OS structure in retinal sections were examined by immunostaining with rod marker Rhodopsin (A), s-cone marker Opn1sw (B), m/l-cone marker Opn1mw/lw (C), and helium ion microscopy (HIM) (G). In Top2b cKO, there was a significant reduction of OS thickness in all photoreceptor cells. OS structure was mostly missing or barely identifiable in HIM. Purple arrows indicate the OS, red arrowheads indicate the processes of photoreceptor cells, and the yellow arrows indicate marker expression inside cell bodies. Images in boxed areas are shown in a higher magnification. The thickness of the ONL and OSL of rod cells were measured as indicated by the bar lines and plotted (D and E). The number of s-cone cells was counted and plotted (F). OLM, outer limiting membrane; ONL, outer nuclear layer; INL, inner nuclear layer; GCL, ganglion cell layer; OS, outer segment; OSL, outer segment layer; RPE, retinal pigment epithelium. Scale bars A-C: 50μm/10μm (enlarged); scale bar in G = 5μm. Data in plots are represented as median ± max/min (n ≥ 3). p-values were determined by Student’s T-test.