Use of CAL Fluor® Dyes

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Applicable Products

2423 3’-CAL Fluor® Orange 560 CPG 500
2424 3’-CAL Fluor® Red 610 CPG 500
2538 CAL Fluor® Orange 560-CE Phosphoramidite
2539 CAL Fluor® Red 590-CE Phosphoramidite
2540 CAL Fluor® Red 610-CE Phosphoramidite

Physical & Dilution Data

Dilution volumes (in ml) are for 0.1M solutions in dry, alcohol-free DCM or acetonitrile (see specifics below). Adjust accordingly for other concentrations. For µmol pack sizes, products should be diluted as 100µmol/ml to achieve 0.1M, regardless of molecular weight.

Item

Mol. Formula

Mol. Wt.

Unit Wt.

250mg

500mg

1g

2423 - - 1095.41 - - -
2424 - - 1171.44 - - -
2538 C40H53F6N5O4P2 843.83 560.60 2.96     5.93 11.85
2539 C42H57F6N5O4P2 871.88 588.65 2.87 5.73 11.47
2540 C46H57F6N5O4P2 919.93 636.69 2.72 5.44 10.87

Dissolution

2538 - Dilute to 0.1M in dry, alcohol-free DCM.

2539 & 2540 - Dilute to 0.1M in dry acetonitrile.

In all cases the use of 4Å molecular sieves is highly recommended.

Coupling

A 15min coupling is recommended. Due to the mild deprotection conditions we recommend using fast deprotecting amidites. Prior to dilution of amidites ensure that all product is at the bottom of the vial. Dilute to the recommended concentration and mix thoroughly in sealed vial to ensure that all contents are dissolved. Once dissolved, add the sieves. We recommend using ETT (3140/2/5) as the activator for these products.

Cleavage & Deprotection

Cleave and deprotect the CAL dye labelled oligo in 2-methoxyethylamine/MeOH (1:3) for 3h at 60ºC (all products) or 6h at 25ºC (CPGs). Filter the sample and evaporate the supernatant. Re-dissolve the CAL dye labelled oligo in water. The use of strong bases, such as concentrated ammonia, causes degradation and must be avoided.

Storage & Stability

The products are light sensitive. Prolonged exposure to light may cause photo-bleaching. The products are also susceptible to oxidation when left out in the open or undesiccated. Store at -20ºC, desiccated. Amidite solutions should be stored under argon, but used within 24h.

Spectral Data

 

Absorbance Max./nm

Emission Max./nm

Fluorescent Region

Dye Replaces

CAL Fluor® Orange 560 537 558 yellow-orange VIC, TET, JOE
CAL Fluor® Red 590 569 591 yellow-orange TAMRA
CAL Fluor® Red 610 590 610 orange-red Texas Red®

Black Hole Quencher, BHQ, CAL Fluor and Quasar, are registered trademarks of Biosearch Technologies, Inc. (BTI). All BTI products offered for sale by LINK are sold under the condition that they be used for research purposes only and are prohibited from use in diagnostic or other applications unless explicitly authorized by separate written agreement with BTI. Black Hole Quencher® (BHQ®) CAL Fluor®, and Quasar® dyes (referred to collectively as "BTI Dyes") are sold to the purchaser for internal R&D use only and are not to be used for clinical or clinical diagnostic purposes. Neither the BTI Dyes nor the compounds synthesized with them are to be re-packaged or re-sold. Separate licenses for other than the aforementioned internal R&D applications of the BTI Dyes may be available. Please inquire via info@biosearchtech.com. The Black Hole Quencher® dye technology is protected in the United States and other countries by U.S. patents and continuations numbered 7,019,129, 7,019,129B1, 7,109,312B2, 7,582,432, 8,410,255B2 and 8,440,399B2 issued to Biosearch Technologies, Inc. The CAL Fluor® technology is covered by U.S. patent number 7,344,701B2. The Quasar® technology is covered by U.S. Patent numbers 7,705,150B2, 7,868, 157B2 and 8,436,153B2.

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