Rabbit anti βiii tubulin

Manufactured by Merck Group

Sourced in United States

Rabbit anti-βIII-tubulin is a primary antibody that specifically recognizes the βIII-tubulin isoform, which is predominantly expressed in neurons. It can be used in various immunodetection techniques, such as Western blotting, immunohistochemistry, and immunocytochemistry, to identify and quantify βIII-tubulin in biological samples.

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Lab products found in correlation

Mouse anti βiii tubulin, by Merck Group (2 mentions) Rabbit anti gfap, by Merck Group (1 mentions) Axioplan 2 microscope, by Zeiss (1 mentions) Slowfade mounting media, by Thermo Fisher Scientific (1 mentions) Goat anti gfap, by Merck Group (1 mentions) Donkey anti chicken cy3, by Jackson ImmunoResearch (1 mentions) Hbo 100 lamp, by Zeiss (1 mentions) Axiovs40 v 4, by Zeiss (1 mentions) Axiocam mrm camera, by Zeiss (1 mentions) Mouse anti ha, by Proteintech (1 mentions) Mouse anti β actin, by Cell Signaling Technology (1 mentions) A4416, by Merck Group (1 mentions) Rabbit anti ha, by Cell Signaling Technology (1 mentions) Mouse anti flag, by Merck Group (1 mentions) Sp8 confocal microscope, by Leica (1 mentions) Rabbit anti s100β, by Agilent Technologies (1 mentions) Rabbit anti gap43, by Abcam (1 mentions) Fluorescent conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions) Mouse anti map2ab, by Merck Group (1 mentions) Rabbit anti laminin, by Merck Group (1 mentions)

Close spelling variants of this product

Anti-βIII-tubulin βIII-tubulin Rabbit anti-tubulin Anti-tubulin Rabbit anti-

7 protocols using rabbit anti βiii tubulin

Immunofluorescence Staining of Cultured Cells

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Cultured cells were fixed with 4% PFA for 15 minutes at room temperature (RT) and block-permeablised using PBST-10% Normal Donkey Serum (NDS) for 1 hour at RT. Primary and secondary antibodies were incubated in 3%NDS overnight (O/N) at 4 °C and 1 hour at RT respectively at the following dilutions; mouse anti-CNPAse, (1:2000; Chemicon, Germany), rabbit anti-GFAP, goat anti-GFAP, mouse anti-βIII-tubulin, rabbit anti-βIII-tubulin (all at 1:300; Sigma-Aldrich), rabbit anti-Pax6 (1:200; Chemicon, Germany), donkey anti-sheep Alexafluor555, donkey anti-rabbit Alexafluor488/555/647 and donkey anti-mouse Alexafluor488/555/647 (all 1:1000; Invitrogen), donkey anti-chickenCy3 (Jackson Laboratories, Bar Harbour, ME). Cells were counterstained with DAPI and mounted with Slow-fade mounting media (Invitrogen). Non-specific staining was controlled by using secondary-only controls. Fluorescence was viewed using the Axioplan2 microscope (Carl Zeiss, Jena, Germany) fitted with an HBO 100 lamp (Carl Zeiss, Jena, Germany). Images were captured using an Axiocam Mrm camera and Axio Vs40 v4.5.0.0 software (Axiovision, Carl Zeiss, Jena, Germany).

Bridges C.R., Tan M.C., Premarathne S., Nanayakkara D., Bellette B., Zencak D., Domingo D., Gecz J., Murtaza M., Jolly L.A, & Wood S.A. (2017). USP9X deubiquitylating enzyme maintains RAPTOR protein levels, mTORC1 signalling and proliferation in neural progenitors. Scientific Reports, 7, 391.

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Antibodies for TDP-43 Protein Analysis

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The following antibodies were used in Western blotting, immunoprecipitation, and immunofluorescence assays: rabbit anti–phospho-TDP-43 (Ser409/Ser410; Sigma-Aldrich; SAB4200225), mouse anti–pTDP-43 (pSer409/Ser410; COSMO BIO CO, TIP-PTD-M01), mouse anti-FLAG (Sigma-Aldrich; F3165), rabbit anti-Flag (Proteintech; 20543–1-AP), mouse anti-HA (Proteintech; 66006–1), rabbit anti-HA (Cell Signaling Technology; 3724), rabbit anti–TDP-43 (Proteintech; 10782–2-AP), mouse anti–TDP-43 (Abcam; ab57105), mouse anti-P62/SQSTM1 (Proteintech; 66184–1-Ig), rabbit anti-LC3B (Abcam; ab48394), rabbit anti-ATG8A/GABARAP (Abcam; ab109364), rabbit anti-Ref(2)P (Abcam; ab178440), rabbit anti-CK1α (Proteintech; 55192–1-AP), rabbit anti-CK1δ (Proteintech; 14388–1-AP), mouse anti-GAPDH (Proteintech; 60004–1), rabbit anti-tubulin (MBL; PM054), mouse anti–β-actin (Cell Signaling Technology; 3700), and rabbit anti–βIII-tubulin (Sigma-Aldrich; T2200). HRP-conjugated secondary antibodies included anti-mouse (Sigma-Aldrich; A4416) and anti-rabbit IgG (Sigma-Aldrich; A9169).

