Probe Reports from the NIH Molecular Libraries Program [Internet].
Bethesda (MD): National Center for Biotechnology Information (US); 2010-.
NCBI Bookshelf. A service of the National Library of Medicine, National Institutes of Health.
Table 6Stepwise protocol of the LDHA counterscreen in 1536-well format
| Sequence | Parameter | Value | Description |
|---|---|---|---|
| 1 | Reagent | 3 μL | LDHA enzyme in assay buffer . Col 1,2-5-48 Assay Buffer. Col 3-4 |
| 2 | Compound | 23 nL | Compound Library |
| 3 | Time | 15 min | Incubation with compounds at room temperature |
| 4 | Reagent | 1 μL | NADH (60 μM [final]) and sodium pyruvate (160 μM [final]) substrate mix |
| 5 | Time | 8 min | Incubation with substrate at room temperature |
| 6 | Reagent | 1 μL | Diaphorase / Resazurin |
| 7 | Time | 10 min | RT incubation |
| 8 | Detector | Fluorescence | Final read (ViewLux; ex 525, em 598, 1 second exposure, Bodipy Mirror) |