Collagen IV Antibody [E5G18]

Catalog No.: F1667

Application: Reactivity: Human

Immunohistochemical analysis of formalin fixed paraffin embedded human placenta tissue with F1667 at 1:125 dilution.

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代表番号: 045-509-1970|電子メール：sales@selleck.co.jp

使用情報

Dilution
IHC1:100-1:250

Application
IHC

Source
Mouse Monoclonal Antibody

Reactivity
Human

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

ポジティブコントロール	Human ovary; Human tongue
ネガティブコントロール

プロトコール

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

生物学的記述

Specificity
Collagen IV Antibody [E5G18] detects endogenous levels of total Collagen IV protein.

タンパク質の局在
基底膜、細胞外マトリックス、細胞外環境

Uniprot ID
P02462

Clone
E5G18

Synonym(s)
Collagen alpha-1(IV) chain, COL4A1

Background
Collagen IV is the primary structural collagen of basement membranes, specialized extracellular matrices that provide mechanical support, regulate cell adhesion, and organize other matrix components. It is a heterotrimer composed of combinations of six α-chains (α1–α6) encoded by distinct genes, typically arranged as two α1 and one α2 chains in most tissues, assembled intracellularly into triple-helical protomers. These protomers are secreted and crosslinked via their NC1 and 7S domains to form a stable, sheet-like network that tethers laminins, proteoglycans, and growth factors. Collagen IV is widely expressed in epithelial and endothelial basement membranes, with tissue-specific isoform composition, and plays essential roles in development, tissue integrity, cell migration, and signaling through integrins such as α1β1 and α2β1. Defects or aberrant crosslinking of collagen IV contribute to diverse pathologies, including Alport syndrome, Goodpasture’s disease, diabetic nephropathy, and certain dermatological disorders.

Background

Collagen IV is the primary structural collagen of basement membranes, specialized extracellular matrices that provide mechanical support, regulate cell adhesion, and organize other matrix components. It is a heterotrimer composed of combinations of six α-chains (α1–α6) encoded by distinct genes, typically arranged as two α1 and one α2 chains in most tissues, assembled intracellularly into triple-helical protomers. These protomers are secreted and crosslinked via their NC1 and 7S domains to form a stable, sheet-like network that tethers laminins, proteoglycans, and growth factors. Collagen IV is widely expressed in epithelial and endothelial basement membranes, with tissue-specific isoform composition, and plays essential roles in development, tissue integrity, cell migration, and signaling through integrins such as α1β1 and α2β1. Defects or aberrant crosslinking of collagen IV contribute to diverse pathologies, including Alport syndrome, Goodpasture’s disease, diabetic nephropathy, and certain dermatological disorders.

References
[1] Boudko SP, et al. Methods Cell Biol. 2018;143:171-185.

PVDFメンブレン孔径参考表

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

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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%