Collagen IV Antikörper

Produktnummer ABIN5596835

Produktdetails anti-Collagen IV Antikörper

Target: Collagen IV (COL4) (Collagen, Type IV (COL4))

Reaktivität: Human, Rind (Kuh), Säugetier

Wirt: Kaninchen

Klonalität: Polyklonal

Konjugat: Dieser Collagen IV Antikörper ist unkonjugiert

Applikation: Western Blotting (WB), ELISA, Immunohistochemistry (IHC), Immunoprecipitation (IP)

Kreuzreaktivität (Details): Typically negligible cross-reactivity against other types of collagens was detected by ELISA against purified standards. Some class-specific anti-collagens may be specific for three-dimensional epitopes which may result in diminished reactivity with denatured collagen or formalin-fixed, paraffin embedded tissues. This antibody reacts with most mammalian Type IV collagens and has negligible cross-reactivity with Type I, II, III, V or VI collagens. Non-specific cross-reaction of anti-collagen antibodies with other human serum proteins or non-collagen extracellular matrix proteins is negligible.

Aufreinigung: Anti-Collagen Type IV has been prepared by immunoaffinity chromatography using immobilized antigens followed by extensive cross-adsorption against other collagens, human serum proteins and non-collagen extracellular matrix proteins to remove any unwanted specificities.

Immunogen:

Immunogen: Collagen Type IV from human and bovine placenta

Immunogen Type: Native Protein

Isotyp: IgG

Produktspezifische Information:

Für welche Zwecke kann der Antikörper ABIN5596835 gegen Kollagen 4 verwendet werden? Dieser polyklonale Kollagen 4-Antikörper ist unkonjugiert und wurde für ELISA, IHC, IP und Western Blotting validiert. Als Immunogen für die Herstellung des COL4-Antikörpers wurde natives Kollagen Typ 4 aus humaner und boviner Plazenta verwendet.

Welche Validierungsdaten liegen für diesen Kollagen-4-Antikörper vor? Der Antikörper wurde bisher in 8 Pubmed-Publikationen referenziert und zeichnet sich durch hohe Zuverlässigkeit aus. Die Referenzen sind unten aufgeführt. 9 Bilder zeigen die Leistung des Produkts in verschiedenen Anwendungen.

Was ist die Funktion von Kollagen 4? Kollagen Typ 4 ist die Hauptstrukturkomponente der glomerulären Basalmembranen und bildet zusammen mit Lamininen, Proteoglykanen und Entactin/Nidogen ein "Hühnerdraht"-Geflecht. (Uniprot). Das Kollagen-4-Protein besteht aus einer aminoterminalen 7S-Domäne, einer kollagenen Domäne, die eine Dreifach-Helix bildet, und einer nicht-kollagenen Domäne am Carboxy-Terminus. Es fungiert als Teil eines Heterotrimers und interagiert mit anderen extrazellulären Matrixkomponenten wie Perlecanen, Proteoglycanen und Lamininen (NCBI)

Anwendungsinformationen

Applikationshinweise:

Immunohistochemistry Dilution: 1:50 - 1:200

Application Note: Anti-Collagen antibodies have been used for indirect trapping ELISA for quantitation of antigen in serum using a standard curve, immunoprecipitation, native (non-denaturing, non-dissociating) PAGE, immunohistochemistry, and western blotting for highly sensitive qualitative analysis.

Western Blot Dilution: 1:1,000 - 1:10,000

Immunoprecipitation Dilution: 1:100

ELISA Dilution: 1:5,000 - 1:50,000

Beschränkungen: Nur für Forschungszwecke einsetzbar

Independent Validation (IHC)

Validierung #300049 (Immunohistochemistry)

by: MS Validated Antibodies

No.: #300049

Datum: 07.08.2023

Antigen: COL4

Chargennummer: 48011

Validierte Anwendung: Immunohistochemistry

Positivkontrolle:

Human TMA

Recombinant rabbit anti-COL4 antibody (MSVA-704R)

Bewertung:

Passed. Staining of collagen type IV using ABIN5596835 is consistent with the expected staining pattern.

Validierungsbilder

IHC staining of lung tissue with anti-collagen IV antibody ABIN5596835 diluted 1:200 shows staining of basement membranes and vessels.

