A VIC & HEX dye alternative—ultra-bright and license-free

License-free fluorescent dyes

Yakima Yellow dye is one in a family of fluorophores—known collectively as the Freedom Dyes—offered by IDT. Since our Freedom Dye program offers license- and royalty-free dyes, you have the assurance that your sequences can be screened in research mode, then easily transitioned to diagnostic or commercial application use, without the requirement for additional licensing.

Alternative to VIC^® and HEX dyes

Originally developed as an excellent alternative to VIC and HEX dyes, Yakima Yellow dye yields ultra-bright results when compared to HEX or ATTO™ 532 (Figure 1), and provides a better spectral match to VIC dye (Figure 2). The excitation and emission wavelength of the Yakima Yellow dye is 525 nm and 549 nm, respectively, and so it emits in the yellow-orange part of the visible spectrum.

Figure 1. Yakima Yellow dye shows higher fluorescence intensity compared to HEX and ATTO 532 dyes. A 13 nt probe, labeled with 1 of 3 dyes—Yakima Yellow, ATTO 532, or HEX dye—was used in PCR amplification of a 490 nt gBlocks^® Gene Fragments template, and fluorescence intensity was compared. Reactions were run in triplicate.

Figure 2. Yakima Yellow dye provides a better spectral match to VIC dye than does HEX dye. (A) Absorption spectra and (B) fluorescence emission spectra for oligonucleotides labeled with HEX, VIC, or Yakima Yellow dye in TE pH 8.0 buffer. The maximum wavelengths of absorption are as follows: HEX = 538 nm, VIC = 529 nm, Yakima Yellow = 525 nm. The maximum wavelengths of emission are as follows: HEX = 554 nm, VIC = 549 nm, Yakima Yellow = 549 nm. These maximum wavelengths can be affected by certain conditions, such as buffer pH, presence of solvent, and whether or not the dyes are conjugated to oligos. The wavelengths reported here represent conditions typical in most biological assays.

Yakima Yellow dye is not pH sensitive; thus, it emits the same fluorescence at pH 7 as it does at pH 9. Figure 3 shows the molecular structure of the Yakima Yellow dye.

Figure 3. Structure of the Yakima Yellow dye.

Applications in genotyping and gene expression assays

A clear advantage of Yakima Yellow dye is that it can be used readily in combination with other dyes for multiple readouts within a single reaction. For example, Yakima Yellow dye is paired with FAM in the rhAmp™ SNP Genotyping System to allow discrimination of 2 different alleles. Similarly, Yakima Yellow dye may be used in multiplex gene expression studies, using this dye on one PrimeTime^® qPCR Probe with compatible dyes on other probes (for assistance, visit the PrimeTime Multiplex Dye Selection Tool).

Using Yakima Yellow dye on common qPCR platforms, such as those from Bio-Rad or Applied Biosystems, will not normally require any additional calibration. Incorporating this VIC alternative into your qPCR assay workflow can be instantly advantageous through its ultra-sensitive spectral profile and versatility as an unrestricted dye.