Phalloidin alexafluor 488

Manufactured by Merck Group

Phalloidin-AlexaFluor 488 is a fluorescent dye conjugate used for the detection and visualization of actin filaments (F-actin) in cells. It binds to F-actin with high affinity, allowing for the labeling and imaging of the actin cytoskeleton.

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2 protocols using phalloidin alexafluor 488

Murine L929 Cell Morphology and Viability on Scaffolds

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The scaffolds (diameter = 1.0 cm) were placed in 48-well plates. Murine L929 cells (3 × 10³ cells/well), were cultured in 1640 Medium with 10% fetal bovine serum (FBS). The cells were cultured on the scaffolds for 1, 3, and 5 days to observe cell morphology. Cell cytoskeleton organization was visualized by fluorescently staining with phalloidin-AlexaFluor 488 (Sigma-Aldrich) and DAPI (Sigma-Aldrich). Images were taken by the laser scanning confocal microscope (Leica, Germany). Cell counting kit-8 (CCK-8) assay was applied to detect cell viability after culture for 1, 3, and 5 days. Cell survival was investigated by live/dead staining at 1, 3, and 5 days (Invitrogen Live/Dead kit), and the quantification result was expressed as the following formula:

Cell ​survival = \frac{The ​number ​of ​living ​cells}{The ​total ​number ​of ​living ​cells ​and ​dead ​cells} \times 100 %

He X., Li W., Liu S., Li Y., Chen Y., Dan N., Dan W, & Zhu M. (2022). Fabrication of high-strength, flexible, porous collagen-based scaffolds to promote tissue regeneration. Materials Today Bio, 16, 100376.

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Inhibition of Cytoskeletal Dynamics

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aIIbb3 inhibition: Abciximab (C7E3-Fab) from Janssen and Tirofiban from Ibigen, myosin IIa inhibition: blebbistatin(−) (Cayman Chemical) (50 µM); blebbistatin(+) (Cayman Chemical) was used as a negative control. To avoid fluorescence-induced toxicity, para-nitroblebbistatin (50 µM, Optopharma) was used in a subset of experiments. Arp2/3 complex inhibition: CK666 (200 µM) and negative control: CK689 (both Merck Millipore). Actin polymerization inhibition: cytochalasin D (2.5 µM) (Sigma). Other: carboxyfluorescein succinimidyl ester (CFSE), Annexin V, busulfan, plasmin, thrombin, Fluo-4 AM, LPS O111:B4, CCL-2 (Sigma) iron (III) chloride (FeCl₃), casein, recombinant human albumin (HSA), fibrinogen from human plasma, unconjugated and conjugated to AlexaFluor 488 and 594, Rhodamine 6G, adenosine diphosphate (ADP), NP40, polyethylene glycol 400 (PEG400), phalloidin-AlexaFluor 488, hexamethyldisilazane (HMDS), paraformaldehyde (PFA), GDA, and LIVE/DEAD BacLight Bacterial Viability Kit were purchased from Sigma-Aldrich. Horm collagen was from Takeda, prostaglandin I2 sodium salt from Abcam, and U46619 from Tocris.

Nicolai L., Schiefelbein K., Lipsky S., Leunig A., Hoffknecht M., Pekayvaz K., Raude B., Marx C., Ehrlich A., Pircher J., Zhang Z., Saleh I., Marel A.K., Löf A., Petzold T., Lorenz M., Stark K., Pick R., Rosenberger G., Weckbach L., Uhl B., Xia S., Reichel C.A., Walzog B., Schulz C., Zheden V., Bender M., Li R., Massberg S, & Gaertner F. (2020). Vascular surveillance by haptotactic blood platelets in inflammation and infection. Nature Communications, 11, 5778.

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