Phalloidin Control , DyLight 554 conjugate

Phalloidin Control for IF, ICC, IHC

Synonyms:
DY554-Phalloidin, F-actin toxin, Amanita phalloides
Details
Type: Control
Label: DyLight 554
Tested Species Reactivity: Many (Many)
Ordering Information
Pierce Phalloidin Control , DyLight 554 conjugate
Product #
21834
Size
300 units
Price
$360.00
Purchase
Add Phalloidin Control , DyLight 554 conjugate to your cart.
Tested Applications Dilution *
Immunofluorescence (IF) 1 - 5 units/ml
Immunocytochemistry (ICC) 1 - 5 units/ml
Immunohistochemistry (IHC) Assay dependent
* Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own experiment using appropriate negative and positive controls.
Form Information
Form: Lyophilized
Concentration: 300units/ml
Purification: purified
Storage Buffer: methanol
Preservative: no preservative
Storage Conditions: -20°C
Product Specific Information
Format: 300 units of lyophilized DyLight 554-Phalloidin (300 units/ml) in methanol. Prepare stock solution by adding 1000 µL of pure methanol to the vial and gently mix.

21834 has been successfully used in immunofluorescence and immunohistochemistry. DyLight 554-Phalloidin has an excitation/emission of 551/572nm and molecular weight of 1313.5 g/mole.

DyLight 554-Phalloidin Stock Solution can be prepared in methanol, DMF, or DMSO and unused Stock Solution should be promptly stored at -20°C to minimize evaporation. Working solutions (per 96-well plate) can be prepared by diluting 20 µL of the Stock Solution in 5.98 ml of PBS and mixing well.

Typical staining procedure adds 50µl of Working Solution (i.e. 1 to 5 unit/ml) to a each well. Incubate cells in the dark for 30 minutes at room temperature (optimal staining times varies from 10 minutes to 3 hours depending on cell type). Aspirate and wash cells three times in PBS after incubation. If desired, probe with specific antibodies or dyes before using DyLight 554-Phalloidin.
General Information
Phalloidin is a bicyclic peptide that belongs to a family of toxins isolated from the deadly Amanita phalloides “death cap” mushroom and is commonly used as a counterstain (similar to DAPI or Hoechst) in cell biology and histology imaging applications to selectively label F-actin in fixed cells, permeabilized cells, and cell-free experiments. Labeled phalloidin conjugates have similar affinity for both large and small filaments and bind in a stoichiometric ratio of about one phallotoxin per actin subunit in both muscle and non-muscle cells. Phalloidins reportedly do not bind to monomeric G-actin, unlike some antibodies against actin. The dynamics of the actin polymerization in cells are important for a variety of cellular processes from cell motility to cell shape, from muscular contraction to cytokinesis, and more.
Product Images
  • Phalloidin Control (21834) in IF
    Phalloidin_DyLight-554_21834_Immunofluorescence_1.jpg

    Immunofluorescence with anti-Phalloidin  Control, DyLight 554 conjugate (21834) (Phalloidin_DyLight-554_21834_Immunofluorescence_1.jpg)

    Immunofluorescence with anti-Phalloidin Control, DyLight 554 conjugate (21834)

    Immunofluorescent analysis of Calreticulin (green) U2OS cells. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in PBS for 10 minutes at room temperature and blocked with 2% BSA (Product # 37525) in PBS + 0.1% Triton X-100 for 30 minutes at room temperature. Cells were probed with a Calreticulin polyclonal antibody (Product # PA3-900) at a dilution of 1:50 for at least 1 hour at room temperature, washed with PBS, and incubated with DyLight 488 goat anti-rabbit IgG secondary antibody (Product # 35552) at a dilution of 1:250 for 30 minutes at room temperature. Actin filaments (red) were stained with DyLight 554 Phalloidin (Product # 21834) at a dilution of 1:300 (1unit/ml final concentration) and nuclei (blue) were stained with Hoechst (Product # 62249) at a concentration of 1ug/mL. Images were taken on a Thermo Scientific ArrayScan VTI at 20X magnification.
  • Phalloidin Control (21834) in IF
    Phalloidin_DyLight-554_21834_Immunofluorescence_2.jpg

    Immunofluorescence with anti-Phalloidin  Control, DyLight 554 conjugate (21834) (Phalloidin_DyLight-554_21834_Immunofluorescence_2.jpg)

    Immunofluorescence with anti-Phalloidin Control, DyLight 554 conjugate (21834)

    Immunofluorescent analysis of Phalloidin in U2OS cells. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 2% BSA in PBS (Product # 37525) containing 0.1% Triton X-100 for 30 minutes at room temperature. Cells were probed with DyLight 554 Phalloidin (Product # 21834) or Alexa Fluor® 555 Phalloidin, each diluted to a final concentration of 1 unit/ml in PBS, for 30 minutes. Nuclei were stained with Hoechst 33342 dye (Product # 62249). Images were taken on a Zeiss Axio Observer Z1 microscope with a 20X objective.
(This product is for In Vitro experimental use only. Not for resale without express authorization.)
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