Alexa fluor 568 conjugated phalloidin

Manufactured by Thermo Fisher Scientific

Sourced in United States, Germany, Canada

Alexa Fluor 568-conjugated phalloidin is a fluorescent dye-labeled molecule used for labeling and visualizing F-actin (filamentous actin) in cells. It binds specifically to F-actin, allowing for the detection and imaging of the actin cytoskeleton.

Automatically generated - may contain errors

Lab products found in correlation

92 protocols using alexa fluor 568 conjugated phalloidin

Actin Stress Fiber Visualization

Check if the same lab product or an alternative is used in the 5 most similar protocols

Polymerized actin stress fibers were stained with Alexa Fluor 568-conjugated phalloidin (1:500), following manufacturer’s instructions (Thermo Fisher Scientific). Cell nuclei were counterstained with DAPI. Olympus BX51 microscope (100× magnification) was used for imaging.

Catalano S., Panza S., Augimeri G., Giordano C., Malivindi R., Gelsomino L., Marsico S., Giordano F., Győrffy B., Bonofiglio D., Andò S, & Barone I. (2019). Phosphodiesterase 5 (PDE5) Is Highly Expressed in Cancer-Associated Fibroblasts and Enhances Breast Tumor Progression. Cancers, 11(11), 1740.

+ Open protocol

+ Expand

Immunofluorescence Staining of Cellular TAZ

Check if the same lab product or an alternative is used in the 5 most similar protocols

Cells were cultured on chamber slide (ThermoFisher Scientific #177437) to appropriate density. Cells were fixed with 4% formaldehyde for 10 min and then wash three times with PBS. After blocking in 5% BSA and permeabilized with 0.1% Saponin for 1 hour, slides were incubated with the TAZ antibody (BD Biosciences #560235) 1:100 diluted in 1% BSA with 0.1% Saponin overnight. After washing with PBS, slides were incubated with Alexa Fluor 488-conjugated secondary antibodies (1:200 dilution, ThermoFisher Scientific #A11001) and Alexa Fluor 568-conjugated phalloidin (ThermoFisher Scientific #A12380) for 1 hour. The slides were then washed and mounted with SlowFade Glod antifade mountant with DAPI ((ThermoFisher Scientific #S36938). Images were acquired using a Leica TCS SP8 confocal microscope equipped with a 40X objective.

Fukayama S., Tashjian AH J.r., Davis J.N, & Chisholm J.C. (1995). Signaling by N- and C-terminal sequences of parathyroid hormone-related protein in hippocampal neurons.. Proceedings of the National Academy of Sciences of the United States of America, 92(22), 10182-10186.

+ Open protocol

+ Expand

Internalization of Fluorescent Microvesicles in Cell Lines

Check if the same lab product or an alternative is used in the 5 most similar protocols

RAW264.7, CMT-93, and Caco-2 cells were plated on a cover glass placed in a 24-well plate. In the case of CMT-93 and Caco-2, cells were starved in serum-free medium for 2 h before the addition of MVs. Cells were then treated with 50 or 100 μg/ml FITC-labeled MVs and incubated at 37°C for 1 h. After the incubation, cells were washed with PBS three times. Regarding immunofluorescence, cells were fixed in 3.7% (w/v) formaldehyde in PBS for 15 min and permeabilized with 0.1% (v/v) Triton X-100 in 1% (w/v) BSA-containing PBS for 15 min. Cells were then treated with Alexa Fluor 568-conjugated phalloidin (Thermo Fisher Scientific) and Hoechst 33342 (DOJINDO). After washing with PBS, cells were observed/photographed using the laser scan confocal microscope IX71 (PLYMPUS). To quantify internalized MVs, cells were lysed with passive lysis buffer (Promega). Hoechst 33342 (final 1/2,000) was added to the cell lysate, and fluorescence intensity was measured with the plate reader Spark (Tecan).

Kobayashi N., Abe K., Akagi S., Kitamura M., Shiraishi Y., Yamaguchi A., Yutani M., Amatsu S., Matsumura T., Nomura N., Ozaki N., Obana N, & Fujinaga Y. (2022). Membrane Vesicles Derived From Clostridium botulinum and Related Clostridial Species Induce Innate Immune Responses via MyD88/TRIF Signaling in vitro. Frontiers in Microbiology, 13, 720308.

