Fig. 2.

Optical evaluation of the axial scanning method. (a) Side-view of the z-variations in FOV size visualized by line scans in a glass cuvette containing a Fluorescein solution. (b) Relative change of the field-of-view (FOV) size with axial focus shift with respect to the FOV size without ETL. The simulated change in FOV size is shown as dashed lines. (c) 2D ray tracing layout of the ETL/OL assembly with the microscope objective (OBJ) attached. The simulation was calculated for optimum filling of the ETL/Objective back aperture (BA). The change in NA with axial focus shift is apparent. (d) Upper panel: PSF measurements with and without the ETL/OL assembly at the zero z-position using a galvanometric scan mirror based two-photon microscope. All values are stated as PSF half-widths in μm. The fluorescent beads used were 500 nm in size (Fluoresbrite, Polysciences Inc.). Lower panel: Simulated Strehl ratios of the entire excitation path (with underfilled back aperture) at 850 nm as a function of distance to the optical axis and axial focus shift. Up to a distance of 200 μm from the optical axis, diffraction-limited performance can be maintained (Strehl ratio > 0.8).