| Product Name | FITC-Annexin V/PI Apoptosis Assay Reagent |
| Catalog No. | CCAT-HMM-0011 |
| Description | Annexin V (membrane-associated protein V) is a Ca²⁺-dependent phospholipid-binding protein with a molecular weight of 35–36 kDa that selectively binds to phosphatidylserine (PS). Phosphatidylserine (PS) is primarily distributed on the inner side of the cell membrane, adjacent to the cytoplasm. During the early stages of apoptosis, various cell types translocate phosphatidylserine to the cell surface, exposing it to the extracellular environment. At this point, FITC-labeled Annexin V (FITC-Annexin V) binds to the translocated phosphatidylserine (PS), enabling direct detection of PS translocation—a key feature of apoptosis—using flow cytometry or fluorescence microscopy. For necrotic or late apoptotic cells, since cellular integrity has been compromised, FITC-Annexin V can enter the cytoplasm and bind to PS located on the inner side of the phospholipid bilayer, thereby also causing necrotic cells to exhibit green fluorescence. |
| Application | This product detects apoptosis levels by labeling early apoptotic cells (green) and necrotic or late apoptotic cells (red). |
| Spectral Parameters | FITC-Annexin V: Ex/Em (nm) = 494/518; PI: Ex/Em (nm) = 535/617 (with DNA) |
| Applicable Instruments | Flow cytometer, fluorescence microscope |
| Materials to Bring | 1 × PBS, pH 7.2-7.6; sterile ddH₂O |
| Storage | Store at 4°C away from light. Do not freeze. |
| Features | Easy to use: Multiple types available, wide range of choices; Good stability: High fluorescence brightness and resistant to quenching; Good grouping effect: Clear grouping and strong specificity. |
| Notes | 1. Before use, briefly centrifuge the product to the bottom of the tube, then proceed with subsequent experiments. 2. To reduce the rate of apoptosis, the incubation process can be performed on ice, but the incubation time should be extended to at least 30 minutes. 3. Since apoptosis is a rapid process, it is recommended that samples be analyzed within 1 hour after staining. 4. For adherent cells, digestion is a critical step. When inducing apoptosis in adherent cells, if there are floating cells, collect both floating and adherent cells and combine them for staining. Handle adherent cells carefully to avoid causing damage. If the trypsin digestion time is too short, cells may require vigorous pipetting to detach, which can damage the cell membrane and lead to excessive PI uptake; if the digestion time is too long, the cell membrane may also be damaged, potentially affecting the binding of phosphatidylserine on the cell membrane with FITC-Annexin V. During digestion, spread the trypsin evenly across the bottom of the plate, gently shake to ensure thorough contact between trypsin and cells, then remove most of the trypsin. Use the remaining small amount of trypsin for further digestion until the intercellular spaces enlarge and the bottom of the bottle exhibits a mottled pattern, at which point digestion can be terminated. Avoid using EDTA in the digestion solution, as EDTA can interfere with the binding of Annexin V to PS. 5. After digesting adherent cells with trypsin, it is recommended to resuspend them in optimal culture conditions and medium for approximately 30 minutes before staining to avoid false positives. 6. To minimize cell loss during washing, use a larger tip over a smaller tip when aspirating. 7. The optimal dye concentration is determined by specific experimental requirements. 8. All fluorescent dyes are subject to photobleaching. Please store and use them in a light-protected environment to minimize photobleaching. 9. For your safety and health, please wear a lab coat and disposable gloves during handling. |
For research use only, not for clinical use.
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