| Usage | 1. Preparation of organoid:
Use 96-well plate suitable for chemiluminescence detection (bottom and lid are transparent, well wall is impermeable, recommended item number: abs7242), inoculate 5ul-10ul organoid matrigel suspension per well, put it in the incubator for more than half an hour, add 100ul organoid culture medium after coagulation (if 384-well plate is used, inoculate 2.5 μl-4μl organoid matrigel suspension per well, depending on different types of 384-well plate), and ensure that the number of cells per well is within 50,000 (if 384-well plate is used). If desired, drugs can be added to treat organoids. In addition, if necessary, the concentration gradient of organoids can also be set to subsequently determine the effectiveness of the use of the kit.
2. Reagent preparation:
Add 1 tube of freeze-dried powder to 1 tube of 10ml buffer, mix well and protect from light for later use.
Note: Use as soon as possible after dissolving,-80 ℃, protected from light, valid for one month
(1) Dissolve the Organoid ATP luminescence detection reagent, and dispense the reagent appropriately if necessary.
(2) According to the amount of 100 μl per well of 96-well plate (25 μl per well of 384-well plate), take an appropriate amount of Organoid ATP luminescence detection reagent, and equilibrate to room temperature.
3. Organoid viability detection:
(1) Take out the organoid culture plate and equilibrate it at room temperature for 10 minutes (usually not more than 30 minutes).
(2) The organoid medium in each well of the 96-well plate was removed, and 100 μl of Organoid ATP luminescence detection reagent was added to each well (25 μl per well of the 384-well plate).
Note: No removal of Matrigel is required.
(3) Violent oscillation at room temperature (microplate constant temperature oscillator, recommended item number: abs72034) for 5 minutes to fully lyse the organoids.
(4) Incubate at room temperature (about 25 °C) for 10 minutes to stabilize the luminescence signal.
(5) Use a multifunctional microplate reader with the function of detecting chemiluminescence for chemiluminescence detection. Please set the corresponding parameters according to the requirements of the instrument. The detection time of each hole is generally 0.25-1 second or longer. The specific adjustment needs to be made according to the detection sensitivity of the instrument.
(6) Calculate the relative viability of organoids directly from the chemiluminescence readings, or calculate the amount of ATP according to the ATP standard curve to calculate the relative viability of organoids.
Note: The detection effect varies depending on the type of organoid. For some organoids with particularly high ATP content, there may not be a linear correlation after the number of cells reaches 50,000, but the chemiluminescence reading will continue to increase. |
| Synonym | ATP; ATP detection; CTG; ATPlite; Luminescent cell viability detection kit |
| Description | This product is an ATP detection kit specially developed for the detection of organoid activity. Its luminescence technology principle is that luciferase uses luciferin, adenosine triphosphate (ATP) and O2Is the substrate, in Mg2+When present, it can convert chemical energy into light energy. ATP is not only an essential substrate for luciferase catalyzed luminescence, but also an energy source for all biological life activities. In the luminescence reaction catalyzed by luciferase, the concentration of ATP has a linear relationship with the luminescence intensity within a certain concentration range.
This kit uses bioluminescent method, uses Firefly luciferase (Firefly luciferase) to catalyze the conversion of substrate-luciferin, and efficiently uses the energy of ATP to emit photons. The luminescence signal is proportional to the amount of ATP present, and ATP is directly proportional to the number of cells present in the organoid.
Designed for multi-well plates, this kit is ideal for automated high-throughput screening (HTS), organoid proliferation and toxicity analysis. The homogenization detection step is to directly add a single reagent to the well plate containing organoid culture medium without removing the matrigel.
The "spike-mix-detect" protocol of homogeneous detection allows organoid lysis and the luminescence signal produced to be proportional to the amount of ATP present, which is directly proportional to the number of cells in the organoid. The unique homogeneous detection protocol avoids the errors that may be introduced by ATP detection methods that require multiple steps.
Product composition:
| Components | 10mL | 100mL | Preservation method | | Organoid ATP Fluorescent dye | 1 stick | 10 sticks | 2-8 ℃, protected from light | | Organoid ATP dye buffer | 10mL | 10 × 10 mL | 2-8 ℃, protected from light |
Note: The product is recommended for immediate use
Product Features:
(1) Simplified organoid activity detection steps: the homogeneous "sample addition-mixing-detection" scheme reduces the operation steps required for other similar detections.
(2) Less amount of organoids: the number of cells below the low detection limit of commonly used colorimetric and fluorescent methods can be accurately detected. The number of cells required per detection reaction is reduced.
(3) Obtain the results quickly: the data can be obtained 10 minutes after adding the reagent.
(4) You can choose your own detection scheme: it can be used for many types of multi-well plate operations. The data can be recorded with a luminescence detector or CCD imaging equipment.
(5) The culture plate can be continuously processed: the luminescence signal is stable, and the samples can be processed in batches. |
| General Notes | 1. The activity of luciferase is sensitive to temperature, so the organoids and detection reagents need to be balanced to room temperature before the reaction before measurement. Please mix the test reagents well before use.
2. The detection reagent of this kit contains luciferase, and repeated freezing and thawing will cause its gradual inactivation. In order to achieve good use effect, it can be appropriately packaged and stored after the first dissolution, but it should be noted that the packaged container should not be contaminated by ATP.
3. When the solvent content of the drug to be tested is high, it may interfere with the luciferase reaction, thus affecting the chemiluminescence signal. Solvent interference can be eliminated by setting organoid culture medium control wells containing solvent.
4. White or black 96-well plates or 384-well plates suitable for organoid culture must be used for testing. If a plain clear 96-well plate or 384-well plate is used, adjacent wells will interfere with each other.
5. This product is limited to scientific research by professionals. It shall not be used for clinical diagnosis or treatment, food or medicine, or stored in ordinary houses.
6. For your safety and health, please wear a lab coat and disposable gloves. |
| Storage Temp. | 2-8 ℃, protected from light, valid for 12 months. |
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