University of Tokyo scattering microscopy sees dynamic biological processes

19 Nov 2025

Unifying detection of forward and backward scatter combines advantages of both approaches.

A project at the University of Tokyo has developed a bidirectional scattering microscopy technique for imaging complex cell structures.

Discussed in Nature Communications, the platform is designed to resolve both micro- and nanoscale features - a "great unified microscope" according to the team.

The new technique has been termed bidirectional quantitative scattering microscopy (BiQSM), and combines aspects of quantitative phase microscopy (QPM) and interferometric scattering (iSCAT).

QPM involves quantifying the phase shift that occurs when light waves pass through a more optically dense object. The variations in phase and optical path length brought about as light moves through a biological sample can generate high contrast images of delicate internal structures.

iSCAT is an interferometry method that detects elastic Rayleigh scattering in addition to reflected or transmission signals from objects being studied. The technique has been applied to the imaging of proteins and sub-wavelength features in cells, along with their size and mass.

"Modern, cutting-edge techniques have had to straddle tradeoffs," commented the University of Tokyo. "QPM leverages forward-scattered light and can detect structures at the microscale, but not smaller. Consequently, this technique has been primarily used to take static pictures of relatively complex cell structures."

Meanwhile iSCAT microscopy exploits back-scattered light and can detect structures as small as single proteins, or be used to track single particles, allowing insight into dynamical changes within the cell. But it cannot provide the comprehensive views that QPM achieves.

Observing the process of cell death

The Tokyo project aimed to find out whether measuring both forward and backward-scattered light simultaneously could overcome this tradeoff, and reveal a wide range of sizes and motions from the same image. A platform using off-axis digital holography with bidirectional illumination and spatial-frequency multiplexing method was constructed for this purpose.

“Our biggest challenge was cleanly separating two kinds of signals from a single image, while keeping noise low and avoiding mixing between them," commented Keiichiro Toda from the University of Tokyo.

In trials, the researchers set out to observe what happened during cell death. BiQSM captured images of dying cells, detecting the microstructural changes in nuclear shape, cell mass and other parameters suggesting the progress of cellular death. New details of the attenuation of motion for both micro- and nanoscale objects during this process were revealed by the BiQSM platform.

The next steps for the researchers will include extending the dynamic range of the process, currently limited by optical shot noise, and perhaps incorporating chemical contrast through combination with Raman or mid-infrared microscopy.

"We plan to study even smaller particles, such as exosomes and viruses, and to estimate their size and refractive index in different samples," said Keiichiro Toda. "We also want to reveal how living cells move toward death by controlling their state and double-checking our results with other techniques."