Modification : AZDye-647
Catalog Reference Number
Category
Modification Code
5 Prime
3 Prime
Internal
Molecular Weight (mw)
Extinction Coeficient (ec)
Technical Info (pdf)
Absorbance MAX
Emission MAX
Absorbance EC
26-6479
Fluorescent Dyes
[AZDye-647-N]
Y
Y
Y
970.1
191
PS26-6479.pdf
655
680
270
| Catalog No | Scale | Price |
| 26-6479-05 | 50 nmol | $621.00 |
| 26-6479-02 | 200 nmol | $621.00 |
| 26-6479-01 | 1 umol | $729.00 |
| 26-6479-03 | 2 umol | $1,166.00 |
| 26-6479-10 | 10 umol | $5,832.00 |
Click here for a list of fluorophores.
This modification is a post synthesis conjugation to a primary amino group thus an additional modification with an amino group is required. A C3, C6 or C12 amino group can be placed at the 5' or for the 3' end a C3 or C7 amino and for internal positions an amino modified base is used, e.g Amino dT C6.
YIELD
NHS based modifications are post synthesis conjugation performed using a primary amino group. The yield is lower as compared to direct automated coupling of modifications that are available as amidites. Approximate yield for various scales are given below.
~2 nmol final yield for 50 nmol scale synthesis.
~5 nmol final yield for 200 nmol scale synthesis.
~16 nmol final yield for 1 umol scale synthesis
AFDye 647 Abs Maxima is at 655 nm and Emission Maxima is at 680 nm.
Human vision is insensitive to light beyond ~650 nm; it is not possible to view near-IR fluorescent dyes.
AZDyes are a set of fluorescent dyes that span the visible electromagnetic spectrum, as well as some of the near-IR. The absorbance range is 346-749 nm, and the emission range is 442-775 nm. Generally speaking, AZDyes are brighter, chemically more stable, and less pH-sensitive than other fluorescent dyes commonly used to label oligonucleotides (1). Because they currently only are in the form of NHS esters, oligos first must be synthesized with an Amino Linker modification (either at the ends or internally). The appropriate AZDye is then manually attached to the oligo through the amino group in a separate reaction post-synthesis. The list of currently available dyes includes AZDye 350, -405, -430, -488, -500, -514, -532, -546, -555, -568, -594, -610, -633, -647, -660, -680, -700, -750, with the number indicating the appropriate absorbance wavelength for the particular dye. AZDyes are suitable for a variety of in vitro and in vivo applications. However, for in vivo experiments, users should note that dyes 350/405, being "blue" dyes, require higher-energy excitations than the other. Users of these particular dyes should confirm that the higher-energy required for excitation does not damage the relevant cells or tissues being used in the in vivo experiments.
-
|
Near Infrared Fluorophore Spectral Data & Quencher Selection Guide
|
|
Fluorophore Name
|
Excitation Max, nm +/-10
|
Emission Max, nm +/-10
|
Extinction Coefficient*
|
Color**
|
Quencher
|
Cy5 |
650 |
665 |
250,000 |
Human vision is insensitive to light beyond ~650 nm. |
BBQ-650 λ (max) = 650nm Range = 550-750 nm |
IRDye 650 NHS |
650 |
665 |
230,000 |
Human vision is insensitive to light beyond ~650 nm. |
BBQ-650 λ (max) = 650nm Range = 550-750 nm |
AZ647 NHS |
655 |
680 |
191,800 |
Human vision is insensitive to light beyond ~650 nm. |
BBQ-650 λ (max) = 650nm Range = 550-750 nm |
Cy5.5 |
684 |
710 |
198,000 |
Human vision is insensitive to light beyond ~650 nm. |
BBQ-650 λ (max) = 650nm Range = 550-750 nm |
IRDye 700 NHS |
684 |
710 |
288,000 |
Human vision is insensitive to light beyond ~650 nm. |
BBQ-650 λ (max) = 650nm Range = 550-750 nm |
Cy7 NHS |
740 |
773 |
199,000 |
Human vision is insensitive to light beyond ~650 nm. |
BBQ-650 λ (max) = 650nm Range = 550-750 nm |
IRDye 750 NHS |
756 |
776 |
260,000 |
Human vision is insensitive to light beyond ~650 nm. |
BBQ-650 λ (max) = 650nm Range = 550-750 nm |
cy7.5 NHS |
788 |
808 |
223,000 |
Human vision is insensitive to light beyond ~650 nm. |
BBQ-650 λ (max) = 650nm Range = 550-750 nm |
IRDye 800 NHS |
795 |
819 |
240,000 |
Human vision is insensitive to light beyond ~650 nm. |
BBQ-650 λ (max) = 650nm Range = 550-750 nm |
|
* Extinction coefficient at λ (max) in cm-1M-1. ** Typical emission color seen through the eyepiece of a conventional fluorescence microscope with appropriate filters. Near-IR region. Human vision is insensitive to light beyond ~650 nm; it is not possible to view near-IR fluorescent dyes.
Click here for a list of fluorophores.
|
|
Click here for list of quenchers.
|
References
1. Panchuk-Voloshina, N., Haugland, R.P., Bishop-Stewart, J., Bhalgat, M.K., Millard, P.J., Mao, F., Leung, W-Y., Haugland, R.P. Alexa Dyes, a Series of New Fluorescent Dyes that Yield Exceptionally Bright, Photostable Conjugates. J. Histochem. Cytochem. (1999), 47: 1179-1188.
Click here for list of quenchers.
Click here for a list of fluorophores.
|
Quencher Spectral Data
|
|
Quencher
|
Absorption Max, nm
|
Quenching Range, nm
|
| Dabcyl |
453 |
380-530 |
| BHQ-0 |
495 |
430-520 |
| BHQ1 |
534 |
480-580 |
| BHQ2 |
579 |
550-650 |
| BHQ3 |
672 |
620-730 |
| BBQ-650 |
650 |
550-750 |
|
Click here for complete list of quenchers and details
|
**Black Hole Quencher License Agreement
Black Hole Quencher License Agreement.
"Black Hole Quencher®, BHQ®, CAL Fluor® and Quasar® are registered trademarks of
Biosearch Technologies, Inc., Petaluma, California.
The BHQ, CAL Fluor and Quasar dye technologies are protected by U.S. and world-wide patents either issued or in application. Compounds
incorporating these dyes are made and sold under agreement with Biosearch Technologies, Inc. for end-user's non-commercial research and development use only.
Their use in commercial applications is encouraged but requires a separate Commercial Use License granted by Biosearch Technologies, Inc."
|
References
1. Didenko, V.V. DNA Probes Using Fluorescence Resonance Energy Transfer (FRET): Designs and Applications. Biotechniques (2001), 31: 1106-1121.
2. Livak, K.J., Flood, S.J.A., Marmaro, J., Giusti, W., Deetz, K. Oligonucleotides with fluorescent dyes at opposite ends provide a quenched probe system useful for detecting PCR product and nucleic acid hybridization.PCR Methods Appl. (1995), 4: 1-6.
3. Thelwell, N., Millington, S., Solinas, A., Booth, J., Brown, T. Mode of action and application of Scorpion primers to mutation detection. Nucleic Acids Res. (2000), 28: 3752-3761.
4. Tyagi, S., Kramer, F.R. Molecular beacons: probes that fluoresce upon hybridization. Nat. Biotechnol. (1996), 14: 303-308.
- AZDye-647
Oligo Purification