NDLI: Effect of acidic and basic pH on Thioflavin T absorbance and fluorescence

Content Provider	Springer Nature Link
Author	Hackl, Ellen V. Darkwah, Joseph Smith, Geoff Ermolina, Irina
Copyright Year	2015
Abstract	Thioflavin T (ThT) is a fluorescent dye able to enhance significantly its fluorescence quantum yield upon binding to protein amyloids. ThT assay is widely used to detect and quantify amyloids in a variety of conditions, including solutions with different pH levels. In the present work, the effect of acidic and basic pH on the conformation of the ThT molecule and its absorption and fluorescence properties was studied. The results show that both acidic and basic pH decrease significantly the intensity of ThT absorption in the visible region and fluorescence emission intensity. Low pHs induce an immediate “all-or-nothing” decrease in the ThT signal, while in alkaline solutions the ThT signal decreases gradually over time. pH-induced signal quenching is less in the presence of glycerol or protein aggregates. Two different mechanisms are responsible for the ThT signal quenching—the ThT hydroxylation at basic pH and protonation of the nitrogen atom of the dimethylamino group at acidic pH. ThT assays should be carefully carried out at basic or acidic pH as strong pH dependence of ThT could be responsible for misinterpretation and false positive/negative experimental results. The potential unsuitability of ThT as a probe in solutions with high pH (>9) has been shown.
Starting Page	249
Ending Page	261
Page Count	13
File Format	PDF
ISSN	01757571
Journal	European Biophysics Journal
Volume Number	44
Issue Number	4
e-ISSN	14321017
Language	English
Publisher	Springer Berlin Heidelberg
Publisher Date	2015-03-22
Publisher Place	Berlin, Heidelberg
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	ThT pH Absorbance Fluorescence NMR Protein aggregation Biochemistry Biophysics and Biological Physics Cell Biology Neurobiology Membrane Biology Nanotechnology
Content Type	Text
Resource Type	Article
Subject	Medicine Biophysics

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