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Anti-NCX1 Mouse Antibody [C23J23]

Catalog No.: F2381

Application: Reactivity: Human

Immunohistochemical analysis of formalin fixed paraffin embedded human gastric cancer tissue with F2381 at 1:1000 dilution.,
Immunohistochemical analysis of formalin fixed paraffin embedded human gastric cancer tissue with F2381 at 1:1000 dilution.,

Usage Information

Dilution
IHC1:200 FCM1:20 - 1:100

Application
IHC, FCM

Reactivity
Human

Source
Mouse

Storage Buffer
PBS, pH 7.2+50% Glycerol+0.05% BSA+0.01% NaN3

Storage (from the date of receipt)
-20°C (avoid freeze-thaw cycles), 2 years

Positive Control	Human kidney; HEK293
Negative Control

Exprimental Methods

IHC
Experimental Protocol： Deparaffinization/Rehydration 1. Deparaffinize/hydrate sections: 2. Incubate sections in three washes of xylene for 5 min each. 3. Incubate sections in two washes of 100% ethanol for 10 min each. 4. Incubate sections in two washes of 95% ethanol for 10 min each. 5. Wash sections two times in dH2O for 5 min each. 6.Antigen retrieval: For Citrate: Heat slides in a microwave submersed in 1X citrate unmasking solution until boiling is initiated; continue with 10 min at a sub-boiling temperature (95°-98°C). Cool slides on bench top for 30 min. Staining 1. Wash sections in dH2O three times for 5 min each. 2. Incubate sections in 3% hydrogen peroxide for 10 min. 3. Wash sections in dH2O two times for 5 min each. 4. Wash sections in wash buffer for 5 min. 5. Block each section with 100–400 µl of blocking solution for 1 hr at room temperature. 6. Remove blocking solution and add 100–400 µl primary antibody diluent in to each section. Incubate overnight at 4°C. 7. Remove antibody solution and wash sections with wash buffer three times for 5 min each. 8. Cover section with 1–3 drops HRPas needed. Incubate in a humidified chamber for 30 min at room temperature. 9. Wash sections three times with wash buffer for 5 min each. 10. Add DAB Chromogen Concentrate to DAB Diluent and mix well before use. 11. Apply 100–400 µl DAB to each section and monitor closely. 1–10 min generally provides an acceptable staining intensity. 12. Immerse slides in dH2O. 13. If desired, counterstain sections with hematoxylin. 14. Wash sections in dH2O two times for 5 min each. 15. Dehydrate sections: Incubate sections in 95% ethanol two times for 10 sec each; Repeat in 100% ethanol, incubating sections two times for 10 sec each; Repeat in xylene, incubating sections two times for 10 sec each. 16. Mount sections with coverslips and mounting medium.

IHC

Experimental Protocol：

Deparaffinization/Rehydration

1. Deparaffinize/hydrate sections:

2. Incubate sections in three washes of xylene for 5 min each.

3. Incubate sections in two washes of 100% ethanol for 10 min each.

4. Incubate sections in two washes of 95% ethanol for 10 min each.

5. Wash sections two times in dH2O for 5 min each.

6.Antigen retrieval: For Citrate: Heat slides in a microwave submersed in 1X citrate unmasking solution until boiling is initiated; continue with 10 min at a sub-boiling temperature (95°-98°C). Cool slides on bench top for 30 min.

Staining

1. Wash sections in dH2O three times for 5 min each.

2. Incubate sections in 3% hydrogen peroxide for 10 min.

3. Wash sections in dH2O two times for 5 min each.

4. Wash sections in wash buffer for 5 min.

5. Block each section with 100–400 µl of blocking solution for 1 hr at room temperature.

6. Remove blocking solution and add 100–400 µl primary antibody diluent in to each section. Incubate overnight at 4°C.

7. Remove antibody solution and wash sections with wash buffer three times for 5 min each.

8. Cover section with 1–3 drops HRPas needed. Incubate in a humidified chamber for 30 min at room temperature.

9. Wash sections three times with wash buffer for 5 min each.

10. Add DAB Chromogen Concentrate to DAB Diluent and mix well before use.

11. Apply 100–400 µl DAB to each section and monitor closely. 1–10 min generally provides an acceptable staining intensity.

