TNFSF9 anticorps (Tumor Necrosis Factor (Ligand) Superfamily, Member 9) (AA 200-250)

N° du produit ABIN705056

Détail du produit anti-TNFSF9 anticorps

Antigène:

TNFSF9

Épitope: AA 200-250

Reactivité: Humain

Hôte: Lapin

Clonalité: Polyclonal

Conjugué: Cet anticorp TNFSF9 est non-conjugé

Application: Immunofluorescence (Paraffin-embedded Sections) (IF (p)), Immunohistochemistry (Paraffin-embedded Sections) (IHC (p)), Western Blotting (WB)

Purification: Purified by Protein A.

Immunogène: KLH conjugated synthetic peptide derived from human TNFSF9

Isotype: IgG

Information d'application

Indications d'application: WB 1:100-1000
IHC-P 1:100-500
IF(IHC-P) 1:50-200

Restrictions: For Research Use only

Independent Validation (IHC)

Validation #029578 (Immunohistochemistry)

by: Immunohistochemistry Core, NYU Langone

No.: #029578

Date: 18.01.2014

Numéro du lot: 999892W

Application validée: Immunohistochemistry

Contrôle positif: Human liver

Contrôle négative: Human placental decidual cells

Conclusion: Signal was detected in positive control tissue and not in negative control tissue.

Validation Images

Figure 1: TNFSF9 staining of human liver (brown). Counterstain in blue.

Figure 2: Isotype control staining of human liver (brown). Counterstain in blue.

Figure 3: Secondary only staining of human liver (brown). Counterstain in blue.

Figure 4: TNFSF9 staining of human placenta (brown). Counterstain in blue.

Figure 5: Isotype control staining of human placenta (brown). Counterstain in blue.

Figure 6: Secondary only staining of human placenta (brown). Counterstain in blue.

Protocole

Anticorps primaire:

• Antibody: Tumor necrosis factor ligand superfamily member 9 (TNFSF9)
• Catalog number: ABIN705056
• Lot number: 999892W

Anticorps secondaire:

• Antibody: Biotinylated goat anti-rabbit/anti-mouse (Kit)
• Lot number: D05923BA

Full Protocol:

• Immunohistochemistry was performed on a Ventana NexES automated platform, instrument manufacturer specific reagents are italicized.
• 1. Slides were preheated in convection oven at 60°C for 30 minutes
• 2. Deparaffinization procedure: - 3 changes of Xylene, 5 minutes each - 3 changes of 100% Ethanol, 3 minutes each - 3 changes of 95% Ethanol, 3 minutes each - Rinsed in distilled water, 3 changes
• 1. Heat retrieval procedure - Slides retrieved in 10.0 mM Citrate, pH6.0 in a 1000W microwave oven (~100°C) for 15 minutes. - Slides were allowed to cool (in citrate) for 30 minutes. - Slides were washed x 3 in Distilled water
• 1. NexES instrument procedure, iVIEW DAB paraffin protocol (*abridged*): - Slide chamber warmed to 37°C
• 1. Slides rinsed with *reaction buffer* x 3
• 1. *iVIEW Inhibitor (H2O2)* applied and incubated for 4 minutes
• 1. Slides rinsed with *reaction buffer*
• 1. Antibody Application - Primary antibody diluted 1:250 in PBS (100 microliters applied/slide) - Ventana Isotype control applied neat - Slides incubated overnight at room temperature (~12 hours ~25°C)
• 1. Slides rinsed with *reaction buffer* x3
• 1. *iVIEW Biotinylated IgG* applied and incubated for 8 minutes
• 1. Slides rinsed with *reaction buffer*
• 1. *iVIEW Streptavidin-Horseradish Peroxidase* applied and incubated for 8 minutes
• 1. Slides rinsed with *reaction buffer*
• 1. *iVIEW DAB/H2O2* applied and incubated for 8 minutes
• 1. Slides rinsed with *reaction buffer*
• 1. *iVIEW Copper* applied and incubated for 4 minutes
• 1. Slides rinsed with *reaction buffer*
• 1. Slides washed in Dawn Detergent/tap water
• 1. Counterstain Procedure - Hematoxylin (Leica 560 MX) 30 seconds - Slides washed in tap water, 1 minute - Decolorized (10% Acetic Acid in 70% ethanol), 1 minute - Slides washed in tap water, 1 minute - Bluing (Austin Clear Ammonia), 1 minute - Slides washed in tap water, 1 minute
• 1. Dehydration/coverslipping procedure: - 3 changes of 95% Ethanol, 3 minutes each - 3 changes of 100% Ethanol, 3 minutes each - 3 changes of Xylene, 5 minutes each - Mounted with Permount
• 1. Imaging: Leica SCN 400F Whole Slide Scanner with Digital Image Hub and Leica Slidepath software

Notes:

• Deviations from protocol/procedure supplied by manufacturer:
• Step 1: Heated tissue 60°C for 30 minutes; manufacturer heats for 45 minutes.
• Step 2: No ethanol wash was performed during deparaffinization; manufacturer includes 1 wash of 80% ethanol for 3 minutes.
• Step 3.1: Slides heated for 15 minutes; manufacturer provides a range of 15-20 minutes.
• Step 3.2: Slides cooled for 30 minutes; manufacturer cools for 20 minutes.
• Step 4: Italicized reagents and incubation time are fixed instrument parameters.
• Step 5: Secondary species-specific serum block not used; manufacturer blocks with 5% normal goat serum for 2 hours.
• Step 8.1: Antibody diluted in PBS at 1:250; manufacture recommends dilution range (1:100-1:500). No primary antibody diluent recommended.
• Step 8.2.1: Primary antibody incubated at room temperature overnight; manufacturer incubates overnight 4°C with agitation.
• Tissue Interpretation (limited): • TNFSF9: Under the staining parameters described above, placenta was weakly cytoplasmic positive (trophoblastic and decidua cells). Liver hepatocytes were weakly stained. Substantial signal detected in limited number of other tissues, including kidney, prostate, and skin.
• • I-NC (Isotype negative control): No signal detected
• • B-NC (Blank negative control): No signal detected
• Signal Localization: - Signal to noise was adequate with predominately cytoplasmic subcellular localization observed. Nuclear staining was rare and no distinct membrane signal was detected.

Stockage

Format: Liquid

Concentration: 1 μg/μL

Buffer: Aqueous buffered solution containing 1 % BSA, 50 % glycerol and 0.09 % sodium azide.

Agent conservateur: Sodium azide

Précaution d'utilisation: This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE, which should be handled by trained staff only.

Stock: -20 °C

Stockage commentaire: Store at -20°C

Date de péremption: 12 months

Détails sur TNFSF9

Antigène:

TNFSF9

Autre désignation: Tnfsf9/Cd137l (TNFSF9 Antibody Extrait)

Sujet:

Cytokine that binds to TNFRSF9. Induces the proliferation of activated peripheral blood T-cells. May have a role in activation-induced cell death (AICD). May play a role in cognate interactions between T-cells and B-cells/macrophages.

Subcellular location: Extracellular

Synonyms: CD137L, 4-1BB-L, Tumor necrosis factor ligand superfamily member 9, 4-1BB ligand, 4-1BBL, TNFSF9

ID gène: 8744

UniProt: P41273

Pathways: Activated T Cell Proliferation, Cancer Immune Checkpoints