Beta tubulin

Manufactured by Developmental Studies Hybridoma Bank

Sourced in United States

Beta-tubulin is a protein that is a component of microtubules, which are cytoskeletal structures found in eukaryotic cells. Microtubules play a crucial role in various cellular processes, including cell division, intracellular transport, and the maintenance of cell shape and structure.

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Lab products found in correlation

Onsds page gels, by Thermo Fisher Scientific (2 mentions) Odyssey, by LI COR (2 mentions) Histone h3, by Santa Cruz Biotechnology (2 mentions) Flag m2, by Merck Group (2 mentions) Gapdh, by Santa Cruz Biotechnology (2 mentions) Protease and phosphatase inhibitor, by Roche (2 mentions) Flag antibody, by Merck Group (1 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions) Flag tag (flag), by Merck Group (1 mentions) Protease inhibitor, by Roche (1 mentions) Protease inhibitor, by Thermo Fisher Scientific (1 mentions) Casein blocking buffer, by Merck Group (1 mentions) Low fluorescent pvdf, by Bio-Rad (1 mentions) Odyssey imaging system, by LI COR (1 mentions) C myc clone 9e10, by Santa Cruz Biotechnology (1 mentions) P p70s6k, by Cell Signaling Technology (1 mentions) 4e bp1, by Cell Signaling Technology (1 mentions) P 4ebp1, by Cell Signaling Technology (1 mentions) P70s6k, by Cell Signaling Technology (1 mentions) Glut1, by Abcam (1 mentions)

5 protocols using beta tubulin

Immunoprecipitation of LRP6 and MESD in HEK293T Cells

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HEK293T cells were transfected with Lrp6 cDNA constructs and MESD-Flag using Lipofectamine ® 2000 reagent (Life Technologies) per manufacturer’s instructions with a 0.5:1 ratio of reagent to DNA. Cells were allowed to incubate with transfection media for 24 hours then were washed with PBS and harvested in TBS with 1% Triton × 100 with protease inhibitors (Roche). Cells were lysed by rocking at 4°C for 2 hours. Total protein concentrations of lysate were determined using a Bradford assay. After normalization to total protein, lysate was incubated with Flag antibody (Sigma) for 2 hours. Protein A/G agarose beads (Pierce) were added to mixture and incubated at 4°C overnight while rocking. Beads were washed 5 times with TBS with 0.5% Tween 20, then boiled in SDS loading buffer and supernatant loaded onto a 10% PAGE gel. Nitrocellulose membranes were used for western blotting. Beta Tubulin (Developmental Studies Hybridoma Bank, 1:500), Lrp6 (Cell Signaling, 1:2000), and Flag (Sigma, 1:500) primary antibodies were used. LiCor secondary antibodies were used at 1:10000 and protein bands were analyzed on a LiCor Odyssey CLx.

Lei Y., Fathe K., McCartney D., Zhu H., Yang W., Ross M.E., Shaw G.M, & Finnell R.H. (2015). Rare LRP6 variants identified in spina bifida patients. Human mutation, 36(3), 342-349.

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Hippocampal Protein Quantification Protocol

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Hippocampal lysates were prepared by homogenizing flash frozen subdissected hippocampi using RIPA buffer (150 mM NaCl, 50 mM Tris, pH 7.5, 1% Triton-X 100, 0.5% sodium deoxycholate, 1% sodium dodecyl sulfate) containing protease inhibitors (Fisher Scientific, Hampton, NH). Following BCA assay (Pierce), 20 μg of lysate was separated through 12% polyacrylamide gels and transferred to low-fluorescent PVDF (Biorad, Hercules, CA). Membranes were blocked with casein blocking buffer (Sigma-Aldrich, St. Louis, MO) in Tris buffered saline with 0.1% Tween 20 (TBST) and incubated with primary antibody (in 0.3% BSA in TBST) at 4°C overnight using antibodies to detect Bax (Fisher Scientific) or beta-tubulin (Developmental Studies Hybridoma Bank). Secondary antibodies conjugated to Alexa-680 (Fisher Scientific) allowed detection and quantification by scanning with an Odyssey Imaging System (Licor Biosciences, Lincoln, NE).

Adlaf E.W., Vaden R.J., Niver A.J., Manuel A.F., Onyilo V.C., Araujo M.T., Dieni C.V., Vo H.T., King G.D., Wadiche J.I, & Overstreet-Wadiche L. (2017). Adult-born neurons modify excitatory synaptic transmission to existing neurons. eLife, 6, e19886.

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Western Blotting Analysis of Protein Expression

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Western blotting of proteins was performed as described previously [20 (link)]. Briefly, for western blotting, cells were washed twice with cold PBS and lysed in radioimmuno precipitation assay (RIPA) lysis buffer by incubating at 4°C on a rotatory shaker for 30 min. Cell debris was removed by centrifugation at 13,000 rpm for 10 min and the supernatant was collected. Protein content was measured by Bradford assay. Equal amounts of denatured proteins were separated by electrophoresis using SDS-PAGE gels and transferred to activated PVDF membranes. The membranes were probed with primary antibodies against GLUT1 (Abcam, Cambridge, UK), c-Myc (clone 9E10; Santa Cruz Biotechnology, Dallas, Texas, USA), HKII, p P70S6K, P70S6K, p 4EBP1, 4EBP1 (Cell Signaling Technology, Danvers, MA, USA) and beta-tubulin (Clone E7 from Developmental Studies Hybridoma Bank, Iowa City, IA).

Shukla S.K., Gunda V., Abrego J., Haridas D., Mishra A., Souchek J., Chaika N.V., Yu F., Sasson A.R., Lazenby A.J., Batra S.K, & Singh P.K. (2015). MUC16-mediated activation of mTOR and c-MYC reprograms pancreatic cancer metabolism. Oncotarget, 6(22), 19118-19131.

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Protein Extraction and Immunoblotting

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Whole cell extracts were harvested using radioimmunoprecipitation (RIPA)
buffer supplemented with protease and phosphatase inhibitors (Roche) and run on
SDS-PAGE gels (Life Technologies) then transferred onto nitrocellulose
membranes. To prepare nuclear extracts, cells were resuspended in hypotonic
lysis buffer (10mM HEPES, pH7.9, 1.5mM MgCl₂, and 10mM KCl) and
dounced 15 times. Isolated nuclei were pelleted and resuspended in RIPA buffer.
Immunoblotting was carried out using antibodies against the following proteins:
GLI1 (Cell Signaling) (Supplementary Fig. 27), beta-tubulin (Developmental Studies
Hybridoma Bank), Flag M2 (Sigma), HA (Abcam) (Supplementary Fig. 28), GAPDH,
Histone H3, SRF, MKL1 (Santa Cruz Biotechnology). All immunoblots imaged using
the LI-COR Odyssey image system.

Whitson R.J., Lee A., Urman N.M., Mirza A., Yao C.Y., Brown A.S., Li J.R., Shankar G., Fry M.A., Atwood S.X., Hollmig S.T., Aasi S.Z., Sarin K.Y., Epstein EH J.r., Tang J.Y, & Oro A.E. (2018). Non-canonical hedgehog pathway activation by MKL1/SRF promotes drug-resistance in basal cell carcinomas. Nature medicine, 24(3), 271-281.

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Protein Extraction and Immunoblotting

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