Anti-Histone H3 (citrulline R17) antibody [EPR20358-120] - BSA and Azide free

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-Histone H3 (citrulline R17) antibody [EPR20358-120] - BSA and Azide free (AB232938)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed HEK-293T (Human epithelial cell line from embryonic kidney) transfected with empty vector (left panel) or PADI4 (WT, right panel), then treated with 10mM CaCl2 and 10μM Ionomycin for 2 hours, labeling Histone H3 (citrulline R17) with ab219407 at 1/500 dilution, followed by Goat anti rabbit IgG (Alexa Fluor^® 647) at 1/2000 dilution.

Positive signal is obtained from HEK-293T cells transfected with WT PADI4 treated with 10mM CaCl2 and 10μM Ionomycin for 2 hours.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab219407).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (citrulline R17) antibody [EPR20358-120] - BSA and Azide free (AB232938)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HEK-293T (Human epithelial cell line from embryonic kidney) cells transfected with GFP only or a GFP-tagged PADI4 expression construct, then treated with 10 mM CaCl₂ for 2 hours, labeling Histone H3 (citrulline R17) with ab219407 at 1/100 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing positive staining on HEK-293T cells transfected with a GFP-tagged PADI4 expression construct, then treated with 10mM CaCl₂ for 2 hours. The nuclear counterstain is DAPI (blue). Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594)) at 1/200 dilution (red).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/1000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab219407).

• IP

Lab

Immunoprecipitation - Anti-Histone H3 (citrulline R17) antibody [EPR20358-120] - BSA and Azide free (AB232938)

Histone H3 (citrulline R17) was immunoprecipitated from 0.35 mg of HEK-293T (Human epithelial cell line from embryonic kidney) transfected with PADI4 (WT), then treated with 10mM CaCl₂ and 10μM Ionomycin for 2 hours, whole cell lysate with ab219407 at 1/30 dilution.

Western blot was performed from the immunoprecipitate using ab219407 at 1/1000 dilution.

VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

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

Lane 2 : ab219407 IP in HEK-293T transfected with PADI4 (WT), then treated with 10mM CaCl₂ and 10μM Ionomycin for 2 hours, whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab219407 in HEK-293T transfected with PADI4 (WT), then treated with 10mM CaCl₂ and 10μM Ionomycin for 2 hours, whole cell lysate.

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Exposure time : 1 second.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab219407).

All lanes:

Immunoprecipitation - Anti-Histone H3 (citrulline R17) antibody [EPR20358-120] (<a href='/en-us/products/primary-antibodies/histone-h3-citrulline-r17-antibody-epr20358-120-ab219407'>ab219407</a>)

Predicted band size: 15 kDa

false

Exposure time: 1s

• I-ELISA

Supplier Data

Indirect ELISA - Anti-Histone H3 (citrulline R17) antibody [EPR20358-120] - BSA and Azide free (AB232938)

This data was developed using ab219407, the same antibody clone in a different buffer formulation.

ELISA analysis of Human Histone H3 (citrulline R17) recombinant protein at 100 ng/ml with ab219407. An Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) at 1/2500 dilution was used as the secondary antibody.

• WB

Lab

Western blot - Anti-Histone H3 (citrulline R17) antibody [EPR20358-120] - BSA and Azide free (AB232938)

All lanes:

Western blot - Anti-Histone H3 (citrulline R17) antibody [EPR20358-120] - BSA and Azide free (ab232938) at 1/50000 dilution

All lanes:

HEK-293T (Human epithelial cell line from embryonic kidney) transfected with PADI4 (WT), then treated with 10mM CaCl2 and 10µM Ionomycin (<a href='/en-us/products/biochemicals/ionomycin-ca2-salt-ca2-ionophore-ab120116'>ab120116</a>) for 2 hours, whole cell lysate at 15 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 1/20000 dilution

Predicted band size: 15 kDa

false

Exposure time: 60s

• ELISA

Unknown

ELISA - Anti-Histone H3 (citrulline R17) antibody [EPR20358-120] - BSA and Azide free (AB232938)

Direct ELISA using ab219407 at 0-1000 ng/ml, followed by Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG(H+L) at 1/2500 dilution. Antigen concentration : 1000 ng/ml.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab219407).