Knockdown of PRMT5 was achieved by transfecting MCF-7 (human breast adenocarcinoma cell line) cells with specific siRNA (Silencer^® select). Immunofluorescence analysis was performed on MCF-7 cells (untransfected, panel a-d), transfected with PRMT5 specific siRNA (panel i-l) or non-specific scrambled siRNA (panels e-h). Cells were fixed, permeabilized, and labeled with Anti-PRMT5 Recombinant Rabbit Multiclonal Antibody (ab277792, 1/100 dilution), followed by Goat anti-Rabbit IgG (H+L) Superclonal^® Secondary Antibody, Alexa Fluor^® 488 conjugate (1/2000). Nuclei (blue) were stained using ProLong Diamond Antifade Mountant with DAPI, and Rhodamine Phalloidin (1/300) was used for cytoskeletal F-actin (red) staining. Significant reduction of the signal was observed upon siRNA mediated knockdown (panel i-l) confirming the specificity of the antibody to PRMT5 (green). The images were captured at 60X magnification.

Knockdown of PRMT5 was achieved by transfecting MCF-7 cells with specific siRNA (Silencer^® select Product # s20377). Immunofluorescence analysis was performed on MCF-7 cells (untransfected, panel a-d), transfected with PRMT5 specific siRNA (panel i-l) or non-specific scrambled siRNA (panels e-h). Cells were fixed, permeabilized, and labeled with Anti-PRMT5 Recombinant Rabbit Multiclonal Antibody (ab277792, 1/100 dilution), followed by Goat anti-Rabbit IgG (H+L) Superclonal^™ Secondary Antibody, Alexa Fluor^® 488 conjugate (1/2000). Nuclei (blue) were stained using ProLong Diamond Antifade Mountant with DAPI(Product # P36962), and Rhodamine Phalloidin (1/300) was used for cytoskeletal F-actin (red) staining. Significant reduction of the signal was observed upon siRNA mediated knockdown (panel i-l) confirming the specificity of the antibody to PRMT5 (green). The images were captured at 60X magnification.

Enrichment of endogenous PRMT5 protein at specific gene loci using Anti-PRMT5 Antibody: Chromatin Immunoprecipitation (ChIP) was performed using Anti-PRMT5 Recombinant Rabbit Multiclonal Antibody (ab277792, 4 μg) on sheared chromatin from 2 million MCF-7 cells using the MAGnify ChIP System kit. Normal Rabbit IgG was used as a negative IP control. The purified DNA was analyzed by qPCR with PCR primer pairs over Promoters of ADIPOQ, BIRC3, RETN (active) and SAT2 satellite repeats (inactive). Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.