Infrared microscopy points to cutting-edge biopsies
Photoacoustic image of myelin in 300-µm-thick slice of the cerebrum from a mouse brain [Caltech]
Researchers have used mid-infrared photoacoustic microscopy, with an ultraviolet laser, to image lipids and proteins in cells, and thick brain slices, at high resolution.
The technique holds huge promise for tissue biopsies as well as cancer screening.
Conventional mid-infrared (MIR) microscopy uses a MIR laser to probe tissues without staining but the long laser wavelength limits lateral resolution while broadband absorption of water prohibits clear imaging of fresh samples.
Given this, Professor Lihong Wang and colleagues from the Caltech Optical Imaging Laboratory, California Institute of Technology, developed ultraviolet-localized MIR photoacoustic microscopy (ULM-PAM), to achieve high resolution and high contrast MIR imaging of fresh biological samples.
As part of their set-up, they use a pulsed MIR laser to thermally excite samples at a focal spot, and then a confocal pulsed ultraviolet laser to photoacoustically detect the resulting temperature rise.
In this way, the magnitude of the MIR absorption by the sample can be measured.
Importantly, by comparing the signals from samples before and after heating, images can be created in which structures such as cancerous cells are differentiated by the heat signatures.
“This detection scheme is based on the fact that a temperature rise in a sample enhances photoacoustic signals, a phenomenon called the Grüneisen relaxation effect,” explains Wang in Nature Photonics. “While our imaging method reveals MIR absorption contrast, its lateral resolution is determined by the ultraviolet wavelength, which is one order of magnitude shorter than the MIR wavelength.”
As Wang adds, the ultraviolet light in the range of 200–230 nm is highly absorbed by most biomolecules, such as lipids, proteins and nucleic acids yet is totally transmissive in water.
This means UV light suppresses the the strong water background of MIR absorption.
What's more, the ultraviolet light can penetrate up to 100 µm greater than MIR in fresh specimens while the photoacoustic signal can propagate in biological tissues with negligible scattering.
Wang and colleagues have used ULM-PAM to map the distribution of intracellular lipids and proteins in mouse fibroblast cells and generate images.
They also studied the structural details of the cerebrum and cerebellum in a mouse brain ex vivo, producing images similar to histology or dye-stained fluorescence images with rich structural information about the nerve fibres or fibre bundles.
The technique is currently at the proof-of-concept stage with analyses taking too long for the system to be useful in a clinical setting.
However, Wang reckons the method can be honed to achieve nanoscale far-field chemical imaging by extending the wavelength of the probe beam to the X-ray regime.
"I want to move this to in vivo. I want to use this to image cancer cells during surgery," he says. "That would be the dream."
Research is published in Nature Photonics.