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Real-Time Sensing of Single-Ligand Delivery with Nanoaperture-Integrated Microfluidic Devices
[Image: see text] The measurement of biological events on the surface of live cells at the single-molecule level is complicated by several factors including high protein densities that are incompatible with single-molecule imaging, cellular autofluorescence, and protein mobility on the cell surface....
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Chemical Society
2017
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5537690/ https://www.ncbi.nlm.nih.gov/pubmed/28782052 http://dx.doi.org/10.1021/acsomega.7b00934 |
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author | Martin, W. Elliott Ge, Ning Srijanto, Bernadeta R. Furnish, Emily Collier, C. Patrick Trinkle, Christine A. Richards, Christopher I. |
author_facet | Martin, W. Elliott Ge, Ning Srijanto, Bernadeta R. Furnish, Emily Collier, C. Patrick Trinkle, Christine A. Richards, Christopher I. |
author_sort | Martin, W. Elliott |
collection | PubMed |
description | [Image: see text] The measurement of biological events on the surface of live cells at the single-molecule level is complicated by several factors including high protein densities that are incompatible with single-molecule imaging, cellular autofluorescence, and protein mobility on the cell surface. Here, we fabricated a device composed of an array of nanoscale apertures coupled with a microfluidic delivery system to quantify single-ligand interactions with proteins on the cell surface. We cultured live cells directly on the device and isolated individual epidermal growth factor receptors (EGFRs) in the apertures while delivering fluorescently labeled epidermal growth factor. We observed single ligands binding to EGFRs, allowing us to quantify the ligand turnover in real time. These results demonstrate that this nanoaperture-coupled microfluidic device allows for the spatial isolation of individual membrane proteins while maintaining them in their cellular environment, providing the capability to monitor single-ligand binding events while maintaining receptors in their physiological environment. These methods should be applicable to a wide range of membrane proteins. |
format | Online Article Text |
id | pubmed-5537690 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | American Chemical Society |
record_format | MEDLINE/PubMed |
spelling | pubmed-55376902017-08-03 Real-Time Sensing of Single-Ligand Delivery with Nanoaperture-Integrated Microfluidic Devices Martin, W. Elliott Ge, Ning Srijanto, Bernadeta R. Furnish, Emily Collier, C. Patrick Trinkle, Christine A. Richards, Christopher I. ACS Omega [Image: see text] The measurement of biological events on the surface of live cells at the single-molecule level is complicated by several factors including high protein densities that are incompatible with single-molecule imaging, cellular autofluorescence, and protein mobility on the cell surface. Here, we fabricated a device composed of an array of nanoscale apertures coupled with a microfluidic delivery system to quantify single-ligand interactions with proteins on the cell surface. We cultured live cells directly on the device and isolated individual epidermal growth factor receptors (EGFRs) in the apertures while delivering fluorescently labeled epidermal growth factor. We observed single ligands binding to EGFRs, allowing us to quantify the ligand turnover in real time. These results demonstrate that this nanoaperture-coupled microfluidic device allows for the spatial isolation of individual membrane proteins while maintaining them in their cellular environment, providing the capability to monitor single-ligand binding events while maintaining receptors in their physiological environment. These methods should be applicable to a wide range of membrane proteins. American Chemical Society 2017-07-25 /pmc/articles/PMC5537690/ /pubmed/28782052 http://dx.doi.org/10.1021/acsomega.7b00934 Text en Copyright © 2017 American Chemical Society This is an open access article published under an ACS AuthorChoice License (http://pubs.acs.org/page/policy/authorchoice_termsofuse.html) , which permits copying and redistribution of the article or any adaptations for non-commercial purposes. |
spellingShingle | Martin, W. Elliott Ge, Ning Srijanto, Bernadeta R. Furnish, Emily Collier, C. Patrick Trinkle, Christine A. Richards, Christopher I. Real-Time Sensing of Single-Ligand Delivery with Nanoaperture-Integrated Microfluidic Devices |
title | Real-Time Sensing of Single-Ligand Delivery with Nanoaperture-Integrated
Microfluidic Devices |
title_full | Real-Time Sensing of Single-Ligand Delivery with Nanoaperture-Integrated
Microfluidic Devices |
title_fullStr | Real-Time Sensing of Single-Ligand Delivery with Nanoaperture-Integrated
Microfluidic Devices |
title_full_unstemmed | Real-Time Sensing of Single-Ligand Delivery with Nanoaperture-Integrated
Microfluidic Devices |
title_short | Real-Time Sensing of Single-Ligand Delivery with Nanoaperture-Integrated
Microfluidic Devices |
title_sort | real-time sensing of single-ligand delivery with nanoaperture-integrated
microfluidic devices |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5537690/ https://www.ncbi.nlm.nih.gov/pubmed/28782052 http://dx.doi.org/10.1021/acsomega.7b00934 |
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