Total internal reflection and transmission illumination fluorescence microscopy imaging system with improved background suppression

A non-transitory computer-readable medium storing instructions that, when executed by one or more processors, cause the one or more processors to perform operations for imaging a sample, the operations comprising:

• obtaining a plurality of intermediate images of the sample, the plurality of intermediate images of the sample captured during an illumination of the sample using a primary beam of light and an interfering beam of light, the interfering beam of light creating interference with respect to the primary beam of light, the interference including an interference pattern, the plurality of intermediate images including a representation on the interference pattern
• detecting changes between each of the plurality of intermediate images based at least in part on the representation of the interference pattern;
• generating a final image of the sample based on the plurality of intermediate images;
• omitting image components of the plurality of intermediate images that remain static from the final image;
• and
• including other image components of the plurality of intermediate images that change in the final image.

An imaging system configured to image a sample, the imaging system comprising:

• a light source configured to generate a beam of light;
• a beam splitter configured to split the beam of light into a primary beam of light and an interfering beam of light;
• one or more mirrors configured to direct the primary beam of light and the interfering beam of light toward the sample, the interfering beam of light being configured to create interference with respect to the primary beam of light during illumination of the sample, the interference including an interference pattern;
• one or more cameras configured to capture a plurality of intermediate images of the sample during the illumination of the sample, the plurality of intermediate images including a representation of the interference pattern;
• and
• one or more processors configured to generate a final image of the sample based on changes detected between each of the plurality of intermediate images based at least in part on the representation of the interference pattern, image components of the plurality of intermediate images that remain static being omitted from the final image and other image components of the plurality of intermediate images that change being included in the final image.