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  5. Real-time pH imaging of macrophage lysosomes using the pH-sensitive probe ApHID.

Real-time pH imaging of macrophage lysosomes using the pH-sensitive probe ApHID.

File(s)
41092905.pdf (10.72 MB)
Permanent Link(s)
https://hdl.handle.net/1813/118149
Collections
Department of Biochemistry
Author
Sole-Domenech, S.
Singh, P.K.
Funes, L.
Ma, C.-I.J.
Warren, J.D.
Maxfield, F.R.
Abstract

Active endolysosomal pH regulation is essential for optimal enzymatic activity. To measure acidification, pH sensors can be delivered to acidic compartments using labeled dextran polymers or proteins. However, commercial probes have limited sensitivity in the acidic endolysosomal range or their fluorescence undergoes degradation. Herein, we introduce the new pH-sensitive probe ApHID, a green-emitting sensor with optimal dynamic range matching the acidity of endosomes and lysosomes. Acid pH indicator dye (ApHID) has a pKa near 5, increasing brightness with acidity, and withstands oxidation and photobleaching. We used ApHID dextrans to measure endolysosomal pH in macrophages and compared it to other commercially available sensors. ApHID reported pH accurately and stably over time in cell culture and was sensitive to subtle variations in organelle acidification in real time. Overall, ApHID circumvents limitations of currently available commercial probes and can provide utility in demanding applications such as intravital imaging of tissues.

Journal / Series
Cell reports methods
Volume & Issue
5(10)
Date Issued
2025-10-14
Publisher
Cell Press
Keywords
WCM Library Coordinated Deposit
•
Lysosomes/metabolism
•
Hydrogen-Ion Concentration
•
Animals
•
Macrophages/metabolism
•
Fluorescent Dyes/chemistry
•
Endosomes/metabolism
•
Mice
•
Dextrans/chemistry
•
ApHID
•
dextrans
•
endolysosomal pH
•
fluorescence probes
•
pH sensors
•
ratiometric imaging
Related DOI
https://doi.org/10.1016/j.crmeth.2025.101203
Previously Published as
Sole-Domenech S, Singh PK, Funes L, Ma C-IJ, Warren JD, Maxfield FR. Real-time pH imaging of macrophage lysosomes using the pH-sensitive probe ApHID. Cell reports methods. 2025;5(10):101203. doi: 10.1016/j.crmeth.2025.101203. PMID: 41092905.
Rights
Attribution 4.0 International
Rights URI
https://creativecommons.org/licenses/by/4.0/
Type
article

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