TiterTACS™ Colorimetric Apoptosis Detection Kit

TUNEL assay cell type - Mouse endothelial cells

Experiment
TUNEL assay cell type - Mouse endothelial cells
Product
TiterTACS™ Colorimetric Apoptosis Detection Kit from Bio-Techne
Manufacturer
Bio-Techne

Protocol tips

Protocol tips
Add Proteinase K solution and incubate for 15 minutes at room temperature.

Add Labeling Reaction mix and incubate at 37°C for 1 hour.

Add TACS-Sapphire at room temperature..

Publication protocol

For ADAM12 augmentation and knockdown in vitro, plasmid containing full-length ADAM12 cDNA or shRNA targeting ADAM12 was transfected into HUVECs using a high-efficiency primary cell-transfection reagent (Cytofect EC transfection kit; Cell Applications), following the manufacturer's protocol. Briefly, cells were cultured to ∼75% confluence in a six-well plate. DNA plasmid (3 μg) was mixed with 2.5 μl Cytofect-2 and 2.5 μl polyethylene reagents and incubated at 37°C for 30 min. This plasmid DNA and transfection reagent complex mixture was added to ECs for 1 h and then replaced with ECGM with 10% serum. Cells were used in subsequent experiments, 24 h after transfection. EC proliferation, apoptosis assay, and in vitro angiogenesis were assessed, as described previously (16). Briefly, proliferation was assessed under baseline conditions (ECGM with 10% serum) using tetrazolium dye incorporation (BioVision, Milpitas, CA), 48 h post-transfection, following the manufacturer's protocol. Apoptosis was assessed in transfected cells, 24 h post-transfection, followed by 24 h exposure to simulated ischemia using the terminal deoxynucleotidyltransferase (TdT)-mediated 2′-deoxyuridine 5′-triphosphate nick end-labeling assay (TiterTACS; Trevigen, Gaithersburg, MD), following the manufacturer's protocol. Nuclease-treated wells were used as positive controls, whereas wells without terminal TdT were used as negative controls. For the angiogenesis assay, the cells were plated on growth factor-enriched matrigel (Catalog #356234; BD Biosciences), 48 h post-transfection, and grown in simulated ischemia for 6 h. Each condition was performed in quadruplicate, and representative pictures were taken from each well at 100× magnification. A blinded observer counted the number of complete tubes, which was expressed as tubes/mm2. All experiments described above were repeated at least three times.

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Discussion

Discussion

5 years ago

Author: Ercole Udinesi Italy

Are 10 week old samples in formalin still usable?

I have left my tumor samples in formalin for an extended period of time (around 10 weeks). Do you think I will be able to use them for TUNEL assay and get results? Thank you for your help in advance

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Papers

Check out relevant papers found by Labettor's AI that are relevant for performing TUNEL assay cell type - Mouse endothelial cells using TiterTACS™ Colorimetric Apoptosis Detection Kit from Bio-Techne.

Paper title
ADAM12: a genetic modifier of preclinical peripheral arterial disease
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Manufacturer protocol

Download the product protocol from Bio-Techne for TiterTACS™ Colorimetric Apoptosis Detection Kit below.

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