SLC12A2 / NKCC1 Recombinant Rabbit Monoclonal Antibody [PSH23-65]
cat.: HA724390
Product Type: Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity: Human, Mouse, Rat
Applications: WB, IF-Cell, IHC-Fr, IHC-P, FC(Intra), IP
Clonality: Monoclonal
Clone number: PSH23-65
Form: Liquid
Storage condition: Shipped at 4℃. Store at +4℃ short term (1-2 weeks). Store at -20℃ long term.
Storage buffer: 1*PBS (pH7.4), 0.1% BSA, 40% Glycerol, 0.2% Proclean 950.
Concentration: 1ug/ul
Purification: Protein A affinity purified.
Molecular weight: Predicted band size: 131 kDa
Isotype: IgG
Immunogen: Recombinant protein within mouse SLC12A2 / NKCC1 aa 101-279.
Positive control: COLO205 (Human colon adenocarcinoma cells) cell lysate, HT-29 (Human colorectal adenocarcinoma cells) cell lysate, HeLa (Human cervical adenocarcinoma cells) cell lysate, NIH/3T3 (Mouse fibroblasts) cell lysate, Mouse cerebellum tissue lysate, Mouse brain tissue lysate, Mouse colon tissue lysate, Mouse small intestine tissue lysate, Rat cerebellum tissue lysate, Rat brain tissue lysate.
Subcellular location: Basolateral cell membrane.
Recommended Dilutions:
  WB
  IF-Cell
  IHC-Fr
  IHC-P
  FC(Intra)
  IP
1:5,000
1:500
1:500
1:5,000
1:1,000
1-2μg/sample
Uniprot #: SwissProt: P55011 Human | P55012 Mouse
Entrez Gene: 83629 Rat
Alternative names: Solute carrier family 12 member 2 Basolateral Na-K-Cl symporter Bumetanide-sensitive sodium-(potassium)-chloride cotransporter 2 (BSC2) Na-K-2Cl cotransporter 1 (hNKCC1) SLC12A2 NKCC1
Images
HA724390_1.jpg Fig1: Western blot analysis of SLC12A2 / NKCC1 on different lysates with Rabbit anti-SLC12A2 / NKCC1 antibody (HA724390) at 1/5,000 dilution.

Lane 1: COLO205 (Human colon adenocarcinoma cells) cell lysate
Lane 2: HT-29 (Human colorectal adenocarcinoma cells) cell lysate
Lane 3: HeLa (Human cervical adenocarcinoma cells) cell lysate
Lane 4: NIH/3T3 (Mouse fibroblasts) cell lysate

Lysates/proteins at 20 µg/Lane.
Exposure time: 20 seconds; ECL: K1801

Blocking: 5% NFDM/TBST, 1 hour at room temperature
Primary antibody: HA724390, 1/5,000 in primary antibody dilution buffer (K1803), overnight at 4 °C
Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature

Predicted band size: 131 kDa
Observed band size: 150-200 kDa
HA724390_2.jpg Fig2: Western blot analysis of SLC12A2 / NKCC1 on different lysates with Rabbit anti-SLC12A2 / NKCC1 antibody (HA724390) at 1/5,000 dilution.

Lane 1: Mouse cerebellum tissue lysate
Lane 2: Mouse brain tissue lysate
Lane 3: Mouse liver tissue lysate
Lane 4: Mouse colon tissue lysate
Lane 5: Mouse small intestine tissue lysate
Lane 6: Rat cerebellum tissue lysate
Lane 7: Rat liver tissue lysate
Lane 8: Rat brain tissue lysate

Lysates/proteins at 20 µg/Lane.
Exposure time: 20 seconds; ECL: K1801

Low expression: Liver (PMID:12000719)

Blocking: 5% NFDM/TBST, 1 hour at room temperature
Primary antibody: HA724390, 1/5,000 in primary antibody dilution buffer (K1803), overnight at 4 °C
Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature

Predicted band size: 131 kDa
Observed band size: 150-200 kDa
HA724390_3.jpg Fig3: Application: Immunocytochemistry (IF-cell)

Species: Human
Sample: HT-29 (Human colorectal adenocarcinoma cells)

Fixation: 4% Paraformaldehyde, 15 minutes at room temperature.
Permeabilization: 0.1% Triton X-100, 15 minutes at room temperature.

Blocking: 1% BSA + 10% normal goat serum, 1 hour at room temperature.
Antibody dilution buffer: 1% BSA in PBST.
Primary antibody: HA724390, 1/500, overnight at 4°C.
Secondary antibody: Goat Anti-Rabbit IgG (iFluor™ 488, HA1121), 45 minutes at room temperature.