12. Immerse slides in dH2O.

13. If desired, counterstain sections with hematoxylin.

14. Wash sections in dH2O two times for 5 min each.

15. Dehydrate sections: Incubate sections in 95% ethanol two times for 10 sec each; Repeat in 100% ethanol, incubating sections two times for 10 sec each; Repeat in xylene, incubating sections two times for 10 sec each.

16. Mount sections with coverslips and mounting medium.

Datasheet & SDS

Biological Description

Specificity
Anti-NCX1 Mouse Antibody [C23J23] detects endogenous levels of total NCX1 protein.

Subcellular Location
Cell membrane, Membrane

Uniprot ID
P32418

Clone
C23J23

Synonym(s)
CNC, NCX1, SLC8A1, Sodium/calcium exchanger 1, Na(+)/Ca(2+)-exchange protein 1, Solute carrier family 8 member 1

Background
NCX1 (sodium–calcium exchanger 1) is a ubiquitously expressed integral membrane protein crucial for cellular Ca²⁺ homeostasis, particularly in the heart and brain, where it exchanges intracellular Ca²⁺ for extracellular Na⁺ to regulate excitation–contraction coupling and neuronal signaling. Structurally, NCX1 contains nine transmembrane segments with two α-repeat regions (involved in ion binding and transport) and a large intracellular loop between TMS 5–6 housing β-repeat regions that bind regulatory Ca²⁺. It exists in multiple splice variants with tissue-specific regulatory phenotypes, with NCX1.1 being the predominant cardiac isoform. NCX1 function is modulated by local factors, ion concentrations, lipids, and phosphorylation by kinases such as PKA and PKC within a macromolecular complex containing mAKAP, phosphatases (PP1, PP2A), and other regulatory proteins. In cardiomyocytes, this organization enables precise β-adrenergic regulation, though alterations such as hyperphosphorylation and attenuated responsiveness occur in heart failure, highlighting its importance in both physiological and pathological conditions.

Background

NCX1 (sodium–calcium exchanger 1) is a ubiquitously expressed integral membrane protein crucial for cellular Ca²⁺ homeostasis, particularly in the heart and brain, where it exchanges intracellular Ca²⁺ for extracellular Na⁺ to regulate excitation–contraction coupling and neuronal signaling. Structurally, NCX1 contains nine transmembrane segments with two α-repeat regions (involved in ion binding and transport) and a large intracellular loop between TMS 5–6 housing β-repeat regions that bind regulatory Ca²⁺. It exists in multiple splice variants with tissue-specific regulatory phenotypes, with NCX1.1 being the predominant cardiac isoform. NCX1 function is modulated by local factors, ion concentrations, lipids, and phosphorylation by kinases such as PKA and PKC within a macromolecular complex containing mAKAP, phosphatases (PP1, PP2A), and other regulatory proteins. In cardiomyocytes, this organization enables precise β-adrenergic regulation, though alterations such as hyperphosphorylation and attenuated responsiveness occur in heart failure, highlighting its importance in both physiological and pathological conditions.

References
https://pubmed.ncbi.nlm.nih.gov/12754202/ https://pubmed.ncbi.nlm.nih.gov/17303833/

Reference Table for Selecting PVDF Membrane Pore Size Specifications

Protein Size (kDa)	PVDF Transfer Membrane Pore Size (μm)
< 10	0.22
10-20	0.22
20-30	0.45
30-45	0.45
45-70	0.45
80-100	0.45
100-140	0.45
> 140	0.45

Tech Support

Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

Handling Instructions

Tel: +1-832-582-8158 Ext:3
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Protein Size (kDa)	Separating Gel Percentage
4-40	20%
12-45	15%
10-70	12.5%
15-100	10%
25-100	8%
60-210	5%