Anti-hnRNP A1 (citrulline R88 + R92) antibody [EPR20175] - BSA and Azide free

• WB

Supplier Data

Western blot - Anti-hnRNP A1 (citrulline R88 + R92) antibody [EPR20175] - BSA and Azide free (AB251486)

This data was developed using ab208027, the same antibody clone in a different buffer formulation.

hnRNP A1 (Uniprot P09651), isoform A1-A (34kD) is twenty times more abundant than isoform A1-B (39kD). The molecular weights of hnRNP A1L2 (Q32P51), hnRNP A0 (Q13151) and hnRNP A2B1 (P22626) are 34kD, 31kD and 37kD respectively.

PAD enzymes (including PADI2 and PADI4) hydrolyze arginine to form citrulline, Ca²⁺ and ionomycin are also required for this catalytic reaction (PMID : 26360112).

Blocking and dilution buffer : 5% NFDM/TBST.

Exposure times : Lanes 1-2 : 1 second; Lanes 3-4 : 2 seconds.

All lanes:

Western blot - Anti-hnRNP A1 (citrulline R88 + R92) antibody [EPR20175] (<a href='/en-us/products/primary-antibodies/hnrnp-a1-citrulline-r88-r92-antibody-epr20175-ab208027'>ab208027</a>) at 1/5000 dilution

Lane 1:

HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) transfected with empty vector (control) then treated with 10 mM CaCl2 and 10 μM Ionomycin for 2 hours, whole cell lysate at 20 µg

Lane 2:

HEK-293T transfected with a PADI4 expression vector then treated with 10 mM CaCl2 and 10 μM Ionomycin for 2 hours, whole cell lysate at 20 µg

Lane 3:

HEK-293T transfected with an empty vector (control) then treated with 10 mM CaCl2 and 10 μM Ionomycin for 2 hours, whole cell lysate at 20 µg

Lane 4:

HEK-293T transfected with a PADI2 expression vector then treated with 10 mM CaCl2 and 10 μM Ionomycin for 2 hours, whole cell lysate at 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

Predicted band size: 38 kDa

Observed band size: 31 kDa,37 kDa,39 kDa

true

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-hnRNP A1 (citrulline R88 + R92) antibody [EPR20175] - BSA and Azide free (AB251486)

This data was developed using ab208027, the same antibody clone in a different buffer formulation.

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) cells transfected with PADI4 expression vector, then treated with 10 mM CaCl2 and 10 μM Ionomycin for 2 hours, labeling hnRNP A1 with ab208027 at 1/5000 dilution (right panel) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (left panel). Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 647) (ab150079) at 1/2000 dilution was used as the secondary antibody.

The CaCl2 and Ionomycin treated GFP positive cell population gives a positive signal. (Right panel, Q2).

• IP

Supplier Data

Immunoprecipitation - Anti-hnRNP A1 (citrulline R88 + R92) antibody [EPR20175] - BSA and Azide free (AB251486)

This data was developed using ab208027, the same antibody clone in a different buffer formulation.

hnRNP A1 was immunoprecipitated from 0.35 mg HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) with ab208027 at 1/30 dilution, whole cell lysate. Western blot was performed from the immunoprecipitate using ab208027 at 1/10000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1 : HEK-293T transfected with PADI4 expression vector, then treated with 10 mM CaCl₂ and 10μM Ionomycin for 2 hours whole cell lysate 10 μg (Input).

Lane 2 : ab208027 IP in HEK-293T transfected with PADI4 expression vector, then treated with 10 mM CaCl₂ and 10 μM Ionomycin for 2 hours whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab208027 in HEK-293T transfected with PADI4 expression vector, then treated with 10 mM CaCl₂ and 10 μM Ionomycin for 2 hours whole cell lysate.

Blocking and dilution buffer : 5% NFDM/TBST.

Exposure time : 1 second.

All lanes:

Immunoprecipitation - Anti-hnRNP A1 (citrulline R88 + R92) antibody [EPR20175] (<a href='/en-us/products/primary-antibodies/hnrnp-a1-citrulline-r88-r92-antibody-epr20175-ab208027'>ab208027</a>)

Predicted band size: 38 kDa

Observed band size: 31 kDa,34 kDa,37 kDa

false

• Dot

Supplier Data

Dot Blot - Anti-hnRNP A1 (citrulline R88 + R92) antibody [EPR20175] - BSA and Azide free (AB251486)

This data was developed using ab208027, the same antibody clone in a different buffer formulation.

Dot Blot analysis of hnRNP A1 (citrulline R88 + R92) labeled with ab208027 at 1/1000 dilution.

Lane 1 : hnRNP A1 (citrulline R88/92) peptide.
Lane 2 : hnRNP A3 (citrulline R109/113) peptide.
Lane 3 : hnRNP A0 (citrulline R81/85) peptide.
Lane 4 : hnRNP A2B1 (citrulline R95/99) peptide.
Lane 5 : CCDC51 (citrulline R142) peptide.
Lane 6 : hnRNP A1 non-citrulline peptide.
Lane 7 : hnRNP A1 (citrulline R88) peptide.
Lane 8 : hnRNP A1 (citrulline R92) peptide.

Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution was used as secondary antibody.

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : 30 seconds.