Sequencing and high resolution imaging

A method of distinguishing a first analyte from a second analyte immobilized on a substrate, said method comprising:

• (a) binding a first probe comprising a first detectable label to said first analyte to generate a first optical signal and binding a second probe comprising a second detectable label to said second analyte to generate a second optical signal, said first analyte and second analyte being immobilized on the substrate at a center-to-center distance that is less than the diffraction limit of light emitted by the first or second detectable label, and said first and second optical signals at least partially overlap;
• (b) detecting said first optical signal and said second optical signal with an optical imaging system;
• and
• (c) processing a first peak intensity of said first optical signal at a first peak location, and a second peak intensity of said second optical signal at a second peak location to distinguish said first analyte from said second analyte separated from the first analyte by said center-to-center distance less than the diffraction limit of light emitted by the first or second detectable label, and to identify at least one of the first analyte or the second analyte.