AlamarBlue® Cell Viability Assay
实验概要
Assess cell viability.
实验原理
Cell health can be monitored by numerous methods. Plasma membrane integrity, DNA synthesis, DNA content, enzyme activity, presence of ATP, and cellular reducing conditions are known indicators of cell viability and cell death. alamarBlue® cell viability reagent functions as a cell health indicator by using the reducing power of living cells to quantitatively measure the proliferation of various human and animal cell lines, bacteria, plant, and fungi allowing you to establish relative cytotoxicity of agents within various chemical classes. When cells are alive they maintain a reducing environment within the cytosol of the cell. Resazurin, the active ingredient of alamarBlue® reagent, is a non-toxic, cell permeable compound that is blue in color and virtually non-fluorescent. Upon entering cells, resazurin is reduced to resorufin, a compound that is red in color and highly fluorescent. Viable cells continuously convert resazurin to resorufin, increasing the overall fluorescence and color of the media surrounding cells.
主要试剂
AlamarBlue®
3% SDS [ Details：pH 7.4, 3% SDS in phosphate buffered saline (PBS) ]
实验材料
cells [ Details：Mammalian or bacterial cells in appropriate medium ]
plates [ Details：Appropriate 96- or 384-well plates ]
实验步骤
Optional: Treat cells with the test compound 24–72 hours prior to performing the alamarBlue® cytotoxicity assay.
1. Add 1/10th volume of alamarBlue® reagent directly to cells in culture medium as described in Table 1
2. Incubate for 1 to 4 hours at 37°C in a cell culture incubator, protected from direct light.
3. Record results using fluorescence or absorbance as follows:
Fluorescence: Read fluorescence using a fluorescence excitation wavelength of 540–570 nm (peak excitation is 570 nm). Read fluorescence emission at 580–610 nm (peak emission is 585 nm).
Absorbance: Monitor the absorbance of alamarBlue® at 570 nm, using 600 nm as a reference wavelength (normalized to the 600 nm value).
4. Optional: Add 50 μL 3% SDS directly to 100 μL of cells in alamarBlue® reagent to stop the reaction.
注意事项
1. Sensitivity of detection increases with longer incubation times. For samples with fewer cells, use longer incubation times of up to 24 hours.
2. Fluorescence mode measurements are more sensitive. When fluorescence instrumentation is unavailable, monitor the absorbance of alamarBlue® reagent. Assay plates or tubes can be wrapped in foil, stored at 4°C, and read within 1–3 days without affecting the fluorescence or absorbance values.
附 件 (共1个附件,占15KB)
picture1.jpg
15KB 查看
