UA-Glo® Luminescent Cell Viability Assay

The UA-Glo Homogeneous Ready-to-Use Cell Viability Assay Kit is used to quantitatively detect the content of ATP in living cells, thereby indirectly determining the number of living cells. This reagent features high signal-to-noise ratio, good repeatability, and excellent stability. The ready-to-use formula eliminates the experimental steps of first lysing and then detecting, thereby further reducing errors caused by frequent sample addition. Moreover, its stable glow signal makes this product particularly suitable for high-throughput tumor cell proliferation detection and compound screening.

The product is filled into 10 ml or 100 ml brown bottles after mixing luciferase, luciferin, and buffer solution.

1.Cell Preparation

1) Seed the cells to be tested at an appropriate density in a 96-well or 384-well cell culture plate. It is recommended to use a white plate.
2) Prepare the test compounds. Add the test compounds at appropriate concentrations to the wells of the cell plate. The concentration of organic solvents in the culture medium should be kept below 1-2%. Set up a control group without added compounds. Continue culturing for an appropriate time according to the project requirements.

2. Cell Viability Assay

1) Take out the UA-Glo Cell Viability Assay Reagent and equilibrate it at room temperature for 20 minutes. Gently shake to mix well.

2) Take out the cell culture plate to be tested and equilibrate it at room temperature for 20 minutes.

3) Add 50 µl of the assay reagent to 100 µl of cells in a 96-well plate, or 10 µl of the reagent to 20 µl of cells in a 384-well plate, gently oscillate for 2 minutes, and place in the dark for further lysis for 10 minutes.

4) Read the luminescent signal on a luminescence microplate reader.

1. Temperature: Unless otherwise specified, luciferase detection is performed at room temperature (22-25°C). Temperature has a significant impact on enzyme reactions.
2. Reagent dosage: It is not recommended to arbitrarily change the dosage of reaction reagents without strict verification.
3. Plate reading time: After adding the reagents, shake for 2 minutes, place in the dark for 10 minutes, and then read the plate. It is recommended to complete the plate reading within 2 hours after adding the reagents.
4. Inter-plate internal reference: If inter-plate data comparison is required, it is recommended to set up two wells of positive controls without added compounds in all reaction plates as inter-plate internal references. The readings of each plate should first be corrected (normalization) using the inter-plate internal reference readings, and the corrected data should then be subjected to downstream processing and analysis.
5. Reaction plate: The level of fluorescence reading may significantly interfere with the readings of adjacent wells, mainly due to the spillover of fluorescence signals. When necessary, opaque-bottom white plates or black plates can be used for experiments or verification.
6. Application scenarios: This product is a detection reagent for live cells. If used in in vitro reaction systems constructed with recombinant proteins, additional ATP needs to be provided.
7. Reagent mixing: It is not recommended to mix reagents from different batches, reagents that have been opened but not used up, and reagents with significantly different storage conditions.
8. Aliquot storage: Aliquot and store the reaction reagents according to the instructions to ensure the stability of the reagents.