HiLyte Fluor - Eurogentec

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HiLyte Fluor 647 Dye, an Excellent Replacement for Cy5 Dye
Cy5 is the preferred dye for preparing near-infrared fluorescent bioconjugates. Compared to the spectra of Cy5
dyes, the spectra of HiLyte Fluor 647 conjugates are only slightly red-shifted. This slight change in absorption spectrum makes
HiLyte Fluor 647 dyes an optimal match of filters designed for Cy5 dyes. In side-by-side comparison of antibody conjugates of
HiLyte Fluor 647 dyes and Cy5 conjugates (supplied by other companies), the total fluorescence of HiLyte Fluor 647 labeled
secondary antibodies is is significantly higher than that of Cy5 conjugates. Extinction coefficient is 250,000 M -1 cm -1 , with a
quantum yield of 0.33. Unlike Cy5 dyes, HiLyte Fluor 647 dyes have very little change in absorption or fluorescence spectra
when conjugated to most proteins, oligonucleotides and nucleic acids, thus yielding greater total fluorescence at the same degree of
substitution.
Normalized Absorbance
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1.0
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A
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Wavelength (nm)
Product Size Cat#
HiLyte Fluor 647 acid 5 mg 81255
HiLyte Fluor 647 amine 1 mg 81257
HiLyte Fluor 647 C2 maleimide 1 mg 81259
HiLyte Fluor 647 hydrazide 1 mg 81258
HiLyte Fluor 647 succinimidyl ester (SE) 1 mg 81256
1.2
1.0
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Normalized Fluorescence
RFU
35000
25000
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5000
HiLyte Fluor 647
Alexa Fluor 647
DyLight 647
Cy5
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Rabbit IgG (ng/well)
DC DE
Figure 7. Absorption and emission spectra of HiLyte Fluor 647 (panel A). Performance of
dye-goat anti-rabbit IgG conjugates (panel B). HiLyte Fluor 647 and Alexa Fluor 647
photostability over time (panel C). 3T3 cells incubated with anti-tubulin antibody and goatanti-rabbit
antibodies, labeled either with HiLyte Fluor 647 (panel D), or Cy5 (panel E),
nuclei were stained with Hoechst 33342 (Cat# 83218).
Figure 8. α-Tubulins of bovine pulmonary artery endothelial cells were probed
with biotin-conjugated mouse anti-α-tubulin, visualized with HiLyte Fluor 647
conjugated streptavidin (Cat# 60667). Mitochondria were detected with mouse
anti-Oxphos V complex, visualized with HiLyte Fluor 488-conjugated goat
anti-mouse (Cat# 28175-H488), nuclei stained with DAPI (Cat# 83210).
0.4
0 50 100 150 200 250 300
Irradiation Time (min)
© 2009 AnaSpec, Inc. • 2149 O’Toole Ave. • San Jose, CA 95131 • 800-452-5530 • www.anaspec.com
B
A/A 0
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C
HiLyte Fluor TM 647
Alexa Fluor TM 647