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Super-resolution microscopy recovers spatial detail beyond the conventional diffraction limit by acquiring information beyond a single ordinary image. Stimulated emission depletion microscopy confines detectable fluorescence with a depletion beam; photoactivated localization microscopy separates emitters in time and estimates each emitter's position before combining the estimates.

The distinction between finding one isolated emitter and resolving many adjacent emitters matters. An isolated point spread function can be localized more precisely than its width if enough photons are collected, but overlapping unknown emitters cannot simply be treated as isolated ones. Label density, photon shot noise and specimen motion therefore constrain the final optical resolution, even when the reconstructed image uses very small pixels.