Goat anti myoc antibody

For all Western blot-related experiments on secreted MYOC, 2% FBS containing media was used to minimize gel warping and epitope masking due to excessive FBS/BSA. Conditioned media (20 μL) was denatured in either reducing or nonreducing Laemmli buffer at 95°C for 5 minutes and loaded onto a Tris-Gly SDS-PAGE gel. Proteins were separated and transferred onto a nitrocellulose membrane using a Pierce G2 Fast Blotter (Thermo Scientific). Membranes then were blocked in Odyssey blocking buffer (LI-COR, Lincoln, NE, USA), and probed with either a goat anti-MYOC antibody (1:1000; Santa Cruz Biotechnology, Santa Cruz, CA, USA), a rabbit anti-GLuc antibody (1:2000; NEB), a mouse anti-FLAG M2 antibody (1:2000, Sigma-Aldrich Corp.), a rabbit anti-GRP94 (1:1500, GeneTex, Irvine, CA, USA), or a mouse anti-β-actin (1:10,000, Sigma-Aldrich Corp.) primary antibody followed by an appropriate near-infrared secondary antibody (1:10,000; LI-COR). A LI-COR Odyssey Fc infrared imager (LI-COR) was used to image the blots, and the Image Quant software was used to quantify protein bands.