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Product Details
| Detection Method | Flow cytometry (Ex = 488 nm; Em = 530 nm). |
|---|---|
| Species Reactivity | Mammalian |
| Applications | Annexin V-FITC kit includes Annexin V-FITC for detecting apoptosis and also sytox green dye for detecting necrosis. |
| Features & Benefits | • Simple one-step procedure; takes only 10 minutes • Fast and convenient • The Annexin VFITC Plus kit includes sytox green dye (instead of PI) which stains necrotic cells with a higher level of green fluorescence than FITC does |
| Kit Components | • Annexin V-FITC • SYTOX Green Dye • Binding Buffer |
| Storage Conditions | 4°C |
| Shipping Conditions | Gel Pack |
| USAGE | For Research Use Only! Not For Use in Humans. |
Details
The Annexin V-FITC Apoptosis Detection Kit Plus is based on the observation that soon after initiating apoptosis, most cell types translocate the membrane phospholipid phosphatidylserine (PS) from the inner face of the plasma membrane to the cell surface. Once on the cell surface, PS can easily be detected by staining with a fluorescent conjugate of Annexin V, a protein that has a strong natural affinity for PS. The one-step staining procedure takes only 10 minutes. In addition, the assay can be directly performed on • Live cells, without the need of fixation. The Annexin V-FITC Apoptosis Detection Kit Plus includes annexin V-FITC, SYTOX green dye, and binding buffer. The SYTOX green dye is impermeant to • Live cells and apoptotic cells, but stains necrotic cells with intense green fluorescence by binding to cellular nucleic acids. After staining a cell population with annexin V-FITC and SYTOX Green dye in the provided binding buffer, apoptotic cells show green fluorescence, dead cells show a higher level of green fluorescence and • Live cells show little or no fluorescence. These populations can easily be distinguished using a flow cytometry with the 488 nm line of an argon-ion laser for excitation. Both annexin V-FITC and SYTOX Green dye emit green fluorescence that can be detected in the FL1 channel, freeing the other channels for the addition of other probes in multi-color labeling experiments.
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Can the Sytox Green interfere with the FITC signal?
Both FITC and Sytox green have similar excitation/emission profiles. SO the same excitation laser can be used for both. There would be 3 cell populations from this assay:
• live cells - with only a low level of fluorescence from FITC alone
• apoptotic cells - with moderate green fluorescence (mostly from FITC)
• and necrotic cells with high-intensity green fluorescence (sytox)
• live cells - with only a low level of fluorescence from FITC alone
• apoptotic cells - with moderate green fluorescence (mostly from FITC)
• and necrotic cells with high-intensity green fluorescence (sytox)
I am looking to investigate cell apoptosis using 96 well plates. I was wondering if your Annexin V-FITC Apoptosis Detection Kit could be adapted to a 96 well plate fluorescent plate reader format?
This assay can be adapted to a plate reader format. For this particular assay, take 10^5 trypsinized cells into the plate wells, resuspend in 100 µl binding buffer, add 1µ Annexin V and sytox green dye respectively, incubate them in the dyes, give a gentle wash and then analyze. The cell number can vary from cell to cell and/or treatment protocol selected. Please take measures to prevent overcrowding in the wells and also aspirate solutions gently since the apoptotic cells tend to detach quickly.
Can you prelabel the cells and then follow apoptosis using this kit?
Unfortunately, the product has not yet been tested in this manner. We are unsure whether it might work and needs significant optimization with known controls and timepoints to determine if the kit is performing correctly in these conditions.
Can we use frozen samples with this assay?
This kit can only work on freshly prepared samples.
Can the kit work on bacteria or yeast cells?
The kit has been standardized for mammalian cells only.
What is the exact volume of sample required for this assay?
There is no specific volume we can recommend for the amount any sample to be used since it is completely sample concentration and quality based. You have to do a pilot expt with multiple sample volumes to determine the optimal volume which gives a reading within the linear range of the standard curve. Please refer to the citations for this product to see what other clients have used with similar sample types.
Do you have trial sizes of this kit?
Unfortunately, we do not have trial sizes of this kit available. However, if you are based in the US or Canada, we will give you a 10% off list price introductory discount on its purchase price. If you are based out of this area please contact yopur regional BioVision distributor.
What is the shelf life of this kit?
The expiry date is typically 1 year from the date of shipment
Can we purchase individual components of this kit?
Yes, you can purchase any of the kit's components without the whole kit. Please refer to the component Cat #s mentioned on the datasheet for ordering.
Can we use an alternate buffer for sample preparation (cell lysis, sample dilutions etc)?
Our assay buffers are optimized for the reactions they are designed for. They not only contain some detergents for efficient lysis of your cells/tissue, but also contain some proprietary components required for the further reactions. Therefore, we highly recommend using the buffers provided in the kit for the best results.
Will trypsinizing the cells removed the phosphatidylserine?
The trypsin used in this kit should in no way affect the PS on the cell membrane.
Shanshan Qi, Hypocrellin A-based photodynamic action induces apoptosis in A549 cells through ROS-mediated mitochondrial signaling pathway. Acta Pharm Sin B., March 2019; 30972277.
Weng et al., Inhibition of miR-17 and miR-20a by Oridonin Triggers Apoptosis and Reverses Chemoresistance by Derepressing BIM-S. Cancer Res., Aug 2014; 74: 4409 - 4419.
Fu et al., YAP regulates cell proliferation, migration, and steroidogenesis in adult granulosa cell tumors. Endocr. Relat. Cancer, Mar 2014; 21: 297 - 310.
Shili Xu et al., Discovery of a Novel Orally Active Small-Molecule gp130 Inhibitor for the Treatment of Ovarian Cancer. Mol. Cancer Ther., Jun 2013; 12: 937 - 949.
Xiaoguang Li et al., Japonicone A Suppresses Growth of Burkitt Lymphoma Cells through Its Effect on NF-
B. Clin. Cancer Res., Jun 2013; 19: 2917 - 2928.
B. Clin. Cancer Res., Jun 2013; 19: 2917 - 2928.For more citations of this product click here
