Higher-Order Nonsaturated Hyperlinear Upconversion Image Scanning Microscopy for Super-Resolution Imaging.

Area of Science:

• Microscopy
• Nanotechnology
• Biophysics

Background:

• Confocal laser scanning microscopy (CLSM) faces resolution-quality trade-offs.
• Image scanning microscopy (ISM) offers improvements but has limited resolution gains.
• Existing super-resolution methods often involve complex procedures or photobleaching.

Purpose of the Study:

• To develop a novel super-resolution microscopy technique using upconversion nanoparticles.
• To overcome the resolution limitations of conventional and ISM techniques.
• To achieve enhanced spatial resolution beyond the diffraction limit.

Main Methods:

• Developed nonsaturated hyperlinear upconversion image scanning microscopy (UISM).
• Employed lanthanide-doped upconversion nanoparticles (UCNPs) with tunable excitation power.
• Utilized near-infrared excitation at 980 nm for image acquisition.

Main Results:

• Achieved spatial resolutions of 133 nm (455 nm emission) and 196 nm (800 nm emission).
• Demonstrated resolution enhancement significantly beyond twice that of CLSM.
• Obtained a shrunken point spread function by tuning UCNP response at the nonsaturated excitation threshold.

Conclusions:

• Nonsaturated hyperlinear UISM offers a novel strategy for super-resolution imaging.
• This technique enables resolution far beyond conventional laser scanning microscopy.
• The approach provides a new pathway for advanced microscopy with enhanced spatial resolution.