Cell Function FAQs

Calcein AM has strong fluorescence only after being hydrolyzed by endogenous esterase in the cell. Free calcein AM in buffer is almost non-fluorescent, so it does not need to be washed before detecting.

It is not recommended to dilute the dye before repackage, since it may lead to hydrolysis of Calcein AM as Calcein AM is more stable in stock solution. Calcein AM stock solution can be divided according to needs, and can be stored at -80°C for a longer shelf life. It is better to prepare the diluted buffer right before use, and use it up within one day.

The minimum number of cells is 1 million, and the protein concentration is between 1 and 4 mg/mL.

sf9 cells are insect cells, it can be used if caspase is indeed involved in the apoptosis pathway of insect cells.

The Caspase 9 kit measures the enzyme activity of caspase 9, which is activated caspase 9, also known as dimers or a tetramers of cleaved caspase 9.

JC-1 needs to detect the changes of fluorescence signal in both Ex/Em = 514/529 and Ex/Em = 585/590 at the same time to determine the changes of mitochondrial membrane potential, and there is fluorescence interference with GFP, so this kit cannot be applied to cells with GFP.

JC-1 exists in monomer and polymer form, and their emission spectra are different. In normal cells with high mitochondrial membrane potentia, JC-1 accumulates in the matrix of mitochondria as polymer, which producing red fluorescence. Since the decreasing of mitochondrial membrane potential during early stage of apoptosis, JC-1 can no longer be accumulated in the mitochondrial matrix, that becomes monomer with green fluorescence. Therefore, both red and green fluorescence need to be observed.

It is not recommended to do so. Cells may suffer apoptosis or damage due to mechanical manipulation during preparation into cell suspension, in which case the results of JC-1 may be inaccurate. In addition, this kit can only detect live single-cell suspensions and cannot be used to slice samples.

MTT assay kits are widely used to detect cell proliferation and cytotoxicity, and MTS, XXT, WST-1 and WST-8 are all upgraded alternatives to MTT, which have obvious advantages compared with MTT. On the one hand, the reaction production of MTT assay is insoluable in water, a specific solvent is required in the assay, products of WST-8, XTT and MTS are water-soluble with easier operation. On the other hand, the solubility of the formazan produced of WST-8 is higher than that of XTT and MTS. What's more, WST-8 is more stable than MTT, XTT, and has a wider linear range and higher sensitivity.

The effect of drug color can be eliminated by changing the medium after the drug incubation and before adding CCK-8.

BrdU method has many steps and requires the use of BrdU antibody, which has many influencing factors and poor stability. Based on EdU incorporation and subsequent click reaction, EdU method does not need to use antibodies, it has convenient operation and high detection sensitivity. It is a new method based on BrdU method, which will gradually replace BrdU method. EdU (5-ethynyl-2-deoxyuridine) is a thymidine analogue that can be incorporated into the newly synthesized DNA instead of thymidine in the process of DNA synthesis. The acetylene group on EdU can covalently react with fluorescence-labeled small molecular azide probes (such as FITC Azide, ElabFluor ®488 Azide, ElabFluor ®594 Azide, ElabFluor ®647 Azide) to form a stable triazole ring catalyzed by univalent copper ions, this reaction is called click reaction (Click reaction).