1. What is propidium iodide
Propidium iodide (PI) is a fluorescent dye that is commonly used in cellular research. It binds to nucleic acids but cannot cross intact cell membranes, making it suitable for separating dead or dying cells from living ones. This is because dead or dying cells' membranes are damaged, allowing the dye to enter the cell and bind to DNA.
2. Characteristics
PI's reluctance to penetrate live cells is owing to its molecular characteristics, which include its relatively large size and positive charge. The lipid bilayer of the cell membrane is hydrophobic and often impenetrable to charged molecules, therefore PI cannot diffuse across it.
3. How it works
PI intercalates between the base pairs of the DNA double helix, forming a stable complex with DNA. It also binds to RNA in a similar manner. This process is facilitated by the planar structure of PI, enabling it to fit between the base pairs with no preference for specific sequences. The intercalation disrupts the local structure of the nucleic acids, and once bound, PI fluoresces when exposed to light of a specific wavelength around 535 nm, emitting red fluorescence at approximately 617 nm.
3. Properties
Molecular weight | 668 g/mol |
Excitation | 535 nm (max) |
Emission / color | 617 nm (max) / emitting red fluorescent light |
Permeability | X |
4. Use
PI is a useful tool for differentiating between live and dead cells because it exclusively stains dying and dead cells. Because of its unique features, PI is frequently used in the following applications:
Viability:
In cell counting, PI selectively stains dead cells, enabling viability calculations when used alongside a permeable dye like acridine orange (AO) to count total cells.
Apoptosis:
PI is often paired with other dyes, such as Annexin V, to analyze apoptosis and necrosis.
Cell Cycle:
PI is also used to study the different phases of the cell cycle.
References:
NanoEntek products using propidium iodide reagent
Cell types | Cell lines, primary cell |
Measuring range | 5x10E4 ~ 4x10E6 cells/mL(PI) 5x10E4 ~ 2x10E7 cells/mL (AO/PI) |
Loading volume | 13 μL |
Staining reagents | Propidium iodide (PI) / Acridine orange (AO) |
Cell types | Cell lines, primary cell |
Measuring range | 5x10E4 ~ 4x10E6 cells/mL(PI) 5x10E4 ~ 2x10E7 cells/mL (AO/PI) |
Loading volume | 13 μL |
Staining reagents | Propidium iodide (PI) / Acridine orange (AO) |
21 CFR Part 11 | Available |
Measuring range | 5x10E4 ~ 5x10E6 cells/mL |
Applications | Total cell/viability Cell cycle Apoptosis Fluorescence expression |
Analysis time | Approx. 2 min ~ 4 min 30 sec (Depends on frames and assay) |
Loading volume | 25 μL |
Channels | Brightfield, GFP, RFP, DAPI |
21 CFR Part 11 | Available |
Comments