Cargando…
Fluorescent nanosensors for intracellular measurements: synthesis, characterization, calibration, and measurement
Measurement of intracellular acidification is important for understanding fundamental biological pathways as well as developing effective therapeutic strategies. Fluorescent pH nanosensors are an enabling technology for real-time monitoring of intracellular acidification. The physicochemical charact...
Autores principales: | , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Frontiers Media S.A.
2014
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3893563/ https://www.ncbi.nlm.nih.gov/pubmed/24474936 http://dx.doi.org/10.3389/fphys.2013.00401 |
_version_ | 1782299717404721152 |
---|---|
author | Desai, Arpan S. Chauhan, Veeren M. Johnston, Angus P. R. Esler, Tim Aylott, Jonathan W. |
author_facet | Desai, Arpan S. Chauhan, Veeren M. Johnston, Angus P. R. Esler, Tim Aylott, Jonathan W. |
author_sort | Desai, Arpan S. |
collection | PubMed |
description | Measurement of intracellular acidification is important for understanding fundamental biological pathways as well as developing effective therapeutic strategies. Fluorescent pH nanosensors are an enabling technology for real-time monitoring of intracellular acidification. The physicochemical characteristics of nanosensors can be engineered to target specific cellular compartments and respond to external stimuli. Therefore, nanosensors represent a versatile approach for probing biological pathways inside cells. The fundamental components of nanosensors comprise a pH-sensitive fluorophore (signal transducer) and a pH-insensitive reference fluorophore (internal standard) immobilized in an inert non-toxic matrix. The inert matrix prevents interference of cellular components with the sensing elements as well as minimizing potentially harmful effects of some fluorophores on cell function. Fluorescent nanosensors are synthesized using standard laboratory equipment and are detectable by non-invasive widely accessible imaging techniques. The outcomes of studies employing this technology are dependent on reliable methodology for performing measurements. In particular, special consideration must be given to conditions for sensor calibration, uptake conditions and parameters for image analysis. We describe procedures for: (1) synthesis and characterization of polyacrylamide and silica based nanosensors, (2) nanosensor calibration and (3) performing measurements using fluorescence microscopy. |
format | Online Article Text |
id | pubmed-3893563 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-38935632014-01-28 Fluorescent nanosensors for intracellular measurements: synthesis, characterization, calibration, and measurement Desai, Arpan S. Chauhan, Veeren M. Johnston, Angus P. R. Esler, Tim Aylott, Jonathan W. Front Physiol Physiology Measurement of intracellular acidification is important for understanding fundamental biological pathways as well as developing effective therapeutic strategies. Fluorescent pH nanosensors are an enabling technology for real-time monitoring of intracellular acidification. The physicochemical characteristics of nanosensors can be engineered to target specific cellular compartments and respond to external stimuli. Therefore, nanosensors represent a versatile approach for probing biological pathways inside cells. The fundamental components of nanosensors comprise a pH-sensitive fluorophore (signal transducer) and a pH-insensitive reference fluorophore (internal standard) immobilized in an inert non-toxic matrix. The inert matrix prevents interference of cellular components with the sensing elements as well as minimizing potentially harmful effects of some fluorophores on cell function. Fluorescent nanosensors are synthesized using standard laboratory equipment and are detectable by non-invasive widely accessible imaging techniques. The outcomes of studies employing this technology are dependent on reliable methodology for performing measurements. In particular, special consideration must be given to conditions for sensor calibration, uptake conditions and parameters for image analysis. We describe procedures for: (1) synthesis and characterization of polyacrylamide and silica based nanosensors, (2) nanosensor calibration and (3) performing measurements using fluorescence microscopy. Frontiers Media S.A. 2014-01-16 /pmc/articles/PMC3893563/ /pubmed/24474936 http://dx.doi.org/10.3389/fphys.2013.00401 Text en Copyright © 2014 Desai, Chauhan, Johnston, Esler and Aylott. http://creativecommons.org/licenses/by/3.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Physiology Desai, Arpan S. Chauhan, Veeren M. Johnston, Angus P. R. Esler, Tim Aylott, Jonathan W. Fluorescent nanosensors for intracellular measurements: synthesis, characterization, calibration, and measurement |
title | Fluorescent nanosensors for intracellular measurements: synthesis, characterization, calibration, and measurement |
title_full | Fluorescent nanosensors for intracellular measurements: synthesis, characterization, calibration, and measurement |
title_fullStr | Fluorescent nanosensors for intracellular measurements: synthesis, characterization, calibration, and measurement |
title_full_unstemmed | Fluorescent nanosensors for intracellular measurements: synthesis, characterization, calibration, and measurement |
title_short | Fluorescent nanosensors for intracellular measurements: synthesis, characterization, calibration, and measurement |
title_sort | fluorescent nanosensors for intracellular measurements: synthesis, characterization, calibration, and measurement |
topic | Physiology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3893563/ https://www.ncbi.nlm.nih.gov/pubmed/24474936 http://dx.doi.org/10.3389/fphys.2013.00401 |
work_keys_str_mv | AT desaiarpans fluorescentnanosensorsforintracellularmeasurementssynthesischaracterizationcalibrationandmeasurement AT chauhanveerenm fluorescentnanosensorsforintracellularmeasurementssynthesischaracterizationcalibrationandmeasurement AT johnstonanguspr fluorescentnanosensorsforintracellularmeasurementssynthesischaracterizationcalibrationandmeasurement AT eslertim fluorescentnanosensorsforintracellularmeasurementssynthesischaracterizationcalibrationandmeasurement AT aylottjonathanw fluorescentnanosensorsforintracellularmeasurementssynthesischaracterizationcalibrationandmeasurement |