Combining confocal and single molecule localisation microscopy: A correlative approach to multi-scale tissue imaging

Area of Science:

• Biophysics
• Cell Biology
• Microscopy

Background:

• Biological investigations often necessitate data across diverse spatial scales, from molecular to tissue levels.
• Understanding cardiac tissue structure and function requires multi-scale imaging capabilities.

Purpose of the Study:

• To develop and validate a correlative multi-scale imaging method for human cardiac tissue.
• To combine nanoscale resolution (dSTORM) with cellular and tissue-level imaging (confocal microscopy).

Main Methods:

• Correlative imaging of human cardiac tissue using direct stochastic optical reconstruction microscopy (dSTORM) and confocal microscopy.
• Utilizing conventional fluorescence dyes for imaging of junctophilin (JPH), ryanodine receptor (RyR), and wheat germ agglutinin (WGA).
• Segmentation of dSTORM data into membrane and non-membrane components for detailed analysis.

Main Results:

• Successfully imaged macro-molecular membrane complexes of JPH and RyR at the nanoscale in human cardiac tissue.
• Demonstrated increased colocalization of RyR with JPH at the plasma membrane compared to intracellular compartments.
• Provided strategies for antibody labeling, quality control, data alignment, and analysis for multi-scale datasets.

Conclusions:

• The combined dSTORM and confocal microscopy approach enables high-resolution, multi-scale analysis of cardiac tissue architecture.
• This method facilitates detailed investigation of protein distribution and interactions within cardiac cells.
• The described strategies support the application of correlative multi-scale imaging in cardiovascular research.