Multi-focal structured illumination microscopy systems and methods

A microscopy system comprising:

• a light source for transmitting a single light beam;
• a beam splitter for splitting the single light beam into a plurality of light beams forming a plurality of multi-focal patterns, wherein each of the multi-focal patterns defines a plurality of focal points;
• a scanner for scanning the plurality of light beams that forms each of the multi-focal patterns onto a sample such that the sample generates a plurality of multi-focal fluorescent emissions in each of the multi-focal patterns that defines a plurality of fluorescence focal points;
• a detector for collecting the plurality of multi-focal fluorescent emissions that results from each of the multi-focal patterns;
• and
• a processing system for processing the plurality of multi-focal fluorescent emissions that results from each of the multi-focal patterns collected from the detector comprising:
• a processor in operative communication with a database for storing the plurality of collected multi-focal fluorescent emissions resulting from the multi-focal patterns, wherein the processor removes out-of-focus fluorescent emissions for each of the plurality of fluorescent emission to leave only in-focus fluorescent emissions resulting from each of the plurality of multi-focal patterns, wherein the processor then scales the in-focus fluorescence emissions resulting from each of the plurality of multi-focal patterns in a local contraction operation in which each of the plurality of fluorescent focal points resulting from each of the multi-focal patterns maintains the same distance from another plurality of fluorescent focal points resulting from the multi-focal pattern as the plurality of fluorescent focal points contract to produce scaled, in-focus fluorescent emissions;
• wherein the processor sums each scaled, in-focus fluorescence emissions to produce a composite image.