Product Pathways - Ca / cAMP / Lipid Signaling
Calcium Ion Regulation Antibody Sampler Kit #8575
|8575S||1 Kit (6 x 40 µl)||---||In Stock||---|
|Kit Includes||Quantity||Applications||Reactivity||Homology†||MW (kDa)||Isotype|
|ATP2A2/SERCA2 (D51B11) Rabbit mAb #9580||40 µl||W, IP||H, M, R, Mk||114||Rabbit IgG|
|Phospho-Phospholamban (Ser16/Thr17) Antibody #8496||40 µl||W||R||H, M, B, Dg, Pg||6 (monomer); 12, 24 (oligomers)||Rabbit|
|Phospholamban Antibody #8495||40 µl||W||H, M, R||6 (monomer); 12, 24 (oligomers)||Rabbit|
|Phospho-PKA C (Thr197) (D45D3) Rabbit mAb #5661||40 µl||W||H, M, R, Mk||42||Rabbit IgG|
|PKA C-α (D38C6) Rabbit mAb #5842||40 µl||W, IP||H, M, R, Hm, Mk||42||Rabbit IgG|
|ATP2A1/SERCA1 (D54G12) Rabbit mAb #12293||40 µl||W||H, M||R||100||Rabbit IgG|
|Anti-rabbit IgG, HRP-linked Antibody #7074||100 µl||W||All||Goat|
†Species predicted to react based on 100% sequence homology.
Applications Key: W=Western Blotting, IP=Immunoprecipitation
Reactivity Key: H=Human, M=Mouse, R=Rat, Mk=Monkey, Hm=Hamster, All=All Species Expected
After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
Western blot analysis of extracts from various cell types using ATP2A2/SERCA2 (D51B11) Rabbit mAb.
Western blot analysis of extracts from HeLa, C6, and COS-7 cells using PKA C-α (D38C6) Rabbit mAb.
Western blot analysis of extracts from NIH/3T3 cells, untreated or λ phosphatase-treated, using Phospho-PKA C (Thr197) (D45D3) Rabbit mAb (upper) or PKA C-α Antibody #4782 (lower).
Western blot analysis of extracts from 16-month old control (WKY) and spontaneous hypertensive (SHR) rat hearts using Phospho-Phospholamban (Ser16/Thr17) Antibody (left), Phospholamban Antibody #8495 (middle), or GAPDH (14C10) Rabbit mAb #2118 (right).
Western blot analysis of extracts from 16-month old control (WKY) and spontaneous hypertensive (SHR) rat hearts using Phospho-Phospholamban (Ser16/Thr17) Antibody #8496 (left), Phospholamban Antibody (middle), or GAPDH (14C10) Rabbit mAb #2118 (right).
The Calcium Ion Regulation Antibody Sampler Kit provides an economical way to investigate the regulation of calcium ions within the cell. The kit contains enough primary and secondary antibodies to perform four western blot experiments per primary antibody.
Specificity / Sensitivity
ATP2A1/SERCA1 (D54G12) Rabbit mAb recognizes endogenous levels of total ATP2A1/SERCA1 protein. ATP2A2/SERCA2 (D51B11) Rabbit mAb recognizes endogenous levels of total ATP2A2/SERCA2 protein. Phospho-Phospholamban (Ser16/Thr17) Antibody recognizes endogenous levels of phospholamban protein only when phosphorylated at Ser16 and Thr17. This antibody does not detect mono- or non-phosphorylated phospholamban. Phospholamban Antibody recognizes endogenous levels of total phospholamban protein. Phospho-PKA C (Thr197) (D45D3) Rabbit mAb recognizes endogenous levels of PKA C (-α, -β, and -γ) only when phosphorylated at Thr197. PKA C-α (D38C6) Rabbit mAb recognizes endogenous levels of total PKA C-α protein.
Source / Purification
Monoclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near the amino terminus of human ATP2A2/SERCA2 protein, residues surrounding Pro995 of human ATP2A1/SERCA1 protein, residues surrounding Thr197 of human PKA C protein, or residues surrounding Ser326 of human PKA C-α protein. Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near the amino terminus of human phospholamban protein (not overlapping Ser16 and Thr17). Phospho-specific polyclonal antibodies are produced by immunizing animals with a synthetic phosphopeptide corresponsing to residues surrounding Ser16/Thr17 of human phospholamban protein. Polyclonal antibodies are purified by protein A and peptide affinity chromatography.
