Effect of Different Fluorescence Iabelling Methods of the Actin Cytoskeleton for Storm Application

Abstract

The Abbe limit was a major challenge in optical microscopy, making it difficult to observe microscopic biological structures. However, the development of STORM, a new fluorescence localization microscopy technique, has enabled much higher resolution imaging by causing dye molecules to blink under laser light. Advances in computational analysis have further improved image processing and resolution, leading to clearer observations and a better understanding of biological structures. Actin, a common protein in eukaryotic cells, forms F-actin, a filament involved in essential processes like cell division, migration, and muscle contraction. Fluorescence labeling is crucial for visualizing F-actin and other cellular structures, allowing scientists to study these important processes. This project explored two methods to visualize actin structures in Hela cells: phalloidin labeling with Alexa-647 dye and DNA-PAINT using GFP. ThunderSTORM analysis produced normalized images for both methods, and resolution values were calculated using FRC. The results indicated that phalloidin labeling provided higher resolution but lower-quality images, while DNA-PAINT with GFP offered better-quality images but with lower resolution.