Anti pdgfr alpha

Frozen murine tissues were fixed in cold acetone and 4% PFA before 8 μm cryosections were cut using a cryostat. Cryosections were then placed onto slides coated with Vectabond. anti-PDGFR-alpha (goat, 1:100, R&D system, MN), anti-Fibronectin (FN1) (rabbit, 1:100, abcam, MA), anti-Collagen VI (COL6) (rabbit, 1:200, abcam, MA), anti-PLAU (rabbit, 1:100, abcam, MA), anti-PROCR (Rabbit, 1:50, Bioss antibodies, MA), anti-BMP7 (Rabbit, 1:100, AVIVA system biology, CA), anti-SEMA3F (Rabbit, 1:50, Bioss antibodies, MA), and anti-PCSK6 (Goat, 1:100, antibodies-online, GA) were the primary antibodies used. Primary and secondary antibodies were diluted in 4% skim milk/1x TBS/0.1% Triton-X114. Each slide was washed twice with 1x TBS and treated with Fluoroshield mounting medium with DAPI (Abcam, ab104139) after incubation with the secondary antibodies. An Olympus FV1000 confocal laser scanning microscope (Olympus, Tokyo, Japan) was used to capture the immunohistochemically stained images and these images were analyzed through Fluoview FV10-ASW 3.1 Viewer software (Olympus, Tokyo, Japan).

Jejunal tissue was analyzed by whole mount and cryostat section staining or GFP fluorescence using confocal microscopy as previously described [25 (link), 26 (link)]. Primary antibodies against the following antigens were used: anti-CACNA1G-C (rabbit, 1:50, SantaCruz, TX), anti-CACNA1G-N (rabbit, 1:50, Alomone Labs, Jerusalem, Israel and goat, 1:50, SantaCruz, TX), anti-PDGFR-alpha for mice (goat, 1:100, R&D system, MN), anti-PDGFR-alpha for humans (goat, 1:50, R&D system, MN), anti-PDGFR-beta (goat, 1:200, R&D system, MN), and anti-ACTA2 (rabbit, 1:200, Abcam, MA). Images were collected using the Fluoview FV10-ASW 3.1 Viewer software (Olympus, Tokyo, Japan) with an Olympus FV1000 confocal laser scanning microscope. Cryostat sections were also stained with β-galactosidase using LacZ Tissue Staining Kit (InvivoGen, San Diego, CA).