Fig. 2.

Holder design for OTLS imaging. a Optical simulations of the system’s point spread function (PSF) and Strehl ratio (S), and ray-tracing spot diagrams, are shown for scenarios in which the optical path difference (Δn × t) is negligible, small, or large, which results in diffraction-limited (S ~ 1.0), near-diffraction-limited (S > 0.8), or aberrated (S < 0.8) imaging performance, respectively (scale bars: 1 μm). The dependence of the Strehl Ratio, S, as a function of Δn × t is plotted, indicating that for diffraction-limited imaging, the condition that Δn × t < 0.002 mm should be maintained. Based on this condition, potential glass and monomer/polymer holder materials are shown in (b). The color scale indicates the maximum material thickness, t_max, that is allowed based upon the intrinsic mismatch, Δn, of those materials with published clearing protocols. Chemically incompatible combinations of materials and chemical reagents are also indicated