AZDye-350

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.

Yield given below are for oligos shorter than 50mer. Please see longer oligos yield at this link Long Oligo Typical Yield.

~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
~32 nmol final yield for 2 umol scale synthesis
~160 nmol final yield for 10 umol scale synthesis
~240 nmol final yield for 15 umol scale synthesis

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.

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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-350