Hjort, Martin and Chen, Haodong and Birey, Fikri and Leal-Ortiz, Sergio A. and Han, Crystal M. and Santiago, Juan G. and Paşca, Sergiu P. and Wu, Joseph C. and Melosh, Nicholas A.},
  issn         = {0027-8424},
  language     = {eng},
  number       = {10},
  pages        = {1866--1874},
  publisher    = {National Academy of Sciences},
  series       = {Proceedings of the National Academy of Sciences of the United States of America},
  title        = {Nondestructive nanostraw intracellular sampling for longitudinal cell monitoring},
  url          = {http://dx.doi.org/10.1073/pnas.1615375114},
  doi          = {10.1073/pnas.1615375114},
  volume       = {114},
  year         = {2017},
}