The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration. PubMed: 26181491
1/1000. Detects a band of approximately 50 kDa (predicted molecular weight: 50 kDa).
1/100. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Tubulin is the major constituent of microtubules. It binds two moles of GTP, one at an exchangeable site on the beta chain and one at a non-exchangeable site on the alpha chain.
Ubiquitously expressed with highest levels in spleen, thymus and immature brain.
Cortical dysplasia, complex, with other brain malformations 6 Skin creases, congenital symmetric circumferential, 1
Belongs to the tubulin family.
The highly acidic C-terminal region may bind cations such as calcium.
Some glutamate residues at the C-terminus are polyglutamylated, resulting in polyglutamate chains on the gamma-carboxyl group (PubMed:26875866). Polyglutamylation plays a key role in microtubule severing by spastin (SPAST). SPAST preferentially recognizes and acts on microtubules decorated with short polyglutamate tails: severing activity by SPAST increases as the number of glutamates per tubulin rises from one to eight, but decreases beyond this glutamylation threshold (PubMed:26875866). Some glutamate residues at the C-terminus are monoglycylated but not polyglycylated due to the absence of functional TTLL10 in human. Monoglycylation is mainly limited to tubulin incorporated into axonemes (cilia and flagella). Both polyglutamylation and monoglycylation can coexist on the same protein on adjacent residues, and lowering glycylation levels increases polyglutamylation, and reciprocally. The precise function of monoglycylation is still unclear. Phosphorylated on Ser-172 by CDK1 during the cell cycle, from metaphase to telophase, but not in interphase. This phosphorylation inhibits tubulin incorporation into microtubules.
IHC image of beta Tubulin staining in a section of formalin-fixed paraffin-embedded normal human colon tissue performed on a Leica BONDTM. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20mins. The section was then incubated with ab21058, 1/100 dilution, for 15 mins at room temperature. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset negative control image is taken from an identical assay without primary antibody.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre
Western blot - Anti-beta Tubulin antibody - Loading Control (HRP) (ab21058)
All lanes : Anti-beta Tubulin antibody - Loading Control (HRP) (ab21058) at 1/1000 dilution
Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate Lane 2 :A431 whole cell lysate (ab7909)
Lysates/proteins at 20 µg per lane.
Developed using the ECL technique.
Performed under reducing conditions.
Predicted band size: 50 kDa Observed band size: 50 kDa
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