Fluorescent Biosensors for Cryogenic Correlative Light and Electron Microscopy
- 1Stanford University
Abstract
Cryogenic electron tomography provides nanometer-scale views of subcellular structures but lacks essential physiological context, such as the metabolic state or ion concentration. To bridge this gap, we incorporated fluorescent biosensors into a cryogenic correlative light and electron microscopy workflow, enabling direct measurement of physiological conditions alongside structural data. Here, we focused on establishing and calibrating biosensors for calcium, pH, and osmotic stress at cryogenic conditions through an in vitro workflow in which biosensors were frozen in defined chemical environments and then imaged ratiometrically. We then applied these biosensors across a range of sample types: an osmotic stress sensor in Saccharomyces cerevisiae, a calcium sensor in human U2OS cells, and a pH sensor in the bacterium Halothiobacillus neapolitanus. This approach links localized physiological changes to the 3D structures of macromolecules and their organization in situ.
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BibTeX
@misc{azaldegui2026fluorescent,
title = {{Fluorescent Biosensors for Cryogenic Correlative Light and Electron Microscopy}},
author = {C. Azaldegui and K. M. Rose and D. Perez and J. A. Summers and C. Mak and W. P. Dwyer and A. G. Vecchiarelli and J. S. Biteen and J. R. Dinneny and P. Dahlberg},
howpublished = {bioRxiv},
year = {2026},
month = oct,
doi = {10.64898/2026.10.15.759416},
url = {https://www.biorxiv.org/content/10.64898/2026.10.15.759416v1},
note = {CC BY},
}
RIS
TY - UNPB
TI - Fluorescent Biosensors for Cryogenic Correlative Light and Electron Microscopy
AU - C. Azaldegui
AU - K. M. Rose
AU - D. Perez
AU - J. A. Summers
AU - C. Mak
AU - W. P. Dwyer
AU - A. G. Vecchiarelli
AU - J. S. Biteen
AU - J. R. Dinneny
AU - P. Dahlberg
PY - 2026
DA - 2026/10/15
PB - bioRxiv
DO - 10.64898/2026.10.15.759416
UR - https://www.biorxiv.org/content/10.64898/2026.10.15.759416v1
AB - Cryogenic electron tomography provides nanometer-scale views of subcellular structures but lacks essential physiological context, such as the metabolic state or ion concentration. To bridge this gap, we incorporated fluorescent biosensors into a cryogenic correlative light and electron microscopy workflow, enabling direct measurement of physiological conditions alongside structural data. Here, we focused on establishing and calibrating biosensors for calcium, pH, and osmotic stress at cryogenic conditions through an in vitro workflow in which biosensors were frozen in defined chemical environments and then imaged ratiometrically. We then applied these biosensors across a range of sample types: an osmotic stress sensor in Saccharomyces cerevisiae, a calcium sensor in human U2OS cells, and a pH sensor in the bacterium Halothiobacillus neapolitanus. This approach links localized physiological changes to the 3D structures of macromolecules and their organization in situ.
ER -