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Purified Mouse Anti-PKA RIIβ
Purified Mouse Anti-PKA RIIβ

Western blot analysis of PKA RIIβ on human endothelial lysate (left). Lane 1: 1:1000, lane 2: 1:2000, lane 3: 1:4000 dilution of PKA RIIβ.

Immunofluorescent staining of SK-N-SH cells (right).  Cells were seeded in a 384 well collagen coated Microplates (Material # 353962) at ~ 8,000 cells per well.  After overnight incubation, cells were stained using the Triton X100 fix/perm protocol (see Recommended Assay Procedure) and the anti- PKARIIb antibody.  The second step reagent was Alexa Fluor® 488 goat anti mouse Ig (Invitrogen).  The image was taken on a Pathway 855 or 435 imager using a 20x objective.  This antibody also stained SH-SY5Y and C6 cells using both the Triton X100 and methanol fix/perm protocols (see Recommended Assay Procedure).

Western blot analysis of PKA RIIβ on human endothelial lysate (left). Lane 1: 1:1000, lane 2: 1:2000, lane 3: 1:4000 dilution of PKA RIIβ.

Immunofluorescent staining of SK-N-SH cells (right).  Cells were seeded in a 384 well collagen coated Microplates (Material # 353962) at ~ 8,000 cells per well.  After overnight incubation, cells were stained using the Triton X100 fix/perm protocol (see Recommended Assay Procedure) and the anti- PKARIIb antibody.  The second step reagent was Alexa Fluor® 488 goat anti mouse Ig (Invitrogen).  The image was taken on a Pathway 855 or 435 imager using a 20x objective.  This antibody also stained SH-SY5Y and C6 cells using both the Triton X100 and methanol fix/perm protocols (see Recommended Assay Procedure).

Product Details
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BD Transduction Laboratories™
Human (QC Testing), Mouse, Rat, Dog, Chicken (Tested in Development)
Mouse IgG1
Human PKA RIIβ, aa 1-418
Western blot (Routinely Tested), Immunofluorescence (Tested During Development), Immunohistochemistry, Immunoprecipitation (Not Recommended)
53 kDa
250 µg/ml
None
AB_397958
Aqueous buffered solution containing BSA, glycerol, and ≤0.09% sodium azide.
RUO


Preparation And Storage

The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. Store undiluted at -20°C.

Recommended Assay Procedures

Methanol Procedure for a 96 well plate:

Remove media from wells. Add 100 µl/well fresh 3.7% Formaldehyde in PBS. Incubate for 10 minutes at room temperature (RT). Flick out and add 100 µl/well 90% methanol. Incubate for 5 minutes at RT. Flick out and wash twice with PBS. Flick out PBS and add 100 µl/well blocking buffer (3% FBS in PBS). Incubate for 30 minutes at RT. Flick out and add diluted antibody (diluted in blocking buffer). Incubate for 1 hour at RT. Wash three times with PBS. Flick out PBS and add second step reagent. Incubate for 1 hour at RT. Wash three times with PBS. Image sample.

Triton-X 100 Procedure for a 96 well plate:

Remove media from wells. Add 100 µl/well fresh 3.7% Formaldehyde in PBS. Incubate for 10 minutes at room temperature (RT). Flick out and add 100 µl/well 0.1% Triton-X 100. Incubate for 5 minutes at RT. Flick out and wash twice with PBS. Flick out PBS and add 100 µl/well blocking buffer (3% FBS in PBS). Incubate for 30 minutes at RT. Flick out and add diluted antibody (diluted in blocking buffer). Incubate for 1 hour at RT. Flick out and wash three times with PBS. Flick out and add second step reagent. Incubate for 1 hour at RT. Flick out and wash three times with PBS. Image sample.

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  3. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
  4. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
610626 Rev. 1
Antibody Details
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45

cAMP-dependent Protein Kinase (PKA) is composed of two distinct subunits: catalytic (C) and regulatory (R). Four regulatory subunits have been identified: RIα, RIß, RIIα, and RIIß.These subunits define type I and II cAMP-dependent protein kinases. Following binding of cAMP, the regulatory subunits dissociate from the catalytic subunits, rendering the enzyme active. Type I and type II holoenzymes have three potential C subunits (Cα, Cß, or Cγ). Type II PKA can be distinguished by autophosphorylation of the R-subunits, while type I PKA binds Mg/ATP with high affinity. Most cells express both type I and type II PKAs. Although the Rα isoforms are ubiquitously expressed, the Rß isoforms are predominant in nervous and adipose tissues. There are indications that the deletion of the gene for PKA RIIß results in lack of long-term potentiation in a select group of hippocampal cells, suggesting an important role for this protein in the neurosciences.

610626 Rev. 1
Format Details
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Purified
Tissue culture supernatant is purified by either protein A/G or affinity purification methods. Both methods yield antibody in solution that is free of most other soluble proteins, lipids, etc. This format provides pure antibody that is suitable for a number of downstream applications including: secondary labeling for flow cytometry or microscopy, ELISA, Western blot, etc.
Purified
610626 Rev.1
Citations & References
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Development References (5)

  1. Casey M, Vaughan CJ, He J, et al. Mutations in the protein kinase A R1alpha regulatory subunit cause familial cardiac myxomas and Carney complex. J Clin Invest. 2000; 106(5):R31-R38. (Clone-specific: Western blot). View Reference
  2. Sandberg M, Skalhegg B, Jahnsen T. The two mRNA forms for the type I alpha regulatory subunit of cAMP-dependent protein kinase from human testis are due to the use of different polyadenylation site signals. Biochem Biophys Res Commun. 1990; 167(1):323-330. (Biology). View Reference
  3. Skalhegg BS, Landmark B, Foss KB, et al. Identification, purification, and characterization of subunits of cAMP-dependent protein kinase in human testis. Reverse mobilities of human RII alpha and RII beta on sodium dodecyl sulfate-polyacrylamide gel electrophoresis compared with rat and bov. J Biol Chem. 1992; 267(8):5374-5379. (Biology). View Reference
  4. Tasken KA, Collas P, Kemmner WA, Witczak O, Conti M, Tasken K. Phosphodiesterase 4D and protein kinase a type II constitute a signaling unit in the centrosomal area. J Biol Chem. 2001; 276(25):21999-22002. (Clone-specific: Immunoprecipitation, Western blot). View Reference
  5. Tavalin SJ, Colledge M, Hell JW, Langeberg LK, Huganir RL, Scott JD. Regulation of GluR1 by the A-kinase anchoring protein 79 (AKAP79) signaling complex shares properties with long-term depression. J Neurosci. 2002; 22(8):3044-3051. (Clone-specific: Western blot). View Reference
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610626 Rev. 1

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Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described


Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

For Research Use Only. Not for use in diagnostic or therapeutic procedures.