Correlated confocal and super-resolution imaging by VividSTORM

Area of Science:

• Life Sciences
• Microscopy
• Cell Biology

Background:

• Single-molecule localization microscopy (SMLM) offers nanoscale precision for visualizing molecular distribution.
• Analyzing SMLM data within the cellular context of tissue preparations remains challenging.

Purpose of the Study:

• To develop an optimized protocol for tissue processing and immunostaining for SMLM.
• To enable correlated confocal and super-resolution imaging of nanoscale protein localization in fixed tissues.
• To introduce open-source software for cell-specific SMLM image analysis.

Main Methods:

• A 5-day optimized tissue processing and immunostaining procedure for SMLM.
• Correlated confocal and 3D-superresolution imaging of fixed mouse brain, heart, and kidney tissues.
• Utilized VividSTORM open-source software for image analysis.

Main Results:

• Demonstrated successful application to fixed mouse brain, heart, and kidney tissues.
• Enabled visualization of nanoscale protein localization within subcellular compartments in minutes.
• Facilitated quantitative analysis of molecular abundance, clustering, and distances in a cell-specific manner.

Conclusions:

• The developed protocol simplifies SMLM data acquisition and analysis in tissue samples.
• Correlated imaging and VividSTORM software allow detailed subcellular and cell-specific molecular studies.
• The protocol requires only basic microscopy and tissue staining skills.