- MA5-15796 - Invitrogen Antibodies CARM1 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of CARM1 was performed by loading 20 µg from p53-/- Mouse Embryonic Fibroblasts (MEFs) treated with a control shRNA (shNT) or four different shRNA’s (sh#1-sh#4) knocking down CARM1 protein per well onto an SDS-PAGE gel. Proteins were transferred to a nitrocellulose membrane and blocked with 5% milk in PBST for 1 hour. The membrane was probed with a CARM1 monoclonal antibody (Product # MA5-15796) at a dilution of 1:1000 at 4°C overnight, washed in PBST, and probed with an HRP-conjugated anti-mouse IgG secondary antibody at a dilution of 1:1000 for 1 hour at room temperature. As a loading control, the same lysates were probed with an antibody against alpha-Tubulin. Detection was performed using ECL substrate. Data courtesy of the Innovators Program.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of CARM1 using CARM1 monoclonal antibody (Product # MA5-15796) in MCF-7 (1), HeLa (2), NIH/3T3 (3), HL-60 (4), LNcap (5), Jurkat (6), PC-3 (7), COS-7 (8), and PC-12 (9) cell lysate.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of PRMT4 was achieved by transfecting PC-3 cells with PRMT4 specific siRNAs (Silencer® select Product # s20579, s20577). Western blot analysis (Fig. a) was performed using whole cell extracts from the PRMT4 knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with PRMT4 Monoclonal Antibody (3H2) (Product # MA5-15796, 1:1000 dilution) and Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.25 µg/mL, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to PRMT4.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis was performed on modified whole cell extracts (1% SDS) (30 µg lysate) of MCF7 (Lane 1), HeLa (Lane 2), LNCaP (Lane 3), PC-3 (Lane 4), NTERA-2 (Lane 5) and COS-7 (Lane 6). The blot was probed with Anti-PRMT4 Monoclonal Antibody (3H2) (Product # MA5-15796, 1:1000 dilution) and detected by chemiluminescence using Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.25 µg/mL, 1:4000 dilution). A 65 kDa band corresponding to PRMT4 was observed in all cell lines tested.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of HeLa cells using CARM1 monoclonal antibody (Product # MA5-15796) (Green). Red: actin filaments have been labeled with phalloidin.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of PRMT4 was performed using 70% confluent log phase HeLa cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with PRMT4 Monoclonal Antibody (3H2) (Product # MA5-15796) at 1:200 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product #A28175) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415). Panel d represents the merged image showing Nuclear localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of paraffin-embedded breast cancer tissues (left) and ovarian cancer tissues (right) using CARM1 monoclonal antibody (Product # MA5-15796) followed with DAB staining.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Flow cytometric analysis of Lovo cells using CARM1 monoclonal antibody (Product # MA5-15796) (green) and negative control (purple).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Enrichment of endogenous PRMT4 protein at specific gene loci using Anti-PRMT4 Antibody: Chromatin Immunoprecipitation (ChIP) was performed using Anti-PRMT4 Monoclonal Antibody (Product # MA5-15796, 8 ul) on sheared chromatin from 2 million HCT 116 cells using the MAGnify ChIP system kit (Product # 49-2024). Normal Rabbit IgG was used as a negative IP control. The purified DNA was analyzed by qPCR with PCR primer pairs over the promoters of CCND1 and PS2 (positive) and SAT2 satellite repeats (negative). Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.

MA5-15796