Simultaneous Apoptotic, Necrotic & Healthy Cell Quantification
Apoptosis occurs via a complex signalling cascade that is tightly regulated at multiple points and there are many opportunities to evaluate and detect the activity of the proteins involved. Consequently, there are a large number of apoptosis assays available to quantify apoptotic cells. However, most assays lack the ability to distinguish apoptosis from necrosis since many features of these processes overlap. Most detection approaches employ two or more distinct assays to confirm that cell death is occurring via apoptosis.
Cambridge Bioscience offers the Apoptotic, Necrotic & Healthy Cells Quantification Kits Plus for the simultaneous, convenient detection and quantification of apoptotic, necrotic and healthy cells within the same cell population by flow cytometry or fluorescence microscopy. This kit, from Biotium, uses Annexin V labelled with CF™488, Ethidium Homodimer III and Hoechst 33342 for labelling apoptotic, necrotic and total cell populations green, red and blue respectively. Annexin CF™488 is brighter and more photostable than FITC-Annexin V and ethidium homodimer III is a superior alternative to both PI and EthD-I.
Benefits of Using the Apoptotic, Necrotic & Healthy Cell Quantification Kit Plus
The apoptotic, necrotic & healthy cell quantification kit plus offers numerous advantages over other detection kits and panels:
• Simultaneous detection of apoptotic, necrotic & healthy cells in a single assay
• Simple to use
• Fast - a single 15-30 minute staining step.
• Sensitive - EthD-III is a superior alternative to both PI and EthD-I for staining necrotic cells
• Uses the photostable & bright CF488™ dye
• Analysis by flow cytometry or fluorescence microscopy
1. Detecting Apoptotic Cells - Green
During apoptosis, phosphatidylserine (PS) is translocated from the inner to the outer surface of the cell for phagocytic cell recognition. Using Annexin V, which has a high affinity for PS, labelled with CF™488, this kit identifies apoptotic cells highlighting them with green fluorescence by binding to PS exposed on the outer leaflet. CF™488A is spectrally similar to fluorescein (FITC) but offers a much brighter and more photostable fluorescence.
2. Detecting Necrotic Cells - Red
During necrosis, both internal organelle and plasma membrane integrity are lost, resulting in spilling of cytosolic and cellular contents into the surrounding environment. Utilising Ethidium homodimer III (EtD-III), a highly positively charged nucleic acid probe which is impermeant to live cells or apoptotic cells, this kit stains and identifies necrotic cells with red fluorescence (λabs/λem = 528/617 nm). EthD-III is a superior alternative to propidium iodide (PI) or ethidium homodimer I used in other kits due to its significantly higher affinity for DNA and higher fluorescence quantum yield.
3. Detecting Healthy Cells - Blue
Utilising Hoechst 33342, a cell membrane-permeant, minor groove-binding DNA stain, this kit identifies healthy cells by emitting bright blue fluorescence upon binding to DNA (λabs/λem = 350/461 nm). Hoechst 33342 stains the nuclei of all cells blue. Healthy cells can be differentiated from apoptotic/necrotic cells as the former healthy cells will be stained by Hoechst only, and the latter will be stained both blue and green (CF™488-Annexin V) and blue and red (EtD-III) respectively.
Note: product availability depends on country. Product detail pages show availability of each product.