Immunofluorescence analysis of beta III tubulin (green) in the ectoderm derived from human ES cells. Embryoid bodies (EBs) were generated from the H9 embryonic stem cell line (WiCell Research Institute, WA09) using Gibco® KnockOut™ Serum Replacement (http://www.lifetechnologies.com/us/en/home/references/protocols/cell-culture/stem-cell-protocols/ipsc-protocols/embryoid-bodies-from-ipscs.html). After four days in suspension culture, EBs were plated on Geltrex™-coated tissue culture-treated polystyrene and continuously cultured for 21 days. EB cultures were then fixed and permeabilized according to the 3-Germ Layer Immunocytochemistry Kit (Product # A25538) and stained with anti-beta III-tubulin monoclonal antibody (Product # MA1-118, 1:200 dilution, 5 uL/mL final) at 4°C overnight. Secondary staining was completed using Alexa Fluor™ 488-conjugated anti-mouse IgG (Product # A11001) and DAPI (Product # D1306) for nuclear DNA (blue) for 1 h at room temperature. Images were taken on EVOS® FL Auto Imaging System at 10X magnification.
|Tested species reactivity||Bovine, Guinea pig, Hamster, Human, Mouse, Pig, Rabbit, Rat|
|Host / Isotype||Mouse / IgG2a|
|Immunogen||A synthetic peptide corresponding to amino acids 436-450 from rat neuronal specific beta-3 tubulin.|
|Storage buffer||PBS with 1mg/ml BSA, 30% glycerol|
|Contains||0.05% sodium azide|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||1:50|
|Immunohistochemistry (Paraffin) (IHC (P))||1:50-1:200|
|Immunoprecipitation (IP)||5 µg|
|Western Blot (WB)||1:500-1:2000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
MA1-118 was successfully used to detect E18 Sparague Dawley primary cortical neurons.
MA1-118 can be used for immunofluorescence analysis of beta III tubulin in the ectoderm derived from human embryonic stem cells.
Western blot analysis of MA1-118 detects an ~50 kDa protein in neuronal-type cells. In bovine, a unknown band at ~32 kDa is also detected. MA1-118 shows specificity to beta-3 Tubulin and is non-reactive to lysates from non-neuronal cell types (e.g. HeLa cell lysate).
Beta tubulins are one of two core protein families (alpha and beta tubulins) that heterodimerize and assemble to form microtubules. Beta-III tubulin is primarily expressed in neurons and may be involved in neurogenesis, axon guidance and maintenance. Mutations in this gene are the cause of congenital fibrosis of the extraocular muscles type 3. Beta-III tubulin was also detected in Sertoli cells of the testis and transiently in non-neuronal embryonic tissues. Glutamate residues at the C-terminus of beta III tubulin can be glutamylated. The precise function of such modifications is unclear. Tubulin is phosphorylated on Ser-172 by CDK1 during cell cycle progression. Ser-172 phosphorylation inhibits tubulin incorporation into microtubules.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.