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Fluorescent Chromeo™ Py-Dyes

simple-to-use, fluorescent chameleon dyes for protein labeling

Chromeo™ Py-Dyes are a unique class of amine-reactive pyrylium dyes that change their color and become fluorescent upon reacting with primary amines. The shift in absorption and the induction of fluorescence effectively eliminate background effects from any unbound dye. The labeled proteins are ready to use immediately, and as the labeling method is fast, easy & reproducible, Chromeo Py-Dyes are suitable for a variety of “no-wash” applications:

  • Pre- and post-stain in SDS-protein electrophoresis
  • Pre-stain in capillary electrophoresis (CE)
  • Pre-stain in capillary isoelectric focusing (IEF)
  • Pre-stain in 2D capillary electrophoresis
  • Label for carbohydrate characterization
  • Small size label of proteins
  • Reagent to monitor amino functionalities on various surfaces

Chromeo Py-Dyes have been shown to work efficiently and with high sensitivity in different methods of protein electrophoresis. Proteins labeled with pyrylium dyes maintain their native charge and isoelectric points, which results in sharp and characteristic bands during electrophoresis. For your convenience, Chromeo Py-Dyes are also included in our complete Capillary Electrophoresis (CE) Dye Kits, containing optimized and quality-controlled buffer systems.

Name Format Cat No. Price  
Chromeo™ P503 1 mg 15106 $245 Buy Now
5 x 1 mg 16106 $825 Buy Now
Chromeo™ P540 1 mg 15107 $245 Buy Now
5 x 1 mg 16107 $825 Buy Now
Chromeo™ P429 1 mg 15108 $245 Buy Now
5 x 1 mg 16108 $825 Buy Now
Chromeo™ P543 1 mg 15109 $245 Buy Now
5 x 1 mg 16109 $825 Buy Now

Pyrylium dyes change their color and fluorescent properties upon reacting with proteins. After binding to primary amines, they undergo a large shortwave spectral shift that is visible to the naked eye (Figure 1). There is also a large increase in quantum yield that changes the non- or only weakly fluorescent dye into a bright fluorescent conjugate. (See the Properties tab for detailed information on each Py-Dye).

Color shift of Chromeo Py-Dyes caused by dye conjugation.
Figure 1: Color shift of Chromeo Py-Dyes caused by dye conjugation.

Chromeo P503 is blue while in its free state. After the short conjugation reaction, the dye's color undergoes a large Stokes shift and becomes red.

The shift of the absorption & emission bands together with the increased fluorescence quantum yield effectively eliminates background from any unbound dye. In addition, because unbound pyrylium dye is hydrolyzed during the labeling procedure, there is no need for purification following the simple one-step, room-temperature incubation. The labeled protein is ready to use immediately, and any possible background from unbound dye is eliminated. Due to the unique conjugation mechanism, proteins labeled with Py-Dyes maintain their native charge and isoelectric point. This is an important improvement over other derivatization-based methods for labeling proteins because it eliminates band or peak broadening in different assays and maintains the native characteristics of the proteins.

Chromeo™ Py-Dye advantages

  • No washing steps required
  • No background from unbound dye
  • Fast and simple procedure
  • Small size
  • No change in net charge of the labeled molecule
  • No interference of excitation and emission wavelength

Because of their unique features, Chromeo Py-Dyes (Chromeo P429, P503, P540 and P543) have been used successfully in a number of “no-wash” applications such capillary electrophoresis, SDS-protein gel electrophoresis, isoelectric focusing or as a label in receptor binding studies. See the Product Citations tab for a sample of published references.

Dye Dye State Absorption Emission ε L/(mol-cm) Quantum Yield*
Chromeo™ P503 Free 612 nm 665 nm 60,000 < 1%
Conjugated 503 nm 600 nm 24,000 ~50%
Chromeo™ P540 Free 587 nm 80,000 0%
Conjugated 533 nm 627 nm 50,000 ~20%
Chromeo™ P429 Free 457 nm 65,000 0%
Conjugated 429 nm 536 nm 75,000 ~10%
Chromeo™ P543 Free 570 nm 110,000 0%
Conjugated 543 nm 590 nm 57,000 ~15%
Table 1: Properties of Chromeo Py-Dyes.
*Quantum Yield will depend on the DPR of the conjugated protein.

Published references of Py-Dye applications


Pre-stain in SDS-gel electrophoresis

Amine-reactive pyrylium dyes have been used as a pre-stain in protein electrophoresis. The high sensitivity with a detection-limit of 16 picograms and the fast, reproducible procedure improve electrophoresis results compared to other commercially available stains like SYPRO Red or silver staining.

Protein stain in capillary electrophoresis (CE)

Pyrylium dyes are useful as a stain in capillary electrophoresis. The detection limit in the picomolar range, the fast reaction time and the preservation of the protein’s native charge make Chromeo Py-Dyes an interesting alternative to commonly used detection techniques.

Stain in microchip applications

On- and off-chip labeling on miniaturized platforms demonstrates the use of pyrylium dyes in the development of detection methods that require small amount of sample and feature fast analysis times.

Protein stain in capillary isoelectric focussing (CIEF)

Pyrylium dyes are excellent stains in capillary isoelectric focussing as they preserve the isoelectric point of the labeled proteins.

Label for carbohydrate characterization

Chromeo P503 (Py-1) was used to label a mixture of ethylenediamine functionalized oligosaccharides. The fluorescent carbohydrates have been analyzed in capillary electrophoresis (CE) and mass spectrometry (MALDI-ToF-MS).

Small protein label for functional analysis

Due to its small size, Chromeo P503 (Py-1) was used to fluorescently label a set of NPY receptor antagonists. The functionality of the labeled structures was shown by flow cytometry and confocal microscopy.

Monitoring of amino functionalities

Chromeo P503 (Py-1) was used to screen and evaluate the surface of polymer films by fluorescence spectroscopy and confocal scanning microscopy. The spectral shift of the bound dye enabled the distinction between covalently bound and nonspecifically adsorbed dye.

General application

Contents & Storage

Each Chromeo Py-Dye is supplied as a dry powder. Guaranteed stable for 6 months when stored properly.