Actively helping customers, employees and the global community during the coronavirus SARS-CoV-2 outbreak.  Learn more >>

Phalloidin Conjugates

Phalloidin is a bicyclic heptapeptide isolated from the poisonous death cap mushroom, Amanita phalloides. It possesses a high binding affinity for the grooves between F-actin subunits over monomeric G-actin. Compared to actin antibodies, non-specific binding of phalloidin is negligible, thus providing minimal background and high-contrast during cellular imaging. Once bound to F-actin, phalloidin shifts the equilibrium of monomers and filaments toward the filaments position, and inhibits ATP-hydrolysis. The interaction stabilizes actin filaments by preventing subunit dissociation, and it promotes actin polymerization by lowering the critical concentration.

When conjugated to fluorescent dyes, Phalloidin can be used at nanomolar concentrations to label and visualize filamentous actin in fixed and permeabilized cells, cell cultures and cell-free experiments, as well as, formaldehyde-fixed and permeabilized tissue sections. Phalloidin conjugates exhibit similar affinity for all types and sizes of actin filaments, binding in a stoichiometric ratio of 1:1 (phallotoxin : actin) in both muscle and nonmuscle cells. In comparison to antibodies, phalloidin derivatives' are small (< 2 kDa) and phalloidin bound filaments do not impede functional properties of the filaments. The small size also permits for denser F-actin labeling producing more detailed stains when imaged at higher resolutions. In addition, because actin is evolutionarily conserved, the binding properties of phalloidin derivatives can be utilized in staining a wide range of animal and plant cells.


Available Phalloidin Conjugates for Staining F-actin

Phalloidin-iFluor™ 488
Figure 1. Actin filaments visualized with Phalloidin-iFluor™ conjugates. F-actin, nucleus and lysosomes were stained, respectively with Phalloidin iFluor™ 488, Nuclear Blue™ DCS1 and LysoBrite™ Red.

Phalloidin iFluor™ Conjugates


Exploiting the excellent fluorescence properties of our iFluor™ dyes, AAT Bioquest has developed a series of phalloidin conjugates spanning the full color spectrum which selectively bind to F-actins. Used at nanomolar concentration, these iFluor™ phalloidin derivatives are convenient for many actin-related assays. Such assays include labeling, identifying, and quantitating F-actins in formaldehyde-fixed and permeabilized tissue sections, cell-cultures, or cell-free experiments. Compared to traditional fluorescein isothiocynanate (FITC) and rhodamine conjugates, Phalloidin-iFluor™ conjugates offer F-actin stains that are superior in brightness and photostability. Phalloidin-iFluor™ conjugates are equivalent, and in most cases superior, to Alexa Fluor® conjugates in performance.

We encourage the research community to try our Phalloidin-iFluor™ conjugates with their respective protocols for a more intense fluorescence in their F-actin staining. Phalloidin-iFluor™ 700, iFluor™-750 and iFluor™-790 conjugates are among the few near-infrared stains available with sufficient spectral separation from commonly used red fluorophores.



Table 1. Overview of fluorescent phalloidin conjugates

Specification
Value
ApplicationFluorescent imaging of filamentous actin (F-actin)
Sample typeFixed & permeabilized cells, cell cultures, cell-free experiments, formaldehyde-fixed & permeabilized tissue sections
Cellular localizatoinF-actin of the cytoskeleton and contractile apparatus of muscle cells (does not bind to monomeric G-actin)
Live/Fixed CellsFixed cells only
PermeabilityCell impermeant
MultiplexingYes
PlatformFluorescence microscope and imaging platforms


Table 2. Phalloidin iFluor™ Conjugates

Conjugate
Mol. Wt.
Ex/Em (nm)
Filter Set (nm)
ε
Φ
Unit Size
Price
Phalloidin-iFluor™ 350∼1300344/448DAPI20,0000.95300 Tests$153
Phalloidin-iFluor™ 405∼1400402/425DAPI37,0000.91300 Tests$153
Phalloidin-iFluor™ 488∼1900491/516FITC75,0000.9300 Tests$153
Phalloidin-iFluor™ 514∼1800527/554TRITC80,0000.83300 Tests$153
Phalloidin-iFluor™ 532∼1800543/563TRITC90,0000.68300 Tests$153
Phalloidin-iFluor™ 555∼1300556/569TRITC100,0000.64300 Tests$153
Phalloidin-iFluor™ 594∼1600587/603Texas Red180,0000.53300 Tests$153
Phalloidin-iFluor™ 6331703.99638/652Texas Red250,0000.29300 Tests$153
Phalloidin-iFluor™ 6471632.87654/669Cy5250,0000.25300 Tests$153
Phalloidin-iFluor™ 680∼2000683/700Cy5.5220,0000.23300 Tests$310
Note:
  1. Extinction Coefficient = molar extinction coefficient at their maximum absorption wavelength (Units = cm-1M-1).
  2. Quantum Yield = fluorescence quantum yield in aqueous buffer (pH 7.2).



Phalloidin-AF Dye Conjugates

The following table outlines the fluorescence properties of available Phalloidin-AF dye conjugates for use in fluorescence imaging applications. For additional information on Phalloidin-AF dye conjugates



Table 3. Available Phalloidin-AF dye conjugates.

Conjugate
Mol. Wt.
Ex/ Em (nm)
Filter Set
Extinction Coefficient¹
Quantum Yield²
Unit Size
Cat No.
XFD350 Phalloidin *XFD350 Same Structure to Alexa Fluor™ 350*∼1100343/441DAPI19000Not Determined300 Tests23150
XFD488 Phalloidin *XFD488 Same Structure to Alexa Fluor™ 488*∼1300499/520FITC730000.92300 Tests23153
XFD594 Phalloidin *XFD594 Same Structure to Alexa Fluor™ 594*∼1600590/618Texas Red920000.66300 Tests23158
Note:
  1. Extinction Coefficient = molar extinction coefficient at their maximum absorption wavelength (Units = cm-1M-1).
  2. Quantum Yield = fluorescence quantum yield in aqueous buffer (pH 7.2).



Classic Dye-Labeled Phalloidin Conjugates

The following table outlines the fluorescence properties of available Phalloidin-AF dye conjugates for use in fluorescence imaging applications. For additional information on Phalloidin-AF dye conjugates



Table 4. Available Classic Dye-Labeled Phalloidin Conjugates.

Conjugate
Mol. Wt.
Ex/ Em (nm)
Filter Set
Extinction Coefficient¹
Quantum Yield²
Unit Size
Cat No.
Phalloidin-AMCA Conjugate∼1000346/434DAPI19000Not Determined300 Tests23100
Phalloidin-Fluorescein Conjugate∼1100498/517FITC800000.79300 Tests23101
Phalloidin-Tetramethylrhodamine Conjugate∼1300552/578TRITC90000Not Determined300 Tests23102
Phalloidin-California Red Conjugate∼1000592609100000Not Determined300 Tests23103
Note:
  1. Extinction Coefficient = molar extinction coefficient at their maximum absorption wavelength (Units = cm-1M-1).
  2. Quantum Yield = fluorescence quantum yield in aqueous buffer (pH 7.2).