+ Open protocol

+ Expand

AhR Localization in Hepatocyte Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Mouse Hepa-1c1c7 or human HepG2 cells were seeded onto eight-well chamber slides followed by incubation in tryptophan-deficient media [Hanks’ balanced salt solution (HBSS) with Ca²⁺ and Mg²⁺, 10% heat-inactivated FBS, 1× nonessential amino acids, 1× sodium pyruvate, and glucose (4.5 g/liter)] for 24 hours. Cells were subsequently treated with vehicle control (DMSO), 10 μM ITE, PY108, or PY109 for 90 min in tryptophan-deficient media. Cells were then fixed in 4% formaldehyde and processed according to a standard immunofluorescence staining protocol. Primary anti-AHR antibody was applied at 1:100 dilution (Enzo, BML-SA210) and Alexa Fluor 488–conjugated secondary antibody was applied at 1:500 dilution (Cell Signaling, 4412S). Actin was stained using Alexa Fluor 568–conjugated phalloidin (Thermo Fisher Scientific). Samples were mounted in anti-fade mounting medium supplemented with 4′,6-diamidino-2-phenylindole for nuclear staining and imaged with confocal microscopy (Leica SP5× MP, 63× oil lens).

Chen J., Haller C.A., Jernigan F.E., Koerner S.K., Wong D.J., Wang Y., Cheong J.E., Kosaraju R., Kwan J., Park D.D., Thomas B., Bhasin S., De La Rosa R.C., Premji A.M., Liu L., Park E., Moss A.C., Emili A., Bhasin M., Sun L, & Chaikof E.L. (2020). Modulation of lymphocyte-mediated tissue repair by rational design of heterocyclic aryl hydrocarbon receptor agonists. Science Advances, 6(3), eaay8230.

+ Open protocol

+ Expand

Hydrogel Immunofluorescence Staining Protocol

Check if the same lab product or an alternative is used in the 5 most similar protocols

Following vibrational stimulation, hydrogels were immediately rinsed with ice-cold PBS, then incubated in 4% PFA solution for 20 minutes. Once fixed, cell/gel constructs were washed twice with PBS containing 0.05% Tween-20 (PBST) then blocked in 3% bovine serum albumin (BSA) in PBS overnight at 4°C. After the blocking step, FITC-conjugated monoclonal α-smooth muscle actin (αSMA, Sigma) antibody (diluted 1:100) and Alexa Fluor 568-conjugated phalloidin (Thermo Fisher, diluted 1:400) in 3% BSA were added and allowed to incubate for 2 h at room temperature. Next, samples were washed in triplicate with PBST, and DAPI (Millipore, 1:500 dilution) was added for 10 minutes at room temperature. Samples were stored in PBS for imaging via confocal laser scanning microscopy (Zeiss LSM 880 with Airyscan).

Zerdoum A.B., Saberi P., Stuffer A.J., Kelly D.J., Duncan R.L., Mongeau L, & Jia X. (2020). Regulation of Stem Cell Function in an Engineered Vocal Fold-Mimetic Environment. Regenerative engineering and translational medicine, 6(2), 164-178.

+ Open protocol

+ Expand

Fixed Cell Cytometric Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

1 × 10⁶ cells in HBS were fixed using 4% paraformaldehyde (PFA) (cat no. 15710; Electron Microscopy Sciences, Hatfield, PA) for 20 min at room temperature. After washing, cell pellets were resuspended in flow buffer (HBS with 1% BSA) and incubated with 0.1% Triton X-100, a 1:250 dilution of Alexa Fluor 568-conjugated phalloidin (cat no. A22287; Thermo Fisher), and 1:1000 dilution of DAPI (cat no. D1306; Thermo Fisher) for 1 hr at room temperature. Cells were then re-pelleted, washed, and suspended in flow buffer. Data were acquired on a FACSCalibur (BD Biosciences, Franklin Lakes, NJ) flow cytometer. Flow cytometric analysis was performed using FlowJo (Ashland, OR) software.

Ullo M.F., D’Amico A.E., Lavenus S.B, & Logue J.S. (2023). The amoeboid migration of monocytes in confining channels requires the local remodeling of the cortical actin cytoskeleton by cofilin-1. bioRxiv.

+ Open protocol

+ Expand

Cofilin-1 Immunofluorescence in Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

After washing with HEPES buffered saline (HBS), cells in poly-L-lysine (cat no. P4707; Sigma Aldrich) coated 6-well glass-bottom plates (cat no. P06–1.5H-N; Cellvis) were fixed with 4% paraformaldehyde (PFA; Electron Microscopy Sciences) in HBS for 20 min at room temperature. Blocking, permeabilization, antibody incubation, and washing were done in HBS with 1% BSA, 1% fish gelatin, 0.1% Triton X-100, and 5 mM EDTA. A 1:250 dilution of cofilin-1 (cat no. MA5–17275; Thermo Fisher) antibody was incubated with cells overnight at 4°C. After extensive washing, a 1:400 dilution of Alexa Fluor 488-conjugated anti-rabbit secondary antibody (cat no. A-21206; Thermo Fisher) was then incubated with cells for 2 hr at room temperature. Cells were then incubated with a 1:250 dilution of Alexa Fluor 568-conjugated phalloidin (cat no. A12380; Thermo Fisher) and a 1:1000 dilution of DAPI (cat no. D1306; Thermo Fisher). Cells were again extensively washed and then imaged in HBS.

