TNKS2 (PARP5B) Colorimetric Assay Kit
The TNKS2 Colorimetric Assay Kit is designed to measure the activity of Tankyrase 2 (TNKS2, also known as PARP5B) for screening and profiling applications. TNKS2 is known to catalyze the NAD-dependent ADP-ribosylation of histones. The key to the TNKS2 Colorimetric Assay Kit is the biotinylated NAD+. First, histone proteins are coated on a 96-well plate. Next, a biotinylated NAD+ mix (termed PARP Substrate Mixture) is incubated with the TNKS2 enzyme. Finally, the plate is treated with streptavidin-HRP followed by addition of the colorimetric HRP substrate to produce color that can be measured using a UV/Vis spectrophotometer microplate reader.
Figure 1. TNKS2 (PARP5B) Colorimetric Assay Kit schematic
*NOTE: As of March 2023, this protocol has been re-optimized for performance. Previous versions of this kit are available upon request.
- DTT (10 mM in water, prepared fresh)
- 1x PBS (phosphate buffer saline)
- PBST: 1x PBS, containing 0.05% Tween-20
- UV/Vis spectrophotometer microplate reader capable of reading absorbance at 450 nm*
- 2 M sulfuric acid (aqueous)
- Adjustable micropipettor and sterile tips
- Rotating or rocker platform
*Alternately, a spectrophotometer reading at 650 nm may be used, but sensitivity of the assay will be greatly reduced.
Catalog # | Name | Amount | Storage | |
80515 | TNKS2* (Tankyrase 2 (PARP5B) [849-1166], GST-Tag) | 1 µg | -80°C | Avoid multiple freeze/ thaw cycles |
52029 | 5x Histone Mixture | 1 ml | -80°C | |
78371 | PARP Substrate Mixture 2 | 250 µl | -80°C | |
80602 | 10x PARP Assay Buffer | 1 ml | -20°C | |
79743 | Blocking Buffer 3 | 25 ml | +4°C | |
80611 | Streptavidin-HRP | 100 µl | +4°C | |
79651 | Colorimetric HRP Substrate | 10 ml | +4°C | |
79964 | 96-well transparent plate | 1 | Room Temp |
*The concentration of the protein is lot-specific and will be indicated on the tube. Excess material has been given for ease of retrieval.