Fluorescent Proteins (FP)

Spectrum of Fluorescent Proteins

  • BFP = Blue Fluorescent Protein
  • CFP = Cyan Fluorescent Protein
  • GFP = Green Fluorescent Protein
  • RFP = Red Fluorescent Protein
  • YFP = Yellow Fluorescent Protein

    • Green Fluorescent Protein

    GFP, a fluorescing protein found in the jellyfish Aequorea of the north Pacific, has proven to be an amazingly effective probe into the inner function of cells and organisms. The GFP chromophore, consisting purely of amino acids, renders it well suited to genetic engineering and protein expression studies, and is often used to monitor localization and dynamics of cellular components and functions. The gene coding for GFP can be introduced into mammalian cells and other DNA to report on gene function and regulation in various tissues and organs.

    Furthermore, a number of flavors have now been engineered in the lab, creating a spectrum of GFP color variants, yielding emission wavelengths from blue (BFP) through to red (RFP, also called DSRed). This has enabled simultaneous study of multiple "FP-tagged" proteins within the living cell or organism, and opens the door to detection of multiple expressed proteins and their interactions. The Fluorescent Resonance Energy Trasnsfer (FRET) technique is often used (see the separate section on FRET in the Bilogy Applications section for more information).

    Widefield, total internal reflection and confocal fluorescence microscopy are all common techniques for visualization of GFP in living or fixed cells or tissues. Light-tight macro-imaging chambers are often used for measurement of localized GFP emission from whole animal models.

    See GFP with EMCCD

    Electron Multiplying CCD (EMCCD) technology is ideal for ultrasensitive detection of GFP and it’s color variants, whether as a key component in confocal live cell imaging systems, or as an EMCCD camera and software solution. The extraordinary Signal to Noise (S/N) offered at rapid rates of imaging is ideally suited to visualization and analysis of GFP in intracellular, tissue or whole animal environments with minimal excitation power. Through minimization of excitation powers, whether laser or lamp based, photobleaching rates of the fluorescent proteins can be dramatically reduced.

    At the single molecule level for example, it is absolutely critical that bleaching of fluorophores is reduced. In living cell environments, these "milder" excitation conditions also reduce the phototoxic effects that would otherwise kill the living cells, enabling them to be studied over longer periods of time. Furthermore, in some instances it can be beneficial to study cells that exhibit reduced expression of GFP or its color variants, since GFP over-expression can result in disruption of the actual cellular mechanisms under study.

    Find out more...