What are the reasons and solutions for FITC staining failure?
Pubtime：2024-05-30
View volume：157
1. The cell clusters did not separate. Solution: It is recommended to set the centrifugal force at 500-1000g. If the centrifugal force is too low, only normal cells will be collected, and apoptotic cells and fragments will be lost, resulting in a low apoptosis rate; 2. No single staining control was conducted. Solution: The fluorescence value and background of each reagent kit are different, and the position of the cross door is also different. For example, compared to Li Ji, domestic American brands tend to lean to the left, while domestic brands tend to lean to the right. Therefore, it is recommended that customers do single dye door marking; 3. Too many cells. Solution: If there is no signal in the picture, it means that it was not detected, which means the signal exceeds the detection range. It is recommended to set the parameter to 10000 cells; 4. FITC cannot be stained. Solution: Generally, freezing will have an impact on FITC, and Li Ji's product recommendation is not to freeze it; 5. The concentration and duration of dye exposure, as well as incubation conditions, are also the main factors affecting staining. Solution: Different cells may not be exactly the same, and there are optimal conditions that need to be explored and confirmed. 6. Insufficient stimulation of cell apoptosis itself. Solution: Adjust the duration and concentration of drug action. 7. The state of the cell itself, such as cells that proliferate excessively and are insensitive to apoptotic stimuli. Solution: Confirm the cell state. 8. The digestion time of adherent cells is too long, the PS on the membrane surface is damaged, and the number of Annexin V binding sites decreases. Solution: Adjust the cell digestion time and avoid excessive digestion
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