IHC staining skeletal muscle tissue with anti-collagen IV antibody ABIN5596835 diluted 1:200 shows collagen IV fibres surrounding skeletal muscle cells.

IHC staining of renal oncocytoma tissue with anti-collagen IV antibody ABIN5596835 diluted 1:300 shows dense collagen IV-positive membranes surrounding tumor cell nests.

Protokoll

Primärantikörper: ABIN5596835

Sekundärantikörper: EnVision Polymer-HRP mouse/rabbit Kit, Dako REAL, K5007

Full Protocol:

• Slide preparation
• • Mount 2,5 µm FFPE tissue sections on superfrost slides.
  • Deparaffinize tissue sections 3x 5 min in xylene.
  • Rehydrate tissue sections in a descending ethanol series for 1 min each 100%, 96%, and 80% ethanol.
  • Rinse tissue sections for 5 min in TBST buffer (DAKO, K8000).
• Epitope retrieval
• • Autoclave tissue sections for 5 min at 121 °C in 1x Tris-EDTA-citrate buffer pH7.8 (20x Tris-EDTA-citrate buffer stock solution: 5 g Trizma base (Sigma-Aldrich, T1503), 10 g EDTA (Merck, 1.08418), 6.4g tri-sodium citrate (Sigma-Aldrich, C0909), adjust to pH 7.8 using HCL 1 M, ad 1 L with dH₂O).
  • Rinse tissue sections for 5 min in TBST buffer.
• Peroxidase blocking
• • Incubate tissue sections for 10 min in Peroxidase-Blocking Solution (Dako REAL, S2023).
  • Rinse tissue sections 2x for 5 min in TBST buffer.
• Antibody incubation
• • Dilute primary rabbit anti-collagen IV antibody (antibodies-online, ABIN5596835, lot 48011) diluted 1:100, 1:200, or 1:300 in antibody diluent (Dako REAL, S2022).
  • Cover tissue section with 100-200 µl diluted antibody.
  • Incubate tissue sections for 1 h at 37 °C in a moist chamber.
  • Rinse tissue sections for 5 min in TBST buffer.
  • Apply EnVision Polymer-HRP mouse/rabbit Kit (Dako REAL, K5007) according to manufacturer’s recommendation.
  • Rinse tissue sections 2x for 5 min in TBST buffer.
• Staining
• • Cover slides for 10 min with DAB-Chromogen (EnVision Polymer-HRP mouse/rabbit Kit, Dako REAL, K5007).
  • Wash slides thoroughly with dH₂O.
  • Counterstain for 15 sec with Hematoxylin (Mayers Hematoxylin: 200ml ddH₂O, 0,2g Hematoxylin (Serva, 24420.02), 10 g aluminium potassium sulfate dodecahydrate (Merck, 1.01047), 0,04 g sodium iodate (Merck, 1.06525), 10 g chloral hydrate (Sigma-Aldrich, 15307)).
  • Develop for 15 sec in H₂O.
  • Dehydrate tissue sections in an ascending ethanol series for 1 min each 80%, 96%, 100% ethanol.
  • Wash tissue sectiona 3x 5 min in xylene.
  • Apply mounting medium and coverslips.
• Image acquisition
• • o Acquire images using a Galileo TMAtic (ISENET).

Anmerkungen:

• For antibody comparison an antibody test TMA was used that contained 80 normal tissues from 21 different organs and 95 neoplastic tissues from 18 different tumor types. Despite some cytoplasmic and stromal background ABIN5596819 identified identical structures as the reference antibody MSVA-704R. An additional occasional staining of nuclei which was only seen by ABIN5596819 must be considered a cross-reactivity.

• According to the rather ubiquitous nature of collagen IV expression, orthogonal validation is not optimally suited for the validation of collagen I antibodies. In agreement with RNA screening studies, including the Human Protein Atlas (HPA) RNA-seq tissue dataset, the FANTOM5 project, and the Genotype-Tissue Expression (GTEx) project (all summarized in https://www.proteinatlas.org/ENSG00000134871-COL4A2/tissue), collagen IV staining by ABIN5596819 was particularly high in the placenta, colon, smooth muscle, and the lung. These are among the tissues with the highest recorded RNA expression.

Handhabung

Format: Liquid

Konzentration: 1.0 mg/mL

Buffer:

Buffer: 0.125 M Sodium Borate, 0.075 M Sodium Chloride, 0.005 M EDTA, pH 8.0

Stabilizer: None

Konservierungsmittel: Sodium azide

Vorsichtsmaßnahmen: This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.

