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Analyst. 2017 Oct 7;142(19):3648-3655. doi: 10.1039/c7an00981j. Epub 2017 Aug 31.

Quantification of green fluorescent protein-(GFP-) tagged membrane proteins by capillary gel electrophoresis.

Author information

1
PharmaCenter Bonn, Pharmaceutical Institute, Pharmaceutical Chemistry I, University of Bonn, An der Immenburg 4, D-53121 Bonn, Germany. christa.mueller@uni-bonn.de.

Abstract

A fast and robust procedure for the quantification of GFP-tagged membrane proteins in cell homogenates was developed employing capillary gel electrophoresis coupled to laser-induced fluorescence detection (CGE-LIF). The new method was found to be highly sensitive and applicable to structurally diverse membrane proteins including synaptic vesicle protein 2A (SV2A), adenosine A2A receptor (A2AAR), and connexin 43 (Cx43). Quantification of SV2A and A2AAR using radioligand binding assays confirmed the results obtained with CGE-LIF. The CGE-LIF method showed significantly higher sensitivity as compared to fluorimetric measurement in a microplate. Importantly, CGE-LIF involves separation of the target proteins and their degradation products prior to quantification and thereby ensures specificity. We anticipate broad applicability of the method for any fluorophore-tagged protein.

PMID:
28858361
DOI:
10.1039/c7an00981j
[Indexed for MEDLINE]

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