Alexa fluor plus 555 phalloidin

Manufactured by Thermo Fisher Scientific

Sourced in United States

Alexa Fluor™ Plus 555 Phalloidin is a fluorescent label used to detect and visualize F-actin, a structural component of the cytoskeleton in eukaryotic cells. It binds specifically to F-actin and emits a bright orange-red fluorescence when excited at the appropriate wavelength.

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Lab products found in correlation

Hoechst 33342, by Solarbio (2 mentions) Cyanine3 (cy3), by Jackson ImmunoResearch (2 mentions) Mitotracker green fm, by Thermo Fisher Scientific (1 mentions) Confocal microscope, by Zeiss (1 mentions) Alexa fluor 488 phalloidin, by Thermo Fisher Scientific (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions) P96 plate wells, by Ibidi (1 mentions) Cell light edu apollo488 in vitro kit, by RiboBio (1 mentions)

Spelling variants of this product

Alexa Fluor 555 (1070 citations) Alexa 555 (299 citations) Alexa Fluor 555 Phalloidin (157 citations) Alexa Fluor Plus 555 (61 citations) Alexa Fluor phalloidin (43 citations) Alexa Fluors (21 citations) Alexa 555-plus (6 citations) Phalloidin 555 (6 citations) PLUSTM (3 citations) Phalloidins (2 citations)

4 protocols using alexa fluor plus 555 phalloidin

Immunofluorescent Staining of Fly Testes

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Testes from flies were dissected in 1 × PBS, fixed in 4% paraformaldehyde (PFA) for 30 min, washed three times using 1% PBS-Triton X-100 (PBST), and incubated for 30 min in 5% bovine serum albumin (BSA). The testes were then incubated with primary antibodies (see Table S4 for details) diluted in 5% BSA for 1 h at 25 °C, and washed thrice using 1% PBST. Secondary antibodies conjugated with Cy3 or A647 (Jackson ImmunoResearch Laboratories, West Grove, PA, USA) were diluted with 5% BSA at 1:400, and incubated with the testes samples for 1 h at 25 °C in the dark. Following three washes with 1% PBST, the testes were stained for 5 min using Hoechst 33342 (1:800, C0031, Solarbio, Beijing, China) before finalizing.
The structure of IC in testes was stained using Alexa Fluor™ Plus555 Phalloidin (1:50; A30106, Invitrogen, Waltham, MA, USA). MitoTracker™ Green FM (M7514, Thermo Fisher Scientific, Waltham, MA, USA) was used to label the Mitochondria. JC-1 (C2006, Beyotime, Shanghai, China) was used as a mitochondrial membrane potential marker. Briefly, fly testes were dissected in 1 × PBS and incubated with related working solutions according to the manufacturer's instructions. Testes were then washed three times with 1 × PBS. Images were captured on Zeiss confocal microscope (Carl Zeiss, Oberkochen, Germany) and processed using Adobe Photoshop CS5 software (Adobe, San Jose, CA, USA).

Yu J., Li Z., Fu Y., Sun F., Chen X., Huang Q., He L., Yu H., Ji L., Cheng X., Shi Y., Shen C., Zheng B, & Sun F. (2023). Single-cell RNA-sequencing reveals the transcriptional landscape of ND-42 mediated spermatid elongation via mitochondrial derivative maintenance in Drosophila testes. Redox Biology, 62, 102671.

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Polymerization and Labeling of F-Actin

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F-actin was polymerized fresh for each experiment from G-actin in G-Mg (2 mM Tris-Cl pH 8.0, 0.5 mM DTT, 0.2 M ATP, 0.1 mM MgCl₂, 0.01% NaN₃) buffer with KMEI (50 mM KCl, 1 mM MgCl₂, 1 mM EGTA, 10 mM imidazole pH 7.0, 1 mM DTT) added to initiate polymerization as described previously (56 ). Polymerization was left to occur at room temperature for 1-2 hours. For TIRF microscopy, polymerized F–actin was incubated with fluorescently labelled phalloidin (Alexa Fluor^™ Plus 555 Phalloidin or Alexa Fluor^™ 488 Phalloidin, Invitrogen) in a 1:1.2 (actin:phalloidin) molar ratio for 10 min at room temperature before being placed on ice.

Levin J.T., Pan A., Barrett M.T, & Alushin G.M. (2023). A platform for dissecting force sensitivity and multivalency in actin networks. bioRxiv.

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Uptake Dynamics of Fluorescent Beads in RPE-iPS Cells

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Protocol was adapted from Peng et al., 2017 [123 (link)] and Toulis et al., 2020 [46 (link)]. Briefly, 2 × 10⁵ cells RPE-iPS cells were seeded in p96 µ-plate wells (IbidiGmbh, Gräfelfing, Germany) and added 5 × 10⁶ FluoSpheres per well (Amine-Modified Microspheres, 0.2 µm, yellow-green fluorescent 505/515, Thermofisher Scientific). Cells were incubated for 4, 8, or 24 h and rinsed with warm PBS six times. Then, they were fixed with 4% paraformaldehyde for 15 min, permeabilized with 1% Triton X-100 for 15 min, and blocked with FBS 20% + 0.1% Triton x-100 for 15 min. Cells were finally stained with alexa Fluor Plus 555 phalloidin (Invitrogen) and DAPI (Thermofisher Scientific). Finally, images were acquired with a ZEISS Axio Vert.A1 (Carl Zeiss Sports Optics) and the cell nuclei and the green spheres were counted with ImageJ.

Navinés-Ferrer A., Ruiz-Nogales S., Navarro R, & Pomares E. (2022). Impaired Bestrophin Channel Activity in an iPSC-RPE Model of Best Vitelliform Macular Dystrophy (BVMD) from an Early Onset Patient Carrying the P77S Dominant Mutation. International Journal of Molecular Sciences, 23(13), 7432.

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Immunofluorescent Analysis of Fly Testes

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Fly testes were dissected in 1 × phosphate-buffered saline (PBS) and fixed for 30 min in 4% paraformaldehyde (PFA). They were washed three times with 0.3% PBS-Triton X-100 (PBST) and incubated in 5% bovine serum albumin (BSA) for 30 min. Primary antibodies (Additional file 4: Table S3) were diluted in 5% BSA solution, and testes were incubated at room temperature for 1 h and then washed three times in 0.3% PBST. Secondary antibodies were conjugated with A488, Cy3, or A647 (Jackson ImmunoResearch Laboratories, West Grove, PA, USA) and were diluted at a ratio of 1:400 with 5% BSA, and incubated at room temperature for 1 h avoiding light. Testes were then washed three times again by 0.3% PBST and stained with Hoechst 33342 (1.0 mg/mL, C0031, Solarbio, Beijing, China) for 5 min before finalizing. According to the manufacturer’s instructions, F-actin and EdU staining were performed with Alexa Fluor™ Plus 555 Phalloidin (1:50; A30106, Invitrogen, Waltham, MA, USA) and Cell-Light™ EdU Apollo488 In Vitro Kit (C10310-3, RiboBio, Guangzhou, China), respectively.

Li Z., Wu Y., Fu Y., Chen X., Zhao X., Wu X., Lu Y., He H., Shen C., Zheng B., Yu J, & Sun F. (2022). Cyst stem cell lineage eIF5 non-autonomously prevents testicular germ cell tumor formation via eIF1A/eIF2γ-mediated pre-initiation complex. Stem Cell Research & Therapy, 13, 351.

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