Lagerung: 4 °C,-20 °C

Informationen zur Lagerung: Store vial at 4° C prior to opening. This product is stable at 4° C as an undiluted liquid. Dilute only prior to immediate use. For extended storage, mix with an equal volume of glycerol, aliquot contents and freeze at -20° C or below. Avoid cycles of freezing and thawing.

Haltbarkeit: 12 months

Referenzen für anti-Collagen IV Antikörper (ABIN5596835)

Ida, Hikage, Itoh, Ida, Ohguro: "Prostaglandin F2α agonist-induced suppression of 3T3-L1 cell adipogenesis affects spatial formation of extra-cellular matrix." in: Scientific reports, Vol. 10, Issue 1, pp. 7958, (2020) (PubMed).

Kumar Gupta, Sarkar, Wertheim, Pan, Carroll, Oxburgh: "Asynchronous mixing of kidney progenitor cells potentiates nephrogenesis in organoids." in: Communications biology, Vol. 3, Issue 1, pp. 231, (2020) (PubMed).

Hwang, Huang, Burwell, Peterson, Connor, Weiss, Yu, Li: "In Situ Imaging of Tissue Remodeling with Collagen Hybridizing Peptides." in: ACS nano, Vol. 11, Issue 10, pp. 9825-9835, (2019) (PubMed).

Tanaka, Ng, Yang Yu, Casco-Robles, Maruo, Tsonis, Chiba: "A developmentally regulated switch from stem cells to dedifferentiation for limb muscle regeneration in newts." in: Nature communications, Vol. 7, pp. 11069, (2016) (PubMed).

Fetting, Guay, Karolak, Iozzo, Adams, Maridas, Brown, Oxburgh: "FOXD1 promotes nephron progenitor differentiation by repressing decorin in the embryonic kidney." in: Development (Cambridge, England), Vol. 141, Issue 1, pp. 17-27, (2014) (PubMed).

Seifert, Kiama, Seifert, Goheen, Palmer, Maden: "Skin shedding and tissue regeneration in African spiny mice (Acomys)." in: Nature, Vol. 489, Issue 7417, pp. 561-5, (2012) (PubMed).

Seifert, Monaghan, Voss, Maden: "Skin regeneration in adult axolotls: a blueprint for scar-free healing in vertebrates." in: PLoS ONE, Vol. 7, Issue 4, pp. e32875, (2012) (PubMed).

Pinheiro, Ferreira, Kitten, da Silveira, da Silva, Santos, Gava, Castro, Magalhães, da Mota, Botelho-Santos, Bader, Alenina, Santos, Simoes E Silva: "Genetic deletion of the angiotensin-(1-7) receptor Mas leads to glomerular hyperfiltration and microalbuminuria." in: Kidney international, Vol. 75, Issue 11, pp. 1184-1193, (2009) (PubMed).

Details zu Collagen IV

Target: Collagen IV (COL4) (Collagen, Type IV (COL4))

Andere Bezeichnung: Collagen Type IV (COL4 Produkte)

Synonyme: collagen, type IV antikoerper, Let-2 antikoerper

Hintergrund:

Synonyms: Arresten antibody, Canstatin antibody, Collagen Of Basement Membrane Alpha 1 Chain antibody

Background: Rockland produces highly active antibodies and conjugates to collagens. Collagens are highly conserved throughout evolution and are characterized by an uninterrupted ''Glycine-X-Y'' triplet repeat that is a necessary part of the triple helical structure. For these reasons, it is often extremely difficult to generate antibodies with specificities to collagens. The development of 'type' specific antibodies is dependent on NON-DENATURED three-dimensional epitopes. Rockland extensively purifies collagens for immunization from human and bovine placenta and cartilage by limited pepsin digestion and selective salt precipitation. This preparation results in a native conformation of the protein. Antibodies are isolated from rabbit antiserum and are extensively cross-adsorbed by immunoaffinity purification to produce 'type' specific antibodies. Greatly diminished reactivity and selectivity of these antibodies will result if denaturing and reducing conditions are used for SDS-PAGE and immunoblotting.

Gene Name: COL4A1

Gen-ID: 1282

UniProt: P02462