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Trapped! A Critical Evaluation of Methods for Measuring Total Cellular Uptake versus Cytosolic Localization


Deprey, Kirsten; Becker, Lukas; Kritzer, Joshua; Plückthun, Andreas (2019). Trapped! A Critical Evaluation of Methods for Measuring Total Cellular Uptake versus Cytosolic Localization. Bioconjugate Chemistry, 30(4):1006-1027.

Abstract

Biomolecules have many properties that make them promising for intracellular therapeutic applications, but delivery remains a key challenge because large biomolecules cannot easily enter the cytosol. Furthermore, quantification of total intracellular versus cytosolic concentrations remains demanding, and the determination of delivery efficiency is thus not straightforward. In this review, we discuss strategies for delivering biomolecules into the cytosol and briefly summarize the mechanisms of uptake for these systems. We then describe commonly used methods to measure total cellular uptake and, more selectively, cytosolic localization, and discuss the major advantages and drawbacks of each method. We critically evaluate methods of measuring "cell penetration" that do not adequately distinguish total cellular uptake and cytosolic localization, which often lead to inaccurate interpretations of a molecule's cytosolic localization. Finally, we summarize the properties and components of each method, including the main caveats of each, to allow for informed decisions about method selection for specific applications. When applied correctly and interpreted carefully, methods for quantifying cytosolic localization offer valuable insight into the bioactivity of biomolecules and potentially the prospects for their eventual development into therapeutics.

Abstract

Biomolecules have many properties that make them promising for intracellular therapeutic applications, but delivery remains a key challenge because large biomolecules cannot easily enter the cytosol. Furthermore, quantification of total intracellular versus cytosolic concentrations remains demanding, and the determination of delivery efficiency is thus not straightforward. In this review, we discuss strategies for delivering biomolecules into the cytosol and briefly summarize the mechanisms of uptake for these systems. We then describe commonly used methods to measure total cellular uptake and, more selectively, cytosolic localization, and discuss the major advantages and drawbacks of each method. We critically evaluate methods of measuring "cell penetration" that do not adequately distinguish total cellular uptake and cytosolic localization, which often lead to inaccurate interpretations of a molecule's cytosolic localization. Finally, we summarize the properties and components of each method, including the main caveats of each, to allow for informed decisions about method selection for specific applications. When applied correctly and interpreted carefully, methods for quantifying cytosolic localization offer valuable insight into the bioactivity of biomolecules and potentially the prospects for their eventual development into therapeutics.

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Additional indexing

Item Type:Journal Article, refereed, further contribution
Communities & Collections:04 Faculty of Medicine > Department of Biochemistry
07 Faculty of Science > Department of Biochemistry
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Scopus Subject Areas:Life Sciences > Biotechnology
Physical Sciences > Bioengineering
Physical Sciences > Biomedical Engineering
Life Sciences > Pharmacology
Life Sciences > Pharmaceutical Science
Physical Sciences > Organic Chemistry
Language:English
Date:17 April 2019
Deposited On:06 Sep 2019 10:15
Last Modified:29 Jul 2020 11:16
Publisher:American Chemical Society (ACS)
ISSN:1043-1802
OA Status:Closed
Publisher DOI:https://doi.org/10.1021/acs.bioconjchem.9b00112
PubMed ID:30882208

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