|IHC - Wholemount||1/3000.|
This antibody appears to stain intercellular bridges between germ cells, as it should. TEX14 staining is present within the testis cords between between DDX4 positive cells (germ cells), but is absent between DDX4 negative cells (Sertoli cells).
Postnatal day 6 mouse testes were fixed with 4% paraformaldehyde for 5 hours at 4°C, then washed overnight in PBS at 4°C. Testes were detunicated and cut in half, then permeabilized with 0.1% Triton X-100 in PBS for 30 min at 20°C, then blocked with 3% BSA with 0.1% Triton X-100 in PBS for 1 hour at 20°C. Sections were incubated with Rabbit Anti-Tex14 (Abcam ab41733) diluted 1/3000 plus Goat Anti-DDX4 diluted 1/800 for 14 hours at 4°C. Tissue was then washed 3X for 30 min each with 0.1% Triton X-100 in PBS. Tissue was then incubated with Donkey Anti-Rabbit 555 (Abcam ab150062) plus Donkey Anti-Goat 647 (Abcam ab150155) diluted 1/500 for 3 hours at 20°C. with Abcam DAPI after washing 3X with 0.1% Triton X-100 in PBS. Tissue was then incubated with Fluoroshield Mounting Medium With DAPI (Abcam ab104139) with mounted onto a slide and coverslipped. Tissue was imaged on an Olympus FV 1000 confocal microscope and represented as a maximum intensity Z-projection of a 5 µM thick Z-stack.