What is the difference between Alexa Fluor 488 and fluorescein?
Share
Differences Between Alexa Fluor 488 and Fluorescein
Alexa Fluor 488 and Fluorescein are both fluorescent dyes commonly used in various applications such as fluorescence microscopy, flow cytometry, and enzyme-linked immunosorbent assays (ELISA). While they share some similarities, they also possess distinct characteristics that make them suitable for different applications.
Chemical Structure
Alexa Fluor 488 is a synthetic fluorophore designed to be more stable and less pH-sensitive than fluorescein. It has a complex structure that includes a sulfonated rhodamine derivative core. On the other hand, fluorescein, also known as FITC (Fluorescein isothiocyanate), has a simpler structure based on the xanthene dye backbone, making it more susceptible to photobleaching and pH changes.
Absorption and Emission Properties
| Dye | Absorption Maximum (nm) | Emission Maximum (nm) |
|---|---|---|
| Alexa Fluor 488 | ~495 | ~519 |
| Fluorescein/FITC | ~494 | ~518 |
Both dyes have similar absorption and emission maxima, which allows them to be excited by 488 nm laser lines commonly used in flow cytometry and confocal microscopy. Nevertheless, Alexa Fluor 488 typically exhibits a higher quantum yield and greater photostability than fluorescein.
", "stability": "Photobleaching and Stability
Alexa Fluor 488 is engineered for enhanced photostability, which makes it resistant to photobleaching, allowing for longer observation times in microscopy. In contrast, fluorescein is more prone to fading under prolonged exposure to light.
", "applications": "Applications
- Fluorescein: Due to its historical use and lower cost, fluorescein is widely utilized despite its limitations in stability. It remains a popular choice for routine applications where high photostability is not required.
- Alexa Fluor 488: Because of its superior photostability and brightness, Alexa Fluor 488 is preferred in applications that involve extensive imaging, or where highly accurate quantitation of fluorescence is necessary.
Conclusion
While Alexa Fluor 488 and fluorescein share many features, including similar excitation and emission spectra, Alexa Fluor 488 is generally superior in terms of photostability, brightness, and resistance to changes in pH. These characteristics often justify its higher cost in applications where they are critical factors.
" } ```