Journal: The Scientific World Journal

Article Title: Knockdown of Zebrafish Blood Vessel Epicardial Substance Results in Incomplete Retinal Lamination

doi: 10.1155/2014/803718

Figure Lengend Snippet: Disruption of zBves affected the dysregulated patterns in the retina. The frozen sections and immunofluorescence staining with different marker antibodies—zn8 for ganglion cells (a), (e), anti-CAII for Müller glia (b), (f), anti-TH for interplexiform neurons (c), (g), and zpr-1 for photoreceptors (d), (h) were performed in eyes of 96 hpf wild-type and zBves MO-injected embryos. Immunofluorescence staining with Crx (i), (j) antibody in the frozen sections was performed in 72 hpf embryos. 37.5% (6/16) of mutant retina showed Crx protein. The ultrastructures of the retina in wild-type (m), (m′) and zBves MO-injected embryos (n), (n′) were analyzed by TEM. Double staining with CtBP and zpr-1 was adopted in frozen crytosections (k), (l). 33.3% (4/12) of mutant retina showed CtBP protein without zpr-1 expression (l). Asterisks indicate the outer segment. Black arrows indicate the inner segment. N, nucleus; S, Scale bar in immunostaining photographs, 50 μ m. Scale bar in TEM picture, 1000 nm.

Article Snippet: The embryos were incubated with the following primary antibodies in blocking buffer at 4°C overnight: affinity purified anti-zBves (1 : 500) [ ], anti-ZO-1 (1 : 200, Zymed, S. San Francisco, CA), anti-PKC ζ (1 : 200, Santa Cruz Biotechnology, Santa Cruz, CA), zn8 (1 : 50, Zebrafish International Resource Center, Eugene, OR), anticarbonic anhydrase II (CA; 1 : 100, Abcam, Cambridge, England), antityrosine hydroxylase (TH; 1 : 100, Millipore, Temecula, CA), anti-CtBP (1 : 200, Santa Cruz Biotechnology, Santa Cruz, CA), anti-Crx (1 : 200, Santa Cruz Biotechnology, Santa Cruz, CA), and zpr-1 (1 : 400, Zebrafish International Resource Center, Eugene, OR).

Techniques: Immunofluorescence, Staining, Marker, Injection, Mutagenesis, Double Staining, Expressing, Immunostaining