710680

antibody from Invitrogen Antibodies

Targeting: FOXA2 HNF3B

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Antibody Data

Product number

710680

Provider

Invitrogen Antibodies

Product name

Phospho-FOXA2 (Thr156) Recombinant Polyclonal Antibody (10HCLC)

Antibody type

Polyclonal

Antigen

Synthetic peptide

Description

This antibody is predicted to react with mouse, bovine and zebrafish based on sequence homology.

Antibody clone number

10HCLC

Concentration

0.5 mg/mL

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis was performed to detect FOXA2 (pT156) protein by loading 30 µg of Hep G2 and Ms. Liver lysates (lane 1, 2) using a Novex®NuPAGE®4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock Electrophoresis System (Product # EI0002), Novex® Sharp Pre-Stained Protein Standard (Product # LC5800) and iBlot® Dry Blotting System (Product # IB21001). Proteins were transferred to a nitrocellulose membrane and blocked with 5% skim milk for 1 hour at room temperature. To confirm specificity; competition was performed with phosphopeptide (10 µg/mL, preincubated with antibody 30 minutes (lane 3 and 4). FOXA2 (pT156) was detected at ~48 kDa using FOXA2 (pT156) Recombinant Rabbit Polyclonal Antibody (Product # 710680) at 2 µg/mL in 2.5% skim milk at 4°C; incubated overnight on a rocking platform. Goat anti-Rabbit IgG-HRP Secondary Antibody (Product # G-21234) was used at 1:5000 dilution and chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescent analysis of FoxA2 (pT156) was done on 70% confluent log phase HepG2 cells. The cells were fixed with 4% paraformaldehyde for 15 minutes; permeabilized with 0.25% Triton X-100 for 10 minutes followed by blocking with 5% BSA for 1 hour at room temperature. The cells were incubated with FoxA2 (pT156) Recombinant Rabbit Polyclonal Antibody (Product # 710680) at a dilution of 1:400 in 1% BSA and incubated for 3 hours at room temperature and then labeled with Alexa Fluor® 488 Goat anti-Rabbit IgG Secondary Antibody (Product # A-11008) at a dilution of 1:400 for 30 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 594 Phalloidin (Product # A12381). Panel d is a merged image showing nuclear localization of FoxA2 (pT156). Panel e is no primary antibody control. Panel e shows competition with FoxA2 (pT156) peptide. The images were captured at 20X magnification.