Cy5 disulfo NHS

Cy5 disulfo NHS modification is a post synthesis conjugation to a primary amino group thus an additional modification with an amino group is required. A 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.

The diSulfo– Cy5 NHS Ester is a hydrophilic version of Cy5 due to the two sulfo groups. This version is particularly helpful when the standard hydrophobic Cy5 version is not appropriate for the desired application. All NHS ester derivative modifications are post synthesis for oligos and requires a primary amino group on the oligo for conjugation. The amino group can be placed at either the 5’ or 3’ ends an internally as well.
Cy5 can be used as a replacement for Alexa Fluor 647 Succinimidyl Ester, DyLight 650 NHS Ester, Colorada 645 XT A - NHS ester, Fluorescentred 647 reactive, CF647 uccinimidyl ester and PromoFluor ® -647, NHS ester for the required applications.

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. 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.
2. 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.
3. Tyagi, S., Kramer, F.R. Molecular beacons: probes that fluoresce upon hybridization. Nat. Biotechnol. (1996), 14: 303-308.
4. Stoecker, K., Dorninger, C., Daims, H., Wagner, M. Double Labeling of Oligonucleotide Probes for Fluorescence In Situ Hybridization (DOPE-FISH) Improves Signal Intensity and Increases rRNA Accessibility. Appl. Environ. Microb.. (2010), 76: 922-926.

Reaction scheme for primary amine labelled oligos with NHS ester is shown in the figure below.

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- Cy5 disulfo NHS