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G1609-100T / 500T Calcein AM Cell Viability Detection Kit (CCK-F)

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G1609-100T / 500T Calcein AM Cell Viability Detection Kit (CCK-F)

Calcein AM is based on Calcein (calcein), introducing an acetomethoxymethyl (AM) group. The AM group not only masks the molecular part of Calcein chelating calcium but also enhances its hydrophobicity, so it can It allows Calcein AM to easily penetrate the membrane of living cells and is cleaved into Calcein by the endogenous esterase of the cell. Calcein that has lost the AM group cannot easily pass through the cell membrane and is thus retained in the cell; in addition, the part of the calcium chelating molecule is exposed, allowing the Calcein probe to bind to calcium ions in living cells and emit strong green fluorescence. The lack of esterase in dead cells does not allow Calcein AM to be hydrolyzed into Calcein, so dead cells cannot be labeled. Calcein AM has low cytotoxicity and has little effect on cell functions, such as cell proliferation or lymphocyte chemotaxis. It is a fluorescent probe very suitable for staining live cells.

 

This product Calcein AM cell viability detection kit is also called Cell Fluorescence Counting Kit (CCK-F), which uses Calcein AM fluorescent probe to label live cells to evaluate cell viability, toxicity, and proliferation. Detection kit. After optimization and debugging, this kit not only has high detection sensitivity and wide linear range, but also only needs a short time of incubation to complete the detection of cells. The operation is simple, does not require radioisotope labeling, and does not require steps such as crystal dissolution, which can reduce errors caused by experimental operations and improve accuracy and repeatability.

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Product Variation:

Item code Description Volume
G1609-100T Calcein AM Cell Viability Detection Kit (CCK-F) 100T
G1609-500T 500T

 

Package Contents:

Item code Description G1609-100T G1609-500T
G1609-1 Calcein AM Solution 10µL 50µL
G1609-2 CCK-FD Detection Buffer 10mL 50mL
  User Manual 1pc 1pc

Technical Specification

Storage and transportation

Storage and transportation at room temperature, valid for 2 years.
 

Operation Steps:

  1. Preparation of Calcein AM detection working solution:
    According to the usage amount of 100 μL per well of 96-well plate, refer to the following table, mix Calcein AM solution and CCK-F detection buffer in proportion to prepare Calcein AM detection working solution.

    Component

    10pcs Samples

    50pcs Samples

    100pcs Samples

    Calcein AM Solution

    1 μL

    5 μL

    10 μL

    CCK-F Detection Buffer

    1 mL

    5 mL

    10 mL

     
  2. Adherent cell detection:
    The following steps mainly correspond to the 96-well plate detection scheme, and other well plate systems need to be adjusted according to the situation.
    a. Cell seeding plate: evenly seed the cells in a 96-well plate at a certain density, and apply drugs or other pre-treatments to them (the seeding density is determined by factors such as cell size, growth rate, etc.);
    b. (Optional) Cell washing: aspirate the original culture medium and wash 1-2 times with PBS buffer (G4202 recommended) to reduce the interference of serum, phenol red or drugs on the experimental results;
    c. Probe labeling: After removing the cell culture medium or PBS buffer, add 100 μL Calcein AM detection working solution, and incubate at 37°C in the dark for 30 min;
    d. (Optional) Cell incubation: Change the cell culture medium and continue to incubate for 15-30 min to ensure that the Calcein AM in the cells is fully hydrolyzed;
    e. Fluorescence detection: Use a microplate reader for fluorescence reading, the maximum excitation wavelength of Calcein AM probe is 501 nm, and the maximum emission wavelength is 521 nm. Cell activity is directly proportional to fluorescence intensity. According to different experimental purposes, cell nuclei can be further counterstained or detected by other instruments (such as fluorescence microscope).
     
  3. Suspension cell detection:
    a. Cell seeding: uniformly seed cells in a 24-well plate at a certain density, and apply drugs or other pre-treatments to them (the seeding density is determined by factors such as cell size, growth speed, etc.);
    b. (Optional) Cell washing: Centrifuge at 1000 g for 3-5 min to remove the original medium, and wash 1-2 times with PBS buffer, each time you need 1000 g centrifugation for 3-5 min to reduce serum, phenol red or drugs Interference with experimental results;
    c. Probe labeling: After centrifugation at 1000 g for 3-5 min, remove the cell culture medium or PBS buffer, add an appropriate amount of Calcein AM detection working solution, and incubate for 30 min at 37°C in the dark;
    d. (Optional) Cell incubation: Change the cell culture medium and continue to incubate for 15-30 minutes to ensure that the Calcein AM in the cells is fully hydrolyzed;
    e. Fluorescence detection: use a microplate reader or flow cytometer for fluorescence detection. The maximum excitation wavelength of Calcein AM probe is 501 nm, and the maximum emission wavelength is 521 nm. Cell activity is directly proportional to fluorescence intensity. According to different experimental purposes, cell nuclei can be further counterstained or detected with other instruments (such as fluorescence microscope).

 

Notes:

  1. Calcein AM solution is centrifuged for a short time before use to reduce reagent loss.
  2. Avoid repeated freezing and thawing of Calcein AM solution. After receiving the product, it is recommended to appropriately aliquot it and store it in a sealed container at -20°C in the dark.
  3. Calcein AM is unstable in aqueous solution. Calcein AM detection working solution needs to be prepared for immediate use, and it will be effective within 24 hours.
  4. Due to different cell states, the incubation time and concentration of Calcein AM detection working solution can be adjusted according to the situation.
  5. When using a microplate reader to measure the fluorescence intensity, please use a black cell culture plate.
  6. Fluorescent dyes have quenching problems. Please operate and store them away from light.
  7. For your health and safety, please wear lab coats and gloves during operation.

 

This product is for scientific research purposes only, not for clinical diagnosis!

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