Deng X., Sun X., Yue W., Duan Y., Hu R., Zhang K., Ni J., Cui J., Wang Q., Chen Y., Li A, & Fang Y. (2021). CHMP2B regulates TDP-43 phosphorylation and cytotoxicity independent of autophagy via CK1. The Journal of Cell Biology, 221(1), e202103033.

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Immunohistochemical Analysis of Neuromuscular Endplates

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Preparation of the gastrocnemius muscles for immunohistochemistry was performed as described [46 (link)] with some modifications. Muscles were stained with α-Bungarotoxin-tetrametylrhodamine (10 μg/ml, Sigma) in PBS for 20 min for acetylcholine receptor (AChR) clusters. Rabbit anti-β-III tubulin (1:500, Sigma-Aldrich) in blocking solution was applied overnight at RT. As secondary Alexa 488-conjugated polyclonal anti-rabbit (1:100, Invitrogen) was diluted in the blocking solution and applied for 5 h at RT. Neuromuscular endplates of Gdap1^+/+ and Gdap^-/- mice were analyzed with a Leica SP8 confocal microscope for morphological analysis.

Barneo-Muñoz M., Juárez P., Civera-Tregón A., Yndriago L., Pla-Martin D., Zenker J., Cuevas-Martín C., Estela A., Sánchez-Aragó M., Forteza-Vila J., Cuezva J.M., Chrast R, & Palau F. (2015). Lack of GDAP1 Induces Neuronal Calcium and Mitochondrial Defects in a Knockout Mouse Model of Charcot-Marie-Tooth Neuropathy. PLoS Genetics, 11(4), e1005115.

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Immunohistochemical Analysis of Protein Markers

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Tissue slides were blocked with 10% donkey serum in PBS (1h) and were incubated with primary antibodies (Rabbit anti-YAP (Cell signaling; AB-2650491); Rabbit anti-TAZ (abclonal; AB-2721146); Rabbit anti-Hoxb7 (Novus; AB-2721144); Rabbit anti-S100β (Dako; AB-10013383); Rabbit anti-GAP43 (Abcam; AB-2247459); Rabbit anti-βIII-tubulin (Sigma-Aldrich AB-262133) diluted in 3% donkey serum (16h, 4°C). The next day, tissue slides were rinsed in PBS (3 × 5 min), were incubated with secondary antibodies coupled to biotin and diluted in 3% donkey serum (1h), were rinsed again in PBS (3 × 5 min), were incubated with a pre-mixture of avidin and biotin (following Vecta Stain Elite ABC kit procedure) (30 min), were rinsed again in PBS (3 × 5 min) and were visualized by adding the DAB substrate (following Vecta Stain Elite ABC kit procedure). Finally, reaction was quenched in distilled water and tissue slides were counterstained with hematoxylin, dehydrated and were cover slipped. A brown precipitate was deposited on positive cells.

Chen Z., Mo J., Brosseau J.P., Shipman T., Wang Y., Liao C.P., Cooper J.M., Allaway R.J., Gosline S.J., Guinney J., Carroll T.J, & Le L.Q. (2018). Spatiotemporal loss of NF1 in Schwann cell lineage leads to different types of cutaneous neurofibroma susceptible to modification by the Hippo pathway. Cancer discovery, 9(1), 114-129.

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Immunostaining of Motoneurons

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Motoneurons were first fixed with 4% paraformaldehyde (PFA) in PBS that was directly added to the culture medium (1:1) for 10 min, and then fixed with 4% PFA in PBS for 15 min on ice. Cells were then washed with PBS and incubated for 1 h in blocking solution containing 4% bovine serum albumin, 4% donkey serum, and 0.1% Triton-X100 in PBS. Coverslips were then incubated overnight at +4 °C with rabbit anti-βIII tubulin (Sigma-Aldrich, Saint-Louis, MO, USA, T2200). Cells were washed 3 times for 10 min each with PBS, incubated with the appropriate fluorescent-conjugated secondary antibody (ThermoFisher Scientific, Waltham, MA, USA), washed, and mounted in Moviol solution [45 (link)]. Image acquisition was carried out on a Zeiss (Car Zeiss AG, Oberkochen, Germany) Axio Imager Z2-module Apotome 2.0. ImageJ (National Institutes of Health, Bethesda, MD, USA) was used for axon tracing.

Younes R., Issa Y., Jdaa N., Chouaib B., Brugioