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Vectors for Expressing Fluorogen Activating Proteins (FAP-tags®) - Spectra Genetics - GPCR Internalization and Trafficking | Spectra Genetics - GPCR Internalization and Trafficking

Vectors for Expressing Fluorogen Activating Proteins (FAP-tags®)

FAP-Tagging Kit for Live Cell Assays (manual)

Vector Sequences and Annotation:  pMFAPalpha2; pMFAPbeta1

SpectraGenetics offers flexible, easy to use vectors for tagging cells and proteins with a series of new fluorescent reporters called fluorogen-activating proteins (FAPs)1.

  • FAP-tags® exhibit fluorescence only in the presence of submicromolar concentrations of cognate small-molecule fluorogens.  The fluorescent FAP/fluorogen complex is called a fluoromodule.  Neither the FAP nor the fluorogen is fluorescent by itself.
  • FAP-tags® allow the user to turn the fluorescent signal on and off by adding or removing fluorogen, or to change the signal wavelength by substituting one fluorogen for another.

Spectragenetics has developed two different types of FAPs, α and β.  Experiments can be designed with both FAPs to measure receptors internalization simultaneously (multiplex) or an individual FAP can be used for more detailed trafficking experiments.  Each FAP has a unique set of available fluorogens.  The α FAP has cell impermeant, permeant, pH sensitive and extra bright available.  This allows measurements of protein at the surface, total protein, internalization 1 color), and trafficking to and from the cell membrane.  The  βFAP has 2 spectrally distinct dyes that allow for measurement of surface protein, internalization (2 color), re-sensitization and pulse/chase experiments. Fluoromodule Properties:

FAP Fluorogen Excitation Max Emission Max Kd Recommended fluorogen working concentration
FAPα2 αRED1-np1 631 nm 650 nm < 1 nM 100 nM
αRED1-p1* 640 nm 668 nm < 1 nM 100 nM
FAPβ1 βGREEN-np1 509 nm 530 nm 3.1 nM 100 nM
βRED-np1 509,650 nm 670 nm ~13nM 100nM

* Membrane permeant fluorogen.  All other fluorogens are membrane-impermeant. When used in conjunction with membrane-impermeable fluorogens, FAP-tags® have proved particularly useful in visualizing the plasma membranes of cultured cells2, and in live-cell assays that monitor the translocation of membrane proteins to or from the cell surface3,4. SpectraGenetics offers vectors for FAP-tagging the plasma membrane of mammalian cells and cloning kits for expressing FAP-tagged fusion proteins, as well as an expanding collection of transfection-ready FAP-tagged genes. FAP-Tagging kits for mammalian surface expression of FAPs.  The vectors in these kits express FAPs that localize to the plasma membrane of transfected cells.  The vectors can also be used to clone and express cell-surface or secreted proteins with extracellular N-terminal FAP tags.

FAP-tag® In vector

Product

Price

FAPα2 pMFAPα2-kit-001 $499
FAPβ1 pMFAPβ1-kit-001 $499

Each kit contains a supply of fluorogen sufficient for 20 experiments with a media volume of 1 ml each, or 100 experiments with a volume of 200 µL each.

Fluorogen Pack Sizes and Prices:

 α

 Fluorogen Catalog Amount (nmol)  Measurements Price
αRED-np  αRED-np-001  2  100  $199
αRED-np-010  20  1,000 $999
 αRED-np-100  200  10,000 $4999
αRED-p  αRED-p-001  2   100  $199
 αRED-p-010  20  1,000 $999
 αRED-p-100  200  10,000 $4999
 αRED-Bright  se-Dyedron5x-001  2  100 $399
se-Dyedron5x-010  20  1,000  $1999
se-Dyedron5x-100  200  10,000  $9999
  αRED-pH  se-RED-pH-001  2  100  $399
 se-RED-pH-010  20  1,000 $1999
  se-RED-pH-100  200  10,000 $9999

β

 Fluorogen Catalog Amount (nmol) Measurements Price
 βGREEN-np βGREEN-np-001  2  100  $199
 βGREEN-np-010  20 1,000 $999
βGREEN-np-100  200 10,000 $4999
βRED-np  βRED-np-001  2  100  $299
βRED-np-010  20  1,000 $1499
 βRED-np-100  200  10,000 $7499

α fluorogens are used with αFAPs whereas β fluorogens are used with β FAPs.

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  1. Szent-Gyorgyi et al., Nature Biotechnology 26:235-240, 2008.  pdf
  2. Holleran et al., Cytometry A 77:776-782, 2010.  pdf
  3. Fisher et al., Journal of Biomolecular Screening15:703-709, 2010.  pdf
  4. Holleran et al., Molecular Medicine 18, 2012.  pdf