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Fluorescence based tubulin polymerization assay kit

Manufactured by Cytoskeleton
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The Fluorescence-based tubulin polymerization assay kit is a laboratory tool used to measure the polymerization of tubulin, a key component of the cytoskeleton. The kit utilizes fluorescence detection to monitor the assembly and disassembly of tubulin polymers in real-time.

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Lab products found in correlation

Dtx880, by Beckman Coulter (1 mentions) Guava viacount, by Merck Group (1 mentions) Anti β actin, by Cell Signaling Technology (1 mentions) Ripa lysis buffer, by Beyotime (1 mentions) Bca protein assay kit, by Beyotime (1 mentions) Apo one homogeneous caspase 3 7 assay kit, by Promega (1 mentions) Wst 1, by Beyotime (1 mentions) Fluoronunc, by Thermo Fisher Scientific (1 mentions) Flexstation 3, by Molecular Devices (1 mentions) Softmax pro software, by Molecular Devices (1 mentions)

Close spelling variants of this product

Tubulin polymerization assay (3 citations) Tubulin Polymerization kit (11 citations)

5 protocols using fluorescence based tubulin polymerization assay kit

1

Fluorescence-based Tubulin Polymerization Assay

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Microtubule polymerization assays were performed using a fluorescence-based tubulin polymerization assay kit (Cytoskeleton, Denver, CO, USA) according to the manufacturer’s instructions. Briefly, tubulin was re-suspended in ice cold G-PEM buffer (80 mM PIPES, 2 mM MgCl2, 0.5 mM EGTA, 1 mM GTP, 20 % (v/v) glycerol) and added to wells on a 96-well plate containing the designated concentration of experimental compounds. The paclitaxel and vincristine were used as polymer stabilizer and depolymerizer controls, respectively. Samples were mixed well and tubulin assembly was measured (emission wavelength is 420 nm; excitation wavelength is 360 nm) using a plate reader (DTX880, Beckman) at 37 °C and recorded every 60 s for 30 min.
Xiang Q., Zhen Z., Deng D.Y., Wang J., Chen Y., Li J., Zhang Y., Wang F., Chen N., Chen H, & Chen Y. (2015). Tivantinib induces G2/M arrest and apoptosis by disrupting tubulin polymerization in hepatocellular carcinoma. Journal of Experimental & Clinical Cancer Research : CR, 34, 118.
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2

Fluorescence-based Tubulin Polymerization

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Microtubule polymerization assays were performed using a fluorescence-based tubulin polymerization assay kit (Cytoskeleton, Denver, CO, USA) according to the manufacturer's protocol.
Lv L.X., Zhou Z.X., Zhou Z., Zhang L.J., Yan R., Zhao Z., Yang L.Y., Bian X.Y., Jiang H.Y., Li Y.D., Sun Y.S., Xu Q.Q., Hu G.L., Guan W.J, & Li Y.Q. (2017). Hispidin induces autophagic and necrotic death in SGC-7901 gastric cancer cells through lysosomal membrane permeabilization by inhibiting tubulin polymerization. Oncotarget, 8(16), 26992-27006.
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3

Molecular Assays for Cell Proliferation

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The cell proliferation reagent WST-1 (#C0036), RIPA lysis buffer (#P0013C) and the BCA protein assay kit (#P0012) were purchased from Beyotime Institute Biotechnology (Songjiang, Shanghai, China). Guava Viacount (#4000–0040), Cell Cycle (#4500–0220) and Nexin reagents (#4500–0450) were purchased from Millipore Corporation (Billerica, MA, USA). Apo-ONE homogeneous caspase-3/7 assay kit (#G7792) was purchased from Promega (Madison, WI, USA). The fluorescence based tubulin polymerization assay kit (#BK011P) was purchased from Cytoskeleton (Denver, CO, USA). The anti-p21 (#2947), anti-PARP (#9542), anti-α-tubulin (#2144) and anti-β-actin (#4967) antibodies were purchased from Cell Signaling Technology (Beverly, MA, USA). The goat anti-mouse (#170–6516) or goat anti-rabbit (#170–6515) secondary antibodies were purchased from LI-COR Biotechnology (Lincoln, NE, USA). VCR and other chemical reagents were obtained from Sigma-Aldrich (St. Louis, MO, USA).
Mu Y., Liu Y., Li L., Tian C., Zhou H., Zhang Q, & Yan B. (2015). The Novel Tubulin Polymerization Inhibitor MHPT Exhibits Selective Anti-Tumor Activity against Rhabdomyosarcoma In Vitro and In Vivo. PLoS ONE, 10(3), e0121806.
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4

Nicotine Effects on Tubulin Polymerization

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The effects of nicotine on tubulin polymerization were assessed using the fluorescence‐based tubulin polymerization assay kit according to the manufacturer's instructions (Cytoskeleton, Denver, CO). Briefly, a buffer containing 2 mg/mL porcine brain tubulin, 80 mM PIPES pH 6.9, 2 mM MgCl2, 0.5 mM EGTA, 1 mM GTP, and 15% v/v glycerol was incubated with either 0.01–10 mM nicotine, 2.3 μM colchicine, or 3 μM paclitaxel in black, clear/flat‐bottom 96‐well plates (FluoroNunc™, Thermo Fisher Scientific) within a fluorimeter (FlexStation® 3; Molecular Devices, San Jose, CA) pre‐warmed to 37°C. Excitation and emission wavelengths of 360 and 420 nm, respectively, were used, and readings were taken every minute for a total of 60 min. Maximum velocity (Vmax) and maximum product values were subsequently extracted from the data using SoftMax® Pro software (v7.0 GxP, Molecular Devices).
Smart D.J., Helbling F.R., Verardo M., McHugh D, & Vanscheeuwijck P. (2019). Mode‐of‐action analysis of the effects induced by nicotine in the in vitro micronucleus assay. Environmental and Molecular Mutagenesis, 60(9), 778-791.
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5

Tubulin Polymerization Inhibition Assay

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A fluorescence-based tubulin polymerization assay kit (Cytoskeleton, Inc. Denver, CO) was used to test CH-2-102’s ability to inhibit tubulin polymerization. Specifically, 5 μL of 10× concentrations of controls and samples were aliquoted into a half-area 96-well plate that had been prewarmed (37 °C) (Corning Costar, Corning, NY). The plate was filled with 50 μL of tubulin mix containing GTP, glycerol, and tubulin protein. The reaction movements were measured using a SpectraMax M5e fluorescent microplate reader at a frequency of 30 s and 90 min. The excitation was at 360 nm, and the fluorescent signal was measured at 450 nm.
Erlich J., Parry G.C., Fearns C., Muller M., Carmeliet P., Luther T, & Mackman N. (1999). Tissue factor is required for uterine hemostasis and maintenance of the placental labyrinth during gestation. Proceedings of the National Academy of Sciences of the United States of America, 96(14), 8138-8143.
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