Impact of Docking Strand Design on Spatial Resolution in DNA-Points Accumulation for Imaging in Nanoscale Topography

Area of Science:

• Biophysics
• Microscopy
• Molecular Biology

Background:

• DNA points accumulation for imaging in nanoscale topography (DNA-PAINT) is a key single-molecule localization microscopy (SMLM) technique.
• Recent repetitive docking strand designs aim to improve imaging speed and multiplexing.
• The impact of these repetitive designs on spatial resolution is not well understood.

Purpose of the Study:

• To systematically quantify the effect of repetitive docking strands on localization distributions and effective resolution in DNA-PAINT.
• To investigate how docking strand architecture influences spatial fidelity in SMLM.
• To provide a framework for optimizing DNA-PAINT for high-precision imaging.

Main Methods:

• Utilized the trimeric proliferating cell nuclear antigen (PCNA) complex as a model system.
• Compared localization precision and distributions between single-motif and repetitive docking strands.
• Analyzed spatial blurring factors including binding site geometry and DNA flexibility.

Main Results:

• Repetitive docking motifs resulted in broadened localization distributions compared to single-motif strands.
• Effective spatial resolution was reduced with repetitive docking strands, despite similar localization precision.
• Factors contributing to spatial blurring include variable binding geometry, rotational flexibility, and imager strand dynamics.

Conclusions:

• Docking strand architecture significantly impacts resolution limits in DNA-PAINT.
• A trade-off exists between multiplexing/speed and structural fidelity in repetitive DNA-PAINT designs.
• Rational design of docking strands is crucial for achieving high-precision imaging of molecular complexes.