Zpr-3 labels green cones as well as rods in immunofluorescence assays. (A) No primary antibody incubation was used as a negative control staining. (B) Double-label immunofluorescence images of zpr-3 and anti-Opn1lw antibody on zebrafish retinal sections. The fluorescence signals of zpr-3 were almost not co-localized with the outer segments of red-cones. Scale bar, 20 μm. N = 3 (biological replicates); n = 3 (technical replicates). (C) No primary antibody incubation was used as a negative control staining. (D) Double-label immunofluorescence images of zpr-3 and anti-Opn1mw antibodies on zebrafish retinal sections. Obvious co-localization between the fluorescence signals of zpr-3 and the outer segments of green cones is shown. Scale bar, 20 μm. N = 3 (biological replicates); n = 3 (technical replicates). INL, inner nuclear layer; NP, no primary antibody control; ONL, outer nuclear layer; OS, outer segment. (E) Phylogenetic tree of zpr-3 (320–354 aa in Rho) was reconstructed using the neighbor-joining method (left). Sequence alignment of the epitope of zpr-3 (320–354 aa in Rho) and its corresponding regions in eight opsins of cones (Opn1mw1-4, Opn1lw1-2, Opn1sw1, and Opn1sw2). The identical amino acid residues are marked in red (right).
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