Hydriscreener NADPH Assay directly measures the NADPH produced by IDH1 Wildtype Enzyme. The NADPH measurements allow researchers to effectively determine the activity of the wildtype IDH1 enzyme. The assay provides a powerful tool for screening compound libraries for IDH1 modulators to help find new therapies for disease.
The kit includes the detection reagents required to measure activity. The IDH1 WT enzyme is not included in the inhibitor screening assay kit but can be purchased through BellBrook Labs.
Looking for a detailed protocol for performing the assay in your lab? Download the application note to get the assay up and running in your lab within hours.
Run your enzyme reaction, add Hydriscreener reagents, and read your plates. The IDH1 wildtype activity assay is compatible with 96, 384, and 1536-well formats.
Z’ measurements using optimized IDH1 wildtype reaction conditions indicate a robust assay. Robust data like this is vital for sizeable, high-throughput screens that are difficult to complete due to the massive sample quantity. Z’ shown here = 0.88.
The assay demonstrates linearity when raw data is converted to NADPH using a standard curve. We used 50 mM Tris pH 8.0, 5 mM MgCl2, and 0.01% Triton X-100 for the buffer, and ran with 250 µM Isocitrate, and 75 µM NADP at 30°C for 1 hour. The Detection Mix was added and incubated at room temperature for 60 minutes and read with a CLARIOstar plate reader.
Hydriscreener Assays are designed to screen compound libraries in a high-throughput format. Follow-up SAR can also be performed using the assay to determine inhibitor potency with ease. Assay conditions for these dose-response assays include 500 pM IDH1 wildtype enzyme, 50 mM Tris pH 8.0, 5 mM MgCl2, and 0.01% Triton X-100 for the buffer, and ran with 250 µM Isocitrate, and 75 µM NADP at 30°C for 1 hour. The enzyme reaction took place for 60 minutes at 30°C.
Interested in moving your program forward but don’t want to bring an assay in-house? Our scientists can help! BellBrook scientists will use their extensive biochemistry and enzymology expertise to work with you and accelerate your IDH1 program.
Please fill out the form below. We will respond quickly to get the conversation moving and learn how we can help. We keep things discrete, confidential, and professional.
| Hydriscreener NADPH Assay | ||||
|---|---|---|---|---|
| 1K | 10K | |||
| Component | Part # | Qty | Part # | Qty |
| Converting Enzyme | 2391 | 1 Bottle | 2391 | 10 Bottles |
| HydriFluor Solution | 2394 | 20 mL | 2394 | 5 x 20 mL |
| NADPH | 2392 | 4 x 1 Tube | 2392 | 40 x 1 Tube |
| NADP | 2393 | 2 x 1 Tube | 2393 | 20 x 1 Tube |
| Stop & Detect Buffer B, 10X | 2027 | 1 mL | 2032 | 10 mL |
| Component | Notes |
|---|---|
| Ultrapure Nuclease Free Water | Some deionized water systems are contaminated with enzymes that can degrade both nucleotide substrates and products, reducing assay performance. Use nuclease free water such as: Invitrogen Part # AM9930 |
| Enzyme | The Hydriscreener NADPH Assay is validated for use with purified dehydrogenase enzymes such as IDH1 Enzyme which is available from BellBrook as a stand-alone product (2395, 2396). |
| Assay Plates | You can purchase assay plates directly through BellBrook Labs. It is important to use assay plates that are entirely black with a nonbinding surface. We recommend Corning® 384-well plates (Cat.# 4514). The suggested plate has a square well top that enables easier robotic pipetting and a round bottom that allows good Z’ factors. It has a recommended working volume of 15–20 µL. |
Find the right resources to help you get to your solution faster.
Please fill out the form below. We will respond quickly to get the conversation moving and learn how we can help. We keep things discrete, confidential, and professional.
BellBrook Labs
1232 Fourier Drive, Suite 115
Madison, Wisconsin 53717 USA
(608) 443-2400
info@bellbrooklabs.com
