Fluorogen Activating Proteins

In the past decade, some interesting fluorescent protein dye complexes have been published. These are antibodies which bind a non-fluorescent dye (a fluorogen) and stabilize it in a conformation that makes it fluorescent – for example [1][2]. I’ve been curious about these for a while but the lack of commercial availability made them inconvenient to work with. It turns out that the scFvs from [2] – they called them fluorogen activating proteins or FAPs  – are now commercially available from SpectraGenetics.  There are two versions, a green (FITC-like) FAP (which is presumably the thiazole orange binding scFv) and a far red (Cy5-like) FAP (which is presumable the malachite green binding FAP). The far red FAP comes with two different fluorogens – one which is cell permeable and one which is cell impermeant (the only green fluorogen is impermeant). This means you can distinguish between protein on the cell surface and protein that is inside the cell [3][4].

There are probably other clever things you can do with these; if you try them out, let me know.


  1. A. Simeonov, M. Matsushita, E.A. Juban, E.H. Thompson, T.Z. Hoffman, A.E. Beuscher, M.J. Taylor, P. Wirsching, W. Rettig, J.K. McCusker, R.C. Stevens, D.P. Millar, P.G. Schultz, R.A. Lerner, and K.D. Janda, "Blue-fluorescent antibodies.", Science (New York, N.Y.), 2000. http://www.ncbi.nlm.nih.gov/pubmed/11030644
  2. C. Szent-Gyorgyi, B.F. Schmidt, B.A. Schmidt, Y. Creeger, G.W. Fisher, K.L. Zakel, S. Adler, J.A.J. Fitzpatrick, C.A. Woolford, Q. Yan, K.V. Vasilev, P.B. Berget, M.P. Bruchez, J.W. Jarvik, and A. Waggoner, "Fluorogen-activating single-chain antibodies for imaging cell surface proteins.", Nature biotechnology, 2007. http://www.ncbi.nlm.nih.gov/pubmed/18157118
  3. G.W. Fisher, S.A. Adler, M.H. Fuhrman, A.S. Waggoner, M.P. Bruchez, and J.W. Jarvik, "Detection and quantification of beta2AR internalization in living cells using FAP-based biosensor technology.", Journal of biomolecular screening, 2010. http://www.ncbi.nlm.nih.gov/pubmed/20488980
  4. J.P. Holleran, M.L. Glover, K.W. Peters, C.A. Bertrand, S.C. Watkins, J.W. Jarvik, and R.A. Frizzell, "Pharmacological rescue of the mutant cystic fibrosis transmembrane conductance regulator (CFTR) detected by use of a novel fluorescence platform.", Molecular medicine (Cambridge, Mass.), 2012. http://www.ncbi.nlm.nih.gov/pubmed/22396015