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Key Specifications Table
| Species Reactivity | Key Applications | Host | Format | Antibody Type |
|---|---|---|---|---|
| B, R, Av, H, Gr, M, Rb, F, Ft, Shk, Ca | EM, ICC, IF, IP, WB | M | Purified | Monoclonal Antibody |
| Description | |
|---|---|
| Catalogue Number | MABS1237 |
| Description | Anti-NKCC1/2 Antibody, clone T4 |
| Alternate Names |
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| Background Information | Solute carrier family 12 member 1 (UniProt Q13621; a.k.a. Bumetanide-sensitive sodium-(potassium)-chloride cotransporter 2) and solute carrier family 12 member 2 (UniProt P55011; a.k.a. Basolateral Na-K-Cl symporter, Bumetanide-sensitive sodium-(potassium)-chloride cotransporter 1) are encoded by the SLC12A1 (a.k.a. NKCC2; Gene ID 6557) and SLC12A2 (a.k.a. NKCC1; Gene ID 6558) gene, respectively, in human. NKCC1 (SLC12A2) and NKCC2 (SLC12A1) belong to the subfamily of cation-chloride cotransporters (CCCs) that mediate electroneutral transport of sodium, potassium and chloride across the plasma membrane and are inhibited by bumetanide and furosemide-type of compounds. NKCCs play an important role in cell volume regulation, where NKCC2 is involved in reabsorption of salt and osmotically-obliged water in renal epithelial cells, while NKCC1 has a major impact on intracellular chloride ion concentration ([Cl-]i) in vascular smooth muscle cells (VSMCs) and neuronal cells. In primary hypertension, NKCC1 activity is increased in VSMCs and paraventricular nucleus (PVN) neurons, leading to elevated peripheral resistance in systemic circulation and activation of sympathetic nerve system (SNS), respectively. Human NKCC1 and NKCC2 are 12-transmembrane proteins having their N- and C-terminal ends exposed at the cytoplasmic side (a.a. 1-285 and 921-1212 of NKCC1; a.a. 1-177 and 814-1099 of NKCC2). |
| Product Information | |
|---|---|
| Format | Purified |
| Presentation | Purified mouse monoclonal IgG1κ in buffer containing 0.1 M Tris-Glycine (pH 7.4) 150 mM NaCl with 0.05% sodium azide |
| Quality Level | MQ100 |
| Applications | |
|---|---|
| Application | This mouse monoclonal Anti-NKCC1/2 Antibody, clone T4, Cat. No. MABS1237 detects levels of NKCC1 & NKCC2, and has been published and validated for use in Electron Microscopy, Immunocytochemistry, Immunofluorescence, Immunoprecipitation, and Western Blotting. |
| Key Applications |
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| Application Notes | Immunohistochemistry Analysis: A 1:3,000 dilution from a representative lot detected NKCC in human prostate cancer, as well as normal colon, small intestine, and testis tissue sections. Electron Microscopy Analysis: A representative lot detected NKCC immunoreactivity in 0.5% glutaraldehyde and 4% paraformaldehyde gerbil inner ear tissue sections (Crouch, J.J., et al. (1997). J. Histochem. Cytochem. 45(6):773-778). Immunofluorescence Analysis: A representative lot detected NKCC-positive cells by fluorescent immunohistochemistry staining of 4% paraformaldehyde- or in 2.5% glutaraldehyde-fixed, paraffin-embedded fish gill filaments (Brauer, P.R., et al. (2005). Anat. Rec. A Discov. Mol. Cell. Evol. Biol. 285(1):600-609). Immunofluorescence Analysis: Representative lots detected brain tissue NKCC immunoreactivity by fluorescent immunohistochemistry staining of free-floating mouse and rat brain sections (Liu, F., et al. (2010). Exp. Neurol. 224(2):356-361; Yan, Y., et al. (2001). J. Cereb. Blood Flow Metab. 21(6):711 721). Immunofluorescence Analysis: A representative lot detected NKCC immunolocalization by fluorescent immunohistochemistry staining of MeOH (80%)/DMSO (20%) fixed rat kidney cryosections (Ginns, S.M., et al. (1996). J. Am. Soc. Nephrol. 7(12):2533-2542). Immunofluorescence Analysis: A representative lot immunostained basolateral margin of the acinar cells, but not the apical pole or cell interior by fluorescent immunohistochemistry staining of PLP-fixed (2% paraformaldehyde, 75 mM lysine, 10 mM sodium periodate) rabbit parotid gland cryosections (Lytle, C., et al. (1995). Am. J. Physiol. 