Revision 1

#8351Store at -20C

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IP, IF-IC

REACTIVITY:

H

SENSITIVITY:

Endogenous

MW (kDa):

160-200

Source/Isotype:

Rabbit IgG

UniProt ID:

#P55011

Entrez-Gene Id:

6558

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:2000
Immunoprecipitation 1:50
Immunofluorescence (Immunocytochemistry) 1:100

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Specificity / Sensitivity

NKCC1 (D13A9) Rabbit mAb recognizes endogenous levels of total NKCC1 protein. This antibody does not cross-react with NKCC2.

Species Reactivity:

Human

Species predicted to react based on 100% sequence homology

Bovine, Pig

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Arg80 of human NKCC1 protein.

Background

The electroneutral cation-chloride-coupled co-transporter (SLC12) gene family comprises bumetanide-sensitive Na+/K+/Cl- (NKCC), thiazide-sensitive Na+/Cl-, and K+/Cl- (KCC) co-transporters. SLC12A1/NKCC2 and SLC12A2/NKCC1 regulate cell volume and maintain cellular homeostasis in response to osmotic and oxidative stress (1). The broadly expressed NKCC1 is thought to play roles in fluid secretion (i.e. salivary gland function), salt balance (i.e. maintenance of renin and aldosterone levels), and neuronal development and signaling (2-7). During neuronal development, NKCC1 and KCC2 maintain a fine balance between chloride influx (NKCC1) and efflux (KCC2), which regulates γ-aminobutyric acid (GABA)-mediated neurotransmission (3). Increased NKCC1 expression in immature neurons maintains high intracellular chloride levels that result in inhibitory GABAergic signaling; KCC2 maintains low intracellular chloride levels and excitatory GABAergic responses in mature neurons (4,5,8). Deletion of NKCC1 impairs NGF-mediated neurite outgrowth in PC-12D cells while inhibition of NKCC1 with bumetanide inhibits re-growth of axotomized dorsal root ganglion cells (6,7). Defective chloride homeostasis in neurons is linked to seizure disorders that are ameliorated by butemanide treatment, indicating that abnormal NKCC1 and NKCC2 expression or signaling may play a role in neonatal and adult seizures (9-12). NKCC1 is found as a homodimer or within heterooligomers with other SLC12 family members. This transport protein associates with a number of oxidative- and osmotic-responsive kinases that bind, phosphorylate, and activate NKCC1 co-transporter activity (13-16). In response to decreased intracellular chloride concentrations, Ste20-related proline-alanine-rich kinase (SPAK) phosphorylates NKCC1 to increase co-transporter activity and promote chloride influx (16-19). Oxidative stress response kinase 1 (OSR1) also phosphorylates and activates NKCC1 in response to oxidative stress (14).

  1. Hebert, S.C. et al. (2004) Pflugers Arch 447, 580-93.
  2. Evans, R.L. et al. (2000) J Biol Chem 275, 26720-6.
  3. Kim, S.M. et al. (2008) Am J Physiol Renal Physiol 295, F1230-8.
  4. Khirug, S. et al. (2008) J Neurosci 28, 4635-9.
  5. Kahle, K.T. et al. (2008) Nat Clin Pract Neurol 4, 490-503.
  6. Nakajima, K. et al. (2007) Biochem Biophys Res Commun 359, 604-10.
  7. Pieraut, S. et al. (2007) J Neurosci 27, 6751-9.
  8. Ben-Ari, Y. (2002) Nat Rev Neurosci 3, 728-39.
  9. Fukuda, A. (2005) Nat Med 11, 1153-4.
  10. Dzhala, V.I. et al. (2005) Nat Med 11, 1205-13.
  11. Jayakumar, A.R. et al. (2008) J Biol Chem 283, 33874-82.
  12. Kahle, K.T. and Staley, K.J. (2008) Neurosurg Focus 25, E22.
  13. Moore-Hoon, M.L. and Turner, R.J. (2000) Biochemistry 39, 3718-24.
  14. Simard, C.F. et al. (2007) J Biol Chem 282, 18083-93.
  15. Piechotta, K. et al. (2002) J Biol Chem 277, 50812-9.
  16. Dowd, B.F. and Forbush, B. (2003) J Biol Chem 278, 27347-53.
  17. Geng, Y. et al. (2009) J Biol Chem 284, 14020-8.
  18. Smith, L. et al. (2008) J Biol Chem 283, 22147-56.
  19. Gagnon, K.B. et al. (2006) Mol Cell Biol 26, 689-98.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation IF-IC: Immunofluorescence (Immunocytochemistry)

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

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Orders: 877-616-CELL (2355)•[email protected]• Support: 877-678-TECH (8324)•[email protected]• Web: cellsignal.com
仅供研究使用。不得用于诊断流程。