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. Author manuscript; available in PMC: 2020 Nov 4.
Published in final edited form as: Nat Methods. 2020 May 4;17(5):531–540. doi: 10.1038/s41592-020-0816-x

Fig. 1. Concept of INSPR.

Fig. 1.

After the single-molecule dataset (left panel) is acquired, a PSF library is obtained. Starting with a constant pupil function, INSPR assigns each detected PSF to a temporary axial position according to its similarity with the template (assignment step, center panel). These axially-assigned PSFs are subsequently grouped, aligned, and averaged to form a 3D PSF stack, which is then used to provide a new pupil estimation through phase retrieval (update step, center panel). The new pupil is used in the next assignment step to generate an updated template. This process iterates until the retrieved 3D PSF model no longer changes, and a final model is obtained (right panel).