269(6 Pt 1):C1469-C1505). Immunohistochemistry Analysis: A representative lot detected NKCC immunoreactivity by immunohistochemistry staining of 4% paraformaldehyde-fixed free-floating rat brain sections (Yan, Y., et al. (2001). J. Cereb. Blood Flow Metab. 21(6):711 721). Immunohistochemistry Analysis: A representative lot detected NKCC immunoreactivity in 10% formalin-fixed, Paraplast Plus-embedded gerbil kidney and inner ear tissue sections (Crouch, J.J., et al. (1997). J. Histochem. Cytochem. 45(6):773-778). Immunohistochemistry Analysis: A representative lot detected Na-K-Cl cotransporters in serous acinar cells, but not along striated ducts in PLP-fixed (2% paraformaldehyde, 75 mM lysine, 10 mM sodium periodate) rabbit parotid gland cryosections (Lytle, C., et al. (1995). Am. J. Physiol. 269(6 Pt 1):C1469-C1505). Immunoprecipitation Analysis: Representative lot immunoprecipitated denatured, but not non-denatured human and canine Na-K-Cl cotransport proteins (Lytle, C., et al. (1995). Am. J. Physiol. 269(6 Pt 1):C1469-C1505; Haas, M., et al. (1995). J. Biol. Chem. 270(48):28955 28961). Western Blotting Analysis: A representative lot detected exogenously expressed ferret NKCC1 & NKCC2A in lysates from transfected HEK293 cells (Hannemann, A., and Flatman, P.W. (2011). PLoS One. 6(3):e17992). Western Blotting Analysis: Representative lots detected mouse, rat, fish, and bovine NKCC proteins (Liu, F., et al. (2010). Exp. Neurol. 224(2):356-361; Brauer, P.R., et al. (2005). Anat. Rec. A Discov. Mol. Cell. Evol. Biol. 285(1):600-609; Abbruscato, T.J., et al. (2004). J. Pharmacol. Exp. Ther. 310(2):459-468; Yan, Y., et al. (2001). J. Cereb. Blood Flow Metab. 21(6):711 721). Western Blotting Analysis: A representative lot detected Na-K-Cl cotransport protein (~160-195 kDa) in membrane extracts from shark rectal gland, mouse medullary thick ascending limb, canine kidney, duck salt gland, human colonic T84 cells, and rabbit parotid salivary gland. N-glycosylase F treatment of the extracts reduced the sizes of target bands (Lytle, C., et al. (1995). Am. J. Physiol. 269(6 Pt 1):C1469-C1505). Note: Clone T4 epitope is not exposed on the surface of Na-K-Cl cotransport protein in its native conformation. Denature NKCC with the addtion of SDS to a final concentration of 1% in test samples, then diluting the denatured samples into 6 v |
| Biological Information | |
|---|---|
| Immunogen | Recombinant human NKCC1 C-terminal fragment. |
| Clone | T4 |
| Concentration | Please refer to lot specific datasheet. |
| Host | Mouse |
| Specificity | Clone T4 recognizes both NKCC1 and NKCC2 by targeting a C-terminal region epitope exposed only under denaturing condition, and unaccessible for antibody binding when NKCC assumes its native conformation (Lytle, C., et al. (1995). Am. J. Physiol. 269(6 Pt 1):C1469-C1505). |
| Isotype | IgG1κ |
| Species Reactivity |
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| Species Reactivity Note | Avian, Bovine, Canine, Ferret, Fish, Gerbil, Human, Mouse, Rabbit, Rat, Shark. |
| Antibody Type | Monoclonal Antibody |
| Entrez Gene Number |
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| Gene Symbol |
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| Purification Method | Protein G purified. |
| UniProt Number |
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| Molecular Weight | ~165 kDa observed. Target bands appear larger than the calculated molecular weights of 131.4/129.7 kDa (human NKCC1 isoform 1/2), 121.5/121.4 kDa (human NKCC2 isoform A/F), 131.0 kDa (mouse NKCC1), 120.4/120.3/120.3 kDa (mouse NKCC2 isoform A/B/F), and 130.4/120.6 kDa (rat NKCC1/NKCC2) due to glycosylation. Uncharacterized bands may be observed in some lysate(s). |
| Product Usage Statements | |
|---|---|
| Quality Assurance | Evaluated by Western Blotting in HeLa cell lysate. Western Blotting Analysis: 0.5 µg/mL of this antibody detected NKCC in 10 µg of HeLa cell lysate. |
| Usage Statement |
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| Storage and Shipping Information | |
|---|---|
| Storage Conditions | Stable for 1 year at 2-8°C from date of receipt. |
| Packaging Information | |
|---|---|
| Material Size | 100 μg |