UA-Glo® 3D Cell Viability Assay

The 3D Cell Viability Assay Kit is used to quantitatively detect the ATP content in cells of 3D models, and the ATP content is proportional to the number of viable 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 3D model cell proliferation detection and compound screening.

Luciferase, luciferin, and buffer are mixed and filled into 10 ml or 100 ml brown bottles, with specifications as follows:

1. Cell Preparation
1) Construction of 3D cell models: Construct relevant 3D cell models according to experimental requirements, such as 3D cell sphere models.
2) If the experiment is to determine the effect of compounds on 3D cell models, add the test compound at an appropriate concentration to the wells of the cell plate. The concentration of organic solvents in the culture medium should be kept below 2%. Continue culturing for an appropriate time according to experimental requirements.

2. Cell Viability Assay
1) Take out the 3D cell viability assay reagent [Note 1], equilibrate to room temperature (22℃-25℃) [Note 2], and mix gently by shaking.
2) Take out the cell culture plate to be tested and equilibrate to room temperature (22℃-25℃). The 3D cell viability assay needs to be performed in a white assay plate. If the cell culture plate to be tested is a transparent or black assay plate, the 3D model needs to be transferred to a white assay plate before testing.
3) Add 50µL of 3D cell viability assay reagent to 100µL of 3D model cells in a 96-well plate, or 10µL of reagent to 20µL of 3D model cells in a 384-well plate [Note 3].
4) Vigorously shake the assay plate for 5 minutes to fully lyse the cells [Note 4], and place it in the dark to equilibrate the reaction for 25 minutes.
5) Read the fluorescent signal on a fluorescence microplate reader [Note 5].

1) After the first use of the reagent, it should be aliquoted and stored in the dark at -20°C or below to ensure the stability of the reagent. The 3D Cell Viability Assay Reagent, after 5 repeated freeze-thaw cycles, shows a signal intensity reduction of <10% with no loss of function. When left at room temperature (22°C) for 8 hours or at 4°C for 72 hours, the signal intensity reduction is <10% with no loss of function.
2) The luciferase reaction in the 3D Cell Viability Assay Reagent is sensitive to temperature changes. The reagent and the assay plate need to be equilibrated to room temperature (22°C-25°C), and the temperature should be kept constant (±1°C) during the test.
3) It is not recommended to arbitrarily change the amount of reaction reagents without strict verification. The volume ratio of cell culture medium to assay reagent should be 2:1.
4) It is very necessary to vigorously shake the assay plate to fully lyse the cells, and at the same time, care should be taken to prevent liquid in the wells from spilling.
5) The 3D Cell Viability Assay Reagent has different signal decay rates for 3D models of different cell types, with a signal half-life between 1.5h and 3h. It is recommended that the plate reading time does not exceed 1h-2h.
6) This product is for research use only.