Cell Function
Cell Function FAQs
In order to provide better customer support and answer customers' questions about cell function assay products, we have compiled and published answers to the frequently-asked questions (FAQs) about cell function assay products, and will continue to update them.
-
Plant tissues have cell walls and chloroplasts, which may interfere with JC-1 detection. We have not validated this application and cannot confirm its effectiveness. If testing, we recommend consulting relevant literature and conducting preliminary experiments.
-
One assay provides 1 mL BMDC differentiation medium (for 24-well plate, 1 mL/well) and 0.5 mL BMDC maturation medium. 200 assays can culture ~1×10^7 mouse bone marrow cells. After 7 days of differentiation, expect 1–3×10^7 immature DCs (70–80% purity). Following 24-hour maturation, yield 1–3×10^7 mature DCs (70–80% purity).
-
For optimal sensitivity: Use ≥1×10^6 cells or ≥50 mg tissue per experiment to achieve a protein concentration of 1–4 mg/mL. Lower amounts may reduce accuracy due to insufficient active caspase 3/7 (OD values below the linear detection range of 0.05–0.6).
-
The copper ions in the EdU kit may quench GFP/RFP/mCherry fluorescence. If you do NOT require GFP detection, the kit will not impact EdU assay results.
-
EDTA-containing trypsin can be used, but ensure thorough washing with PBS after digestion to remove residual EDTA.
-
Yes, but the fixation process may have certain effects on the intensity of the CFSE fluorescence signal. After fixation, it is best to perform detection as soon as possible, as over time, the fixative may lead to attenuation of the fluorescence signal or changes in cell morphology.
-
Regarding real - time video capture: The TUNEL assay itself is a detection method for a specific apoptotic marker (DNA fragmentation). It doesn't directly enable real - time visualization of the apoptotic process. The apoptotic process involves a series of complex biochemical and morphological changes that occur over time. The TUNEL kit labels cells that have already reached a certain stage of apoptosis (with DNA breaks). It doesn't continuously monitor the process from a healthy cell to an apoptotic cell. While the Leica DMi8 S Live Cell Microscope is a powerful tool for live - cell imaging, the TUNEL - labeled cells are apoptotic cells at a given moment. You would not be able to use the TUNEL - labeled cells to observe the entire apoptotic process in real - time using this method. To observe the apoptotic process in real - time, other methods such as using fluorescent probes that can monitor changes in mitochondrial membrane potential (which is an early event in apoptosis), caspase activation probes, or other dynamic markers related to the apoptotic pathway need to be employed. In conclusion, simply using the TUNEL - labeled cells obtained with the One - step TUNEL In Situ Apoptosis Kit and the Leica DMi8 S Live Cell Microscope is not sufficient to observe the apoptotic process in real - time.
-
Triton X-100 can disrupt cell membranes, allowing antibodies and TUNEL reagents to penetrate cells more easily and label target molecules. Theoretically, it can be used as a permeabilization agent. However, since you are performing TUNEL and IF co-staining, the concentration and incubation time need to be optimized experimentally. Typically, concentrations ranging from 0.1% to 0.5% are used.
-
https://www.elabscience.cn/resource-cell_function-detail-293
-
Hoechst can stain fixed cell nuclei, so it is compatible with TUNEL staining.
-
FITC-11-dUTP: The "11" in the name indicates that the fluorophore is connected to the dUTP base through a flexible carbon chain (or polyethylene glycol chain) of 11 atoms. Fluorescein-12-dUTP: The "12" indicates a longer linker chain (12 atoms), which may increase the distance between the fluorophore and the enzyme's active center, reducing steric hindrance.
-
Regarding your inquiry about performing in situ TUNEL assay (using our One-step TUNEL In Situ Apoptosis Kit, E-CK-A322) and IHC co-staining: The E-CK-A322 kit is based on a fluorescent method , while IHC is typically a chromogenic method.There are significant differences in the detection methods and intermediate operational procedures between these two assays. Therefore, we do not recommend co-staining TUNEL (E-CK-A322) with IHC. However, co-staining with IF is theoretically feasible, as both are fluorescent methods. Please note that we have not conducted specific validation tests for this combination using our kit. Implementing this would requirecustomers to explore and optimize the experimental conditions on their own.