Counterstain: Beta tubulin (HA601187, red), 1/100, overnight at 4℃. The Nuclear counterstain was DAPI (Blue).
HA724390_4.jpg Fig4: Application: Immunofluorescence (IHC-Fr)

Species: Mouse
Tissue: Colon
Sample: Frozen section

Antigen retrieval: Not required

Wash buffer: 1× TBST
Blocking: 10% normal goat serum + 0.5 % Triton X-100 + 0.3 M Glycine in PBS, 10 minutes at room temperature.
Primary antibody: HA724390, 1/500, overnight at 4°C.
Secondary antibody: Goat Anti-Rabbit IgG (iFluor™ 488, HA1121), 1.5 hours at room temperature.
HA724390_5.jpg Fig5: Application: Immunofluorescence (IHC-Fr)

Species: Rat
Tissue: Colon
Sample: Frozen section

Antigen retrieval: Not required

Wash buffer: 1× TBST
Blocking: 10% normal goat serum + 0.5 % Triton X-100 + 0.3 M Glycine in PBS, 10 minutes at room temperature.
Primary antibody: HA724390, 1/500, overnight at 4°C.
Secondary antibody: Goat Anti-Rabbit IgG (iFluor™ 488, HA1121), 1.5 hours at room temperature.
HA724390_6.jpg Fig6: Application: Immunohistochemistry (IHC-P)

Species: Human
Tissue: Colon
Sample: Paraffin-embedded section

Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.

Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724390, 1/5,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
HA724390_7.jpg Fig7: Application: Immunohistochemistry (IHC-P)

Species: Human
Tissue: Cerebellum
Sample: Paraffin-embedded section

Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.

Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724390, 1/5,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
HA724390_8.jpg Fig8: Application: Immunohistochemistry (IHC-P)

Species: Mouse
Tissue: Colon
Sample: Paraffin-embedded section

Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.

Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724390, 1/5,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
HA724390_9.jpg Fig9: Application: Immunohistochemistry (IHC-P)

Species: Mouse
Tissue: Cerebellum
Sample: Paraffin-embedded section

Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.

Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724390, 1/5,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
HA724390_10.jpg Fig10: Application: Immunohistochemistry (IHC-P)

Species: Mouse
Tissue: Liver
Sample: Paraffin-embedded section

Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95℃.

Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724390, 1/5,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.

Low expression: Liver (PMID:12000719).
HA724390_11.jpg Fig11: Application: Immunohistochemistry (IHC-P)

Species: Rat
Tissue: Colon
Sample: Paraffin-embedded section

Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.

Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724390, 1/5,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
HA724390_12.jpg Fig12: Application: Immunohistochemistry (IHC-P)

Species: Rat
Tissue: Cerebellum
Sample: Paraffin-embedded section

Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.

Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724390, 1/5,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
HA724390_13.jpg Fig13: Application: Flow Cytometry (Intra)

Species: Human
Sample: HT-29 (Human colorectal adenocarcinoma cells)

Fixation: 4% Paraformaldehyde, 15 minutes at room temperature.
Permeabilization: 1x Permeabilization Buffer (eBioscience), 15 minutes at room temperature.

Blocking: 1% BSA + 10% normal goat serum, 15 minutes at room temperature.
Antibody dilution buffer: 1x PBS.
Primary antibody: HA724390 (1/1,000) (Red) compared with Rabbit IgG Isotype Control (HA722127, Green), 15 minutes at room temperature.
Secondary antibody: Goat Anti-Rabbit IgG (iFluor™ 488, HA1121), 15 minutes at room temperature.
HA724390_14.jpg Fig14: Immunoprecipitation (IP)

SLC12A2 / NKCC1 was immunoprecipitated in 0.2 mg COLO205 (Human colon adenocarcinoma cells) cell lysate with HA724390 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA724390 at 1/5,000 dilution. Anti-Rabbit IgG for IP Nano-secondary antibody (NBI01H) at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: COLO205 cell lysate (input)
Lane 2: HA724390 IP in COLO205 cell lysate
Lane 3: Rabbit IgG instead of HA724390 in COLO205 cell lysate

Exposure time: 7 seconds
Blocking: 5% NFDM/TBST, 1 hour at room temperature

Primary dilution: HA724390, 1/5,000 in primary antibody dilution buffer (K1803), 2 hours at room temperature

Predicted band size: 131 kDa
Observed band size: 150-200 kDa
Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.