Sarcoplasmic and endoplasmic reticulum Ca2+ ATPases (SERCA) are members of a highly conserved family of Ca2+ pumps (1). ATP2A1 (SERCA1) is a fast-twitch, skeletal muscle sarcoplasmic reticulum (SR) Ca2+ ATPase (2). Multiple ATP2A2 (SERCA2) isoforms have been isolated, with ATP2A2a (SERCA2a) found predominantly in the SR of muscle cells and ATP2A2b (SERCA2b) more ubiquitously expressed in the ER of most cell types (3). Post-translational modification of ATP2A2, including phosphorylation and tyrosine nitration, modify Ca2+ -ATPase activity and calcium transport (4,5).
Phospholamban (PLN) was identified as a major phosphoprotein component of the SR (6). Despite very high expression in cardiac tissue, phospholamban is also expressed in skeletal and smooth muscle (7). Localization of PLN is limited to the SR, where it serves as a regulator of the sarco-endoplasmic reticulum calcium ATPase, SERCA (8). PLN binds directly to SERCA and effectively lowers its affinity for calcium, thus reducing calcium transport into the SR. Phosphorylation of PLN at Ser16 by PKA or myotonic dystrophy protein kinase and/or phosphorylation at Thr17 by Ca2+/calmodulin-dependent protein kinase results in release of PLN from SERCA, relief of this inhibition, and increased calcium uptake by SR (reviewed in 9,10). It has long been held that phosphorylation at Ser16 and Thr17 occurs sequentially, but increasing evidence suggests that phosphorylation, especially at Thr17, may be differentially regulated (reviewed in 11,12).
The second messenger cyclic AMP (cAMP) activates cAMP-dependent protein kinase (PKA or cAPK) in mammalian cells and controls many cellular mechanisms such as gene transcription, ion transport, and protein phosphorylation (13). Inactive PKA is a heterotetramer composed of a regulatory subunit (R) dimer and a catalytic subunit (C) dimer. In this inactive state, the pseudosubstrate sequences on the R subunits block the active sites on the C subunits. Three C subunit isoforms (C-α, C-β, and C-γ) and two families of the regulatory subunits (RI and RII) with distinct cAMP binding properties have been identified. Upon binding of cAMP to the R subunits, the auto-inhibitory contact is eased and active monomeric C subunits are released. PKA shares substrate specificity with Akt (PKB) and PKC, which are characterized by an arginine at position -3 relative to the phosphorylated serine or threonine residue (14). PKA phosphorylation is involved in the regulation of Ca2+ channels, including Cav1.1 in skeletal muscle and Cav1.2 in the heart (reviewed in 15).
- Hovnanian, A. (2007) Subcell Biochem 45, 337-63.
- Odermatt, A. et al. (1996) Nat Genet 14, 191-4.
- de Smedt, H. et al. (1991) J Biol Chem 266, 7092-5.
- Hawkins, C. et al. (1995) Mol Cell Biochem 142, 131-8.
- Viner, R.I. et al. (1999) Biochem J 340 ( Pt 3), 657-69.
- Kirchberber, M.A. et al. (1975) Recent Adv Stud Cardiac Struct Metab 5, 103-15.
- Fujii, J. et al. (1991) J Biol Chem 266, 11669-75.
- Tada, M. and Kirchberger, M.A. Recent Adv Stud Cardiac Struct Metab 11, 265-72.
- Traaseth, N.J. et al. (2008) Biochemistry 47, 3-13.
- Bhupathy, P. et al. (2007) J Mol Cell Cardiol 42, 903-11.
- Hagemann, D. and Xiao, R.P. (2002) Trends Cardiovasc Med 12, 51-6.
- Mattiazzi, A. et al. (2005) Cardiovasc Res 68, 366-75.
- Montminy, M. (1997) Annu Rev Biochem 66, 807-22.
- Dell'Acqua, M.L. and Scott, J.D. (1997) J Biol Chem 272, 12881-4.
- Dai, S. et al. (2009) Physiol Rev 89, 411-52.
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This product is intended for research purposes only. The product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
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