Single-Objective Lattice Light Sheet Microscopy with Microfluidics for Single-Molecule Super-Resolution Imaging of...

Area of Science:

• Biophysics
• Optical Microscopy
• Cellular Imaging

Background:

• Single-molecule localization microscopy (SMLM) enables nanoscale imaging beyond the diffraction limit.
• Light sheet microscopy enhances SMLM by improving signal-to-background and reducing photodamage.
• Lattice light sheet (LLS) microscopy offers superior sectioning and imaging depth in scattering samples but often requires dual objectives.

Purpose of the Study:

• To develop and characterize a novel single-objective lattice light sheet (soLLS) imaging platform.
• To overcome the limitations of conventional dual-objective LLS microscopy.
• To demonstrate improved 3D super-resolution imaging in challenging biological samples.

Main Methods:

• Implementation of LLS illumination within a reflective single-objective geometry (soLLS) inside a microfluidic chip.
• Quantitative characterization of soLLS propagation properties in scattering media.
• Integration of soLLS with point spread function engineering for 3D super-resolution imaging.

Main Results:

• The soLLS platform utilizes a single high numerical aperture objective for both illumination and detection.
• soLLS demonstrates superior field of view and sectioning compared to Gaussian light sheets in scattering samples.
• The platform enables improved 3D single-molecule super-resolution imaging of multiple targets across multiple cells.

Conclusions:

• The soLLS imaging platform provides a versatile solution for nanoscale investigations in challenging biological and biomedical samples.
• This technology expands the scope of SMLM applications by mitigating constraints of dual-objective setups.
• The soLLS system facilitates deeper insights into cellular architecture and molecular dynamics at the nanoscale.