Cyan fluorescent protein TagCFP

- Bright cyan fluorescence
- Monomeric protein with successful performance in fusions
- High pH stability and photostability
- Recommended for protein labeling

TagCFP is a cyan monomeric protein generated on the basis of the wild-type GFP-like protein from jellyfish Aequorea macrodactyla [Xia et al., 2002]. It possesses bright fluorescence with excitation/emission maxima at 458 and 480 nm, respectively. TagCFP is significantly brighter than commonly used ECFP. TagCFP is mainly intended for protein labeling in protein localization and interaction studies. It can also be used for cell and organelle labeling and for tracking the promoter activity.

Main properties

TagCFP normalized excitation (thin line) and emission (thick line) spectra. Download TagCFP spectra (xls)

CHARACTERISTIC *Brightness is a product of extinction coefficient and quantum yield, divided by 1000. Molecular weight, kDa 26.7 Polypeptide length, aa 239 Fluorescence color cyan Excitation maximum, nm 458 Emission maximum, nm 480 Quantum yield 0.57 Extinction coefficient, M-1cm-1 37 000 Brightness* 21.1 Brightness, % of EGFP 64 pKa 4.7 Structure monomer Aggregation no Maturation rate at 37°C fast Photostability high Cell toxicity not observed Main advantages bright cyan monomeric fluorescent protein

Recommended filter sets and antibodies

TagCFP can be recognized using Anti-Tag(CGY)FP antibody (Cat.# AB121-AB122) available from Evrogen.

TagCFP can be detected using fluorescence filter sets for ECFP and the similar. Recommended Omega Optical filter sets are XF114-2 and XF130-2.

Performance and use

TagCFP can be easily expressed and detected in a wide range of organisms. Mammalian cells transiently transfected with TagCFP expression vectors give bright fluorescent signals within 10-12 hrs after transfection. No cell toxic effects and visible protein aggregation are observed.

TagCFP performance in fusions has been demonstrated in human cytoplasmic β-actin, α-tubulin, and mitochondria models.

		TagCFP use for protein labeling. Confocal microscopy of transiently transfected mammalian cells expressing TagCFP-tagged fusions: (A) α-tubulin; (B) β-actin. See also 3D vizualization of TagCFP-α-tubulin fusion

TagCFP suitability to generate stably transfected cells has been proven by Marinpharm company. Cell line expressing TagCFP fusion with mitochondrial targeting sequence (MTS) is commercially available.

		TagCFP use for labeling of cells and cellular orgenelles. (A) Confocal microscopy of transiently transfected mammalian cells expressing TagCFP; (B) Mitochondria-targeted TagCFP in stably transfected U-205 cell line. Image was kindly provided by Dr. Christian Petzelt (Marinpharm).

TagCFP can be used in multicolor labeling applications with green, yellow, red, and far-red fluorescent dyes.

TagCFP use in multicolor labeling of mammalian cells. (A) TagCFP-tagged β-actin (cyan), TagYFP-tagged α-tubulin (green), TagFP635-H2B fusion (red), and Golgi-targeted TagRFP (orange); (B) mitochondria-targeted TagCFP (cyan), peroxisome-targeted PhiYFP-m (yellow), and membrane-targeted KillerRed (red); (C) TagCFP-tagged α-tubulin (cyan), TagFP635-clathrin fusion (red), mitochondria-targeted TagYFP (green). Images (A, C) were kindly provided by Michael W. Davidson (Florida State University).

Available variants and fusions

TagCFP codon usage is optimized for high expression in mammalian cells [Haas et al., 1996], but it can be successfully expressed in many other heterological systems.

TagCFP-mito fusion: A mitochondrial targeting sequence (MTS) is linked to the TagCFP N-terminus. MTS was derived from the subunit VIII of human cytochrome C oxidase [Rizzuto et al., 1989; Rizzuto et al., 1995]. When expressed in mammalian cells, this variant provides cyan fluorescent labeling of mitochondria.

TagCFP-actin fusion: Human β-actin is linked to the TagCFP C-terminus. When expressed in mammalian cells, this fusion provides cyan fluorescent labeling of actin filaments.

TagCFP-tubulin fusion: Human α-tubulin is linked to the TagCFP C-terminus. When expressed in mammalian cells, this fusion provides cyan fluorescent labeling of tubulin filaments.