Anti-CARM1 antibody [EPR26711-36] - BSA and Azide free

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-CARM1 antibody [EPR26711-36] - BSA and Azide free (AB307092)

This data was developed using ab307091, the same antibody clone in a different buffer formulation. Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized Wild-type HEK-293T (human embryonic kidney epithelial cell, Right) / CARM1 knockout HEK-293T (Left) cells labeling CARM1 with ab307091 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution was used as the secondary antibody. Positive staining on HEK-293T cells (ab255449), while no staining on CARM1 knockout HEK-293T cells (ab266557).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-CARM1 antibody [EPR26711-36] - BSA and Azide free (AB307092)

This data was developed using ab307091, the same antibody clone in a different buffer formulation. Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized CARM1 KO HEK293T (CARM1 knockout human embryonic kidney epithelial cell)(ab266557) cells labeling CARM1 with ab307091 at 1/500 dilution (1.026 ug/ml), followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (2 ug/ml) (Green). Confocal image showing nuclear and weak cytoplasmic staining in Parental HEK293T cell line, and no staining in CARM1 KO HEK293T cell line. Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (2.5 ug/ml) (Red). The nuclear counterstain was DAPI (Blue). Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (2 ug/ml).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-CARM1 antibody [EPR26711-36] - BSA and Azide free (AB307092)

This data was developed using ab307091, the same antibody clone in a different buffer formulation. Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized MCF7 (human breast adenocarcinoma epithelial cell) cells labeling CARM1 with ab307091 at 1/500 dilution (1.026 ug/ml), followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (2 ug/ml) (Green). Confocal image showing nuclear and weak cytoplasmic staining in MCF7 cell line. Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (2.5 ug/ml) (Red). The nuclear counterstain was DAPI (Blue). Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (2 ug/ml).

• IP

Supplier Data

Immunoprecipitation - Anti-CARM1 antibody [EPR26711-36] - BSA and Azide free (AB307092)

This data was developed using ab307091, the same antibody clone in a different buffer formulation. CARM1 was immunoprecipitated from 0.35 mg 293T (human embryonic kidney epithelial cell) whole cell lysate 10 ug with ab307091 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab307091 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution. Lane 1 : 293T whole cell lysate 10 ug Lane 2 : ab307091 IP in 293T whole cell lysate Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab307091 in 293T whole cell lysate Blocking and dilution buffer and concentration : 5% NFDM/TBST. Exposure time : 41 seconds.

All lanes:

Immunoprecipitation - Anti-CARM1 antibody [EPR26711-36] (<a href='/en-us/products/primary-antibodies/carm1-antibody-epr26711-36-ab307091'>ab307091</a>) at 1/1000 dilution

Lane 1:

293T (human embryonic kidney epithelial cell) whole cell lysate 10 μg

Lane 2:

293T whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 63 kDa

false

Exposure time: 41s

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-CARM1 antibody [EPR26711-36] - BSA and Azide free (AB307092)

This data was developed using ab307091, the same antibody clone in a different buffer formulation. Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (mouse embryonic fibroblast) cells labeling CARM1 with ab307091 at 1/500 dilution (1.026 ug/ml), followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (2 ug/ml) (Green). Confocal image showing nuclear and weak cytoplasmic staining in NIH/3T3 cell line. Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (2.5 ug/ml) (Red). The nuclear counterstain was DAPI (Blue). Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (2 ug/ml).

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-CARM1 antibody [EPR26711-36] - BSA and Azide free (AB307092)

This data was developed using ab307091, the same antibody clone in a different buffer formulation. Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized NIH/3T3 (mouse embryonic fibroblast) cells labeling CARM1 with ab307091 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution was used as the secondary antibody.

