Label-free mid-infrared photothermal microscopy revisits intracellular thermal dynamics: what do fluorescent nanothermometers measure?

[2406.16265] Label-free mid-infrared photothermal microscopy revisits intracellular thermal dynamics: what do fluorescent nanothermometers measure? Skip to main content Search Submit Donate Log in Search arXiv Press Enter to search · Advanced search Physics > Biological Physics arXiv:2406.16265 (physics) [Submitted on 24 Jun 2024 ( v1 ), last revised 12 Apr 2026 (this version, v4)] Title: Label-free mid-infrared photothermal microscopy revisits intracellular thermal dynamics: what do fluorescent nanothermometers measure? Authors: Keiichiro Toda , Masaharu Takarada , Genki Ishigane , Hiroyuki Shimada , Venkata Ramaiah Badarla , Kohki Okabe , Takuro Ideguchi View a PDF of the paper titled Label-free mid-infrared photothermal microscopy revisits intracellular thermal dynamics: what do fluorescent nanothermometers measure?, by Keiichiro Toda and 6 other authors View PDF Abstract: Fluorescent nanothermometry has revealed pronounced intracellular temperature heterogeneity, establishing the field of single-cell thermal biology. However, these observations have sparked a controversy known as the "10^5 gap issue", because heat conduction calculations in aqueous environments predict that such large temperature distributions cannot be sustained within cells. Here, we address this issue using label-free mid-infrared photothermal microscopy. This technique quantifies heat-induced temperature changes under local thermal equilibrium (LTE), in accordance with the conventional thermodynamic and statistical-mechanical definition of temperature, by detecting refractive index variations. From transient thermal decay measurements, we determined that intracellular thermal diffusivity corresponds to 93-94% that of water. This result indicates that intracellular heat conduction is essentially water-like and rules out the hypothesis that anomalously slow intracellular heat conduction underlies the 10^5 gap discrepancy. We then directly compared fluorescent nanothermometry with our label-free thermometry. Under seconds-long heating, the label-free method exhibited a rapid temperature response consistent with water-like heat conduction. In contrast, fluorescent nanothermometers showed not only a similarly fast response but also an additional slow variation that was absent in the label-free readout. This slow component cannot be explained by temperature changes defined under LTE, but instead likely reflects slower intracellular processes not governed by conductive heat transfer. These results suggest that the "10^5 gap issue" stems from comparing two fundamentally distinct physical quantities: the LTE-defined temperature and a slowly-varying, long-lived non-conductive signal detected by fluorescent nanothermometers. Subjects: Biological Physics (physics.bio-ph) ; Optics (physics.optics) Cite as: arXiv:2406.16265 [physics.bio-ph] (or arXiv:2406.16265v4 [physics.bio-ph] for this version) https://doi.org/10.48550/arXiv.2406.16265 Focus to learn more arXiv-issued DOI via DataCite Submission history From: Takuro Ideguchi [ view email ] [v1] Mon, 24 Jun 2024 02:04:37 UTC (1,944 KB) [v2] Sun, 16 Mar 2025 08:25:16 UTC (2,560 KB) [v3] Mon, 1 Sep 2025 08:23:15 UTC (2,191 KB) [v4] Sun, 12 Apr 2026 01:51:51 UTC (4,160 KB) Full-text links: Access Paper: View a PDF of the paper titled Label-free mid-infrared photothermal microscopy revisits intracellular thermal dynamics: what do fluorescent nanothermometers measure?, by Keiichiro Toda and 6 other authors View PDF view license Current browse context: physics.bio-ph < prev | next > new | recent | 2024-06 Change to browse by: physics physics.optics References & Citations NASA ADS Google Scholar Semantic Scholar export BibTeX citation Loading... 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