NCX1 Antibody [C23J23]

적용: 반응성:Human

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

사용 정보

희석
IHC1:200 FCM1:20 - 1:100

응용
IHC, FCM

반응성
Human

출처
Mouse Monoclonal Antibody

보관 완충액
PBS, pH 7.2+50% Glycerol+0.05% BSA+0.01% NaN3

보관 (수령일로부터)
-20°C (avoid freeze-thaw cycles), 2 years

양성 대조군	Human kidney; HEK293
음성 대조군

실험 방법

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

생물학적 설명

특이성
NCX1 Antibody [C23J23] detects endogenous levels of total NCX1 protein.

세포 내 위치
Cell membrane, Membrane

Uniprot ID
P32418

클론
C23J23

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

배경
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.

배경

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.

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

단백질 크기	PVDF 전이막 기공 크기
< 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

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단백질 크기	분리 젤 퍼센트
4-40	20%
12-45	15%
10-70	12.5%
15-100	10%
25-100	8%
60-210	5%