• IP

Supplier Data

Immunoprecipitation - Anti-CARM1 antibody [EPR26711-36] - BSA and Azide free (AB307092)

This data was developed using ab307091, the same antibody clone in a different buffer formulation. CARM1 was immunoprecipitated from 0.35 mg NIH/3T3 (mouse embryonic fibroblast) whole cell lysate 10 ug with ab307091 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab307091 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution. Lane 1 : NIH/3T3 whole cell lysate 10 ug Lane 2 : ab307091 IP in NIH/3T3 whole cell lysate Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab307091 in NIH/3T3 whole cell lysate Blocking and dilution buffer and concentration : 5% NFDM/TBST. Exposure time : 41 seconds.

All lanes:

Immunoprecipitation - Anti-CARM1 antibody [EPR26711-36] (<a href='/en-us/products/primary-antibodies/carm1-antibody-epr26711-36-ab307091'>ab307091</a>) at 1/1000 dilution

Lane 1:

NIH/3T3 (mouse embryonic fibroblast) whole cell lysate 10 μg

Lane 2:

NIH/3T3 whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 63 kDa

false

Exposure time: 41s

• WB

Supplier Data

Western blot - Anti-CARM1 antibody [EPR26711-36] - BSA and Azide free (AB307092)

This data was developed using ab307091, the same antibody clone in a different buffer formulation. Blocking and diluting buffer and concentration : 5% NFDM/TBST. In lane 1-4, the lysates were freshly made and used for Western Blotting immediately to minimize protein degradation. In lanes 5-8, the lysates were stored at -80degC prior to Western Blotting.  Bands above 250 kDa are aggregates of PRMT4. Exposure time : Lane 1,2 : 26 seconds Lane 3,4 : 48 seconds Lane 5-8 : 3 minutes

All lanes:

Western blot - Anti-CARM1 antibody [EPR26711-36] (<a href='/en-us/products/primary-antibodies/carm1-antibody-epr26711-36-ab307091'>ab307091</a>) at 1/1000 dilution

Lane 1:

HeLa (human epithelial cell line from cervical adenocarcinoma), whole cell lysate 20 μg

Lanes 2 and 5:

293T (human embryonic kidney epithelial cell) whole cell lysate 20 μg

Lanes 3 and 7:

NIH/3T3 (mouse embryonic fibroblast), whole cell lysate 20 μg

Lanes 4 and 8:

PC-12 (rat adrenal gland pheochromocytoma), whole cell lysate 20 μg

Lane 6:

HCT116 (human colorectal carcinoma epithelial cell), whole cell lysate 20 μg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 63 kDa

false

Exposure time: 26s

• WB

Supplier Data

Western blot - Anti-CARM1 antibody [EPR26711-36] - BSA and Azide free (AB307092)

This data was developed using ab307091, the same antibody clone in a different buffer formulation. Blocking and diluting buffer and concentration : Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS. Lysates at 20 µg per lane. Performed under reducing conditions. False colour image of Western blot : Anti-CARM1 antibody EPR26711-36 ab307091 staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody 6C5 loading control staining at 1/20000 dilution, shown in red. In Western blot, ab307091 was shown to bind specifically to CARM1. A band was observed at 63 kDa in wild-type HEK293T cell lysates with no signal observed at this size in CARM1 knockout cell line (knockout cell lysate). To generate this image, wild-type and CARM1 knockout HEK293T cell lysates were analyzed. First, samples were run on an SDS-PAGE gel then transferred onto an immobilon-FL PVDF membrane. Membranes were blocked in in Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-CARM1 antibody [EPR26711-36] (<a href='/en-us/products/primary-antibodies/carm1-antibody-epr26711-36-ab307091'>ab307091</a>) at 1/1000 dilution

Lane 1:

Wild-type HEK293T (human embryonic kidney epithelial cell), whole cell lysate

Lane 2:

CARM1 knockout HEK293T whole cell lysate

Lane 3:

HeLa (human epithelial cell line from cervical adenocarcinoma), whole cell lysate

Lane 4:

HCT116 (human colorectal carcinoma epithelial cell), whole cell lysate

Secondary

Lanes 1 - 4:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Lanes 1 - 4:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Observed band size: 63 kDa

false