711722

antibody from Invitrogen Antibodies

Targeting: PRKACB PKACb

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Antibody Data

Product number

711722

Provider

Invitrogen Antibodies

Product name

Phospho-PKA alpha/beta (Thr197) Recombinant Polyclonal Antibody (6HCLC)

Antibody type

Polyclonal

Antigen

Synthetic peptide

Description

This antibody is predicted to react with Monkey, Rat, Mouse, Bovine and Pig

Antibody clone number

6HCLC

Concentration

0.5 mg/mL

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis was performed on Whole cell extracts (30 µg lysate) of HeLa (Lane 1), Neuro-2a (Lane 2) and HEK-293 (Lane 3). The blots were probed with Phospho-PKA alpha/beta (Thr197) Recombinant Rabbit Polyclonal Antibody (Product # 711722, 1-2 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2,500 dilution). A 40 kDa band corresponding to PKA CAT alpha/beta was observed across the cell lines tested. Known quantity of protein samples were electrophoresed using Novex®NuPAGE®4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

For immunofluorescence analysis, HeLa cells were fixed and permeabilized for detection of endogenous PKA CAT alpha/beta using Phospho-PKA alpha/beta (Thr197) Recombinant Rabbit Polyclonal Antibody (Product # 711722, 2 µg/mL) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2,000). Panel a) shows representative cells that were stained for detection and localization of PKA CAT alpha/beta (green), Panel b) is stained for nuclei (blue) using SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). Panel c) represents cytoskeletal F-actin staining using Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d) is a composite image of Panels a, b and c clearly demonstrating cytoplasmic localization of PKA CAT alpha/beta. Panel e) represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.