The Alexa Fluor family of fluorescent dyes is produced by Molecular Probes, Inc., a wholly owned subsidiary of Life Technologies Corporation. Alexa Fluor dyes are frequently used as cell and tissue labels in fluorescence microscopy and cell biology.
The excitation and emission spectra of the Alexa Fluor series cover the visible spectrum and extend into the infrared. The individual members of the family are numbered according roughly to their excitation maxima (in nm).
Alexa Fluor dyes are synthesized through sulfonation of coumarin, rhodamine, xanthene (such as fluorescein), and cyanine dyes. Sulfonation makes Alexa Fluor dyes negatively charged and hydrophilic. Alexa Fluor dyes are generally more stable, brighter, and less pH-sensitive than common dyes (e.g. fluorescein, rhodamine) of comparable excitation and emission, and to some extent the newer cyanine series. However, they are also more expensive. They are patented by Invitrogen (which acquired the company that developed the Alexa Fluor dyes, Molecular Probes).
|Quantum Yield |
|Alexa Fluor 350||blue||346||442||410||19,000||-|
|† = approximate color of the emission spectrum
ε = extinction coefficient
Comparison with other dyes
While extinction coefficients are known (see the table above), quantum yields and life times are not. Comparisons with other dyes should be considered depending on the conditions (technique) used and performance (signal, background, stability) needed.
The Alexa Fluor series dyes are less pH-sensitive and more photostable than the original dyes (fluorescein, rhodamine, etc.) from which they were synthesized. Brightness comparisons are also generally favorable. Comparisons with other dyes are less consistent, and also even more delicate, depending on the conditions (technique) used. A third party has compared Alexa Fluor 647 dye with Cy5 (similar wavelength), conjugated to DNA. This study found that Cy5 is brighter, but less photostable than Alexa Fluor 647. Other providers claim better brightness of photostability (i.e. Alexa Fluor 488 compared to Dylight488) and Fluoprobes488. These findings have been both confirmed and contested in scientific literature marking the difficulty in quantitatively comparing dyes. Alexa Fluor dyes remain excellent dyes in many cases, but empirical testing is needed for optimal results in each application.
- "Alexa Fluor Dyes Spanning the Visible and Infrared Spectrum". 2007-06-06. Archived from the original on 11 August 2007. Retrieved 2007-08-13.
- "The Alexa Fluor Dye Series". Molecular Probes, Inc. 2006-04-06. Archived from the original on 15 August 2007. Retrieved 2007-08-13.
- Panchuk-Voloshina N, Haugland RP, Bishop-Stewart J, et al. (1 September 1999). "Alexa dyes, a series of new fluorescent dyes that yield exceptionally bright, photostable conjugates". J. Histochem. Cytochem. 47 (9): 1179–88. doi:10.1177/002215549904700910. PMID 10449539.
- Berlier JE, Rothe A, Buller G, et al. (1 December 2003). "Quantitative comparison of long-wavelength Alexa Fluor dyes to Cy dyes: fluorescence of the dyes and their bioconjugates". J. Histochem. Cytochem. 51 (12): 1699–712. doi:10.1177/002215540305101214. PMID 14623938.
- "Fluorescence quantum yields (QY) and lifetimes (τ) for Alexa Fluor dyes—Table 1.5". Retrieved 2011-04-26.
- Ballard JL, Peeva VK, deSilva CJ, Lynch JL, Swanson NR (July 2007). "Comparison of Alexa Fluor and CyDye for practical DNA microarray use". Retrieved 23 October 2010.
- comparison of Alexa Fluor 488, 555 and 647 to Dylight 488, 549 and 649
- AlexaFluor 488, 546 and 555 and 647 compared to Fluorescein, Cy3, Cy5 and other dyes in flow cytometry or Confocal Microscopy