High-throughput computational microscopy imaging

Organelles are involved in a variety of life activities
of cells.
Their dysfunction is closely related
to the development and metastasis of cancer.
The exploration of subcellular structures and their abnormal states can lead to a better understanding of pathological mechanisms and provide more effective treatments
for early diagnosis.

The optical microscope, invented more than 400 years ago, has become an indispensable and ubiquitous instrument for the study
of miniature objects in many scientific and technical fields.
In particular, fluorescence microscopy technology has achieved a leap from two-dimensional wide field of view, to three-dimensional confocal to super-resolution fluorescence microscopy, which has greatly promoted the development of
modern life sciences.

With conventional microscopy, researchers currently struggle to generate sufficient intrinsic contrast
for unstained cells due to their low absorption or weak scattering properties.
Specific dyes or fluorescent labels can help visualize, but long-term observation of living cells is still difficult to achieve
.

In recent years, quantitative phase imaging (QPI) has shown promise
with its unique ability to quantify the phase delay of unlabeled samples in a non-destructive manner.
However, the throughput of an imaging platform is fundamentally limited by the spatial bandwidth product (SBP) of its optical system, while the SBP increase of the microscope is fundamentally confounded
by the scale-dependent geometric aberration of its optics.
This leads to a trade-off
between achievable image resolution and field of view (FOV).

To achieve accurate detection and quantification of subcellular features and events, a method
for label-free, high-resolution, and large-field microscopy imaging is required.
To this end, researchers from Nanjing University of Science and Technology (NJUST) and the University of Hong Kong have recently developed a label-free, high-throughput microscopy method
based on mixed light and dark field illumination.
The Hybrid Brightfield-Darkfield Intensity Transport (HBDTI) method for high-throughput quantitative phase microscopy significantly expands the accessible sample space frequencies in Fourier space, extending the maximum achievable resolution by about 5 times the diffraction limit
of coherent imaging, according to Advanced Photonics.

Based on the principle of illumination multiplexing and synthetic aperture, a forward imaging model
of nonlinear brightfield and darkfield intensity transmission was established.
This model gives HBDTI the ability to
provide features that exceed the coherence diffraction limit.
The team demonstrated HBDTI high-throughput imaging using a commercial microscope with a 4x 0.
16NA objective, achieving a half-wide imaging resolution of 488 nm over a field of view of approximately 7.
19 ^mm2, an increase of 25 ×
in SBP with coherent illumination.

Non-invasive high-throughput imaging techniques can delineate subcellular structures
in large-scale cell studies.
Corresponding author Chao Zuo, principal investigator at Northwestern Polytechnic University's Intelligent Computational Imaging Laboratory (SCILab), said, "HBDTI provides a simple, high-performance, low-cost, and versatile quantitative imaging tool
for life science and biomedical research.
Given its high-throughput QPI capabilities, HBDTI promises to provide a powerful solution
for the detection and analysis of subcellular structures in large populations of cells across scales.
Zuo noted that further efforts are needed to facilitate the high-speed implementation
of HBDTI in large populations of live cell analysis.