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Updated: Jul 6, 2025

Conducting Multiple Imaging Modes with One Fluorescence Microscope

Conducting Multiple Imaging Modes with One Fluorescence Microscope

Published on: October 28, 2018

Bond-selective fluorescence imaging with single-molecule sensitivity.

Haomin Wang¹, Dongkwan Lee¹, Yulu Cao¹

¹Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California 91125, USA.

Nature Photonics

|January 1, 2024

Summary

This summary is machine-generated.

We developed Bond Selective Fluorescence-Detected Infrared-Excited (BonFIRE) spectral microscopy for sensitive bioimaging. This technique provides bond-specific chemical information in cells and tissues with single-molecule sensitivity.

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Area of Science:

Optical imaging
Vibrational spectroscopy
Molecular biology

Background:

Bioimaging requires high chemical specificity and sensitivity.
Mid-infrared (mid-IR) vibrational spectroscopy offers rich chemical information but lacks sensitivity for bioimaging.
Existing methods struggle to provide detailed molecular insights in complex biological systems.

Purpose of the Study:

To develop a highly sensitive bioimaging technique for bond-selective chemical analysis.
To overcome the sensitivity limitations of mid-IR vibrational spectroscopy for biological applications.
To enable detailed molecular interrogation within cells, neurons, and tissues.

Main Methods:

Developed Bond Selective Fluorescence-Detected Infrared-Excited (BonFIRE) spectral microscopy.
Utilized two-photon excitation in mid-IR and near-IR to upconvert vibrational excitations.
Employed tuneable narrowband picosecond pulses for high sensitivity and biocompatibility.
Demonstrated spectral imaging in fingerprint and cell-silent windows with single-molecule sensitivity.

Main Results:

Achieved single-molecule sensitivity for fluorescent dyes using BonFIRE.
Successfully imaged intracellular targets in fixed and live cells, neurons, and tissues.
Demonstrated time-lapse BonFIRE microscopy of live HeLa cells using high-frequency modulation.
Showcased the potential for vibrational multiplexing for advanced analysis.

Conclusions:

BonFIRE spectral microscopy significantly enhances sensitivity for vibrational bioimaging.
The technique provides unprecedented bond-specific molecular information.
BonFIRE expands the bioimaging toolbox for functional imaging and sensing in biological investigations.