+ Open protocol

+ Expand

3D Collagen Gel Contraction Assay

Check if the same lab product or an alternative is used in the 5 most similar protocols

hSMECs or hS/PCs were suspended at a density of 3 × 10⁵ cells/ml in a collagen I solution (Corning Inc.) (3 mg/mL) at 4°C, and aliquots of 200 µl of cell suspension were added to BSA-coated wells of a 24-well plate (Cell Treat, Pepperell, MA). Samples were incubated at 37°C for 30 min before the addition of 500 µl FBS-free RPMI cell culture media. Cells were allowed to spread in the collagen gel overnight before the initiation of contraction with the addition of FBS. After 16 h of incubation at 37°C, the cell culture media was changed with the same media supplied with 10% FBS, and images of the gels were taken at 0 min. Gels were then photographed at indicated time points, and the average gel diameter for each experimental group (n=8) was calculated. Following the completion of the experiments, collagen gels immediately fixed using 4% paraformaldehyde for 30 min and followed by three PBS washes. The collagen gels were subsequently treated with the permeabilization solution (PBS containing 0.1% TritonX-100) for 45 min, stained with AlexaFluor 568-conjugated phalloidin and DAPI (Thermo Fisher) for 1 h and 30 min, respectively. After multiple PBS washes. samples were imaged with ZEISS LSM 880 (ZEISS, Pleasanton, CA) confocal microscope under fluorescence and differential reflectance modes to visualize collagen fibril, cytoskeleton and nuclei.

Ozdemir T., Srinivasan P.P., Zakheim D.R., Harrington D.A., Witt R.L., Farach-Carson M.C., Jia X, & Pradhan-Bhatt S. (2017). Bottom-up Assembly of Salivary Gland Microtissues for Assessing Myoepithelial Cell Function. Biomaterials, 142, 124-135.

+ Open protocol

+ Expand

Immunostaining of Cytoskeletal Proteins

Check if the same lab product or an alternative is used in the 5 most similar protocols

3T3-L1 cells grown on a cover slip were fixed with 3% (w/v) paraformaldehyde for 10 minutes at room temperature or with 100% methanol for 2 minutes at -20°C, and immunostained with anti-β-tubulin and anti-GEF-H1 primary antibodies and Alexa Fluor 488- or Alexa Fluor 568-conjugated secondary antibodies as described previously [13 (link)]. The actin filaments were stained with Alexa Fluor 568-conjugated phalloidin (Thermo Fisher Scientific). Cells were also stained with 4',6-diamidino-2-phenylindole (DAPI) to visualize the nuclei. Immunofluorescence images were obtained with FluoView FV1000 confocal microscope system (Olympus, Tokyo).

Masubuchi Y., Nakagawa Y., Medina J., Nagasawa M., Kojima I., Rasenick M.M., Inagaki T, & Shibata H. (2017). T1R3 homomeric sweet taste receptor regulates adipogenesis through Gαs-mediated microtubules disassembly and Rho activation in 3T3-L1 cells. PLoS ONE, 12(5), e0176841.

+ Open protocol

+ Expand

Fixation and Staining of Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

1 × 10⁶ cells in HBS were fixed using 4% paraformaldehyde (PFA) (cat no. 15710; Electron Microscopy Sciences, Hatfield, PA) for 20 min at room temperature. After washing, cell pellets were resuspended in flow buffer (HBS with 1% BSA) and incubated with 0.1% Triton X-100, a 1:250 dilution of Alexa Fluor 568-conjugated phalloidin (cat no. A22287; Thermo Fisher), and 1:1000 dilution of DAPI (cat no. D1306; Thermo Fisher) for 1 h at room temperature. Cells were then re-pelleted, washed, and suspended in flow buffer. Data were acquired on a FACSCalibur (BD Biosciences, Franklin Lakes, NJ) flow cytometer. Flow cytometric analysis was performed using FlowJo (Ashland, OR) software.

Ullo M.F., D’Amico A.E., Lavenus S.B, & Logue J.S. (2024). The amoeboid migration of monocytes in confining channels requires the local remodeling of the cortical actin cytoskeleton by cofilin-1. Scientific Reports, 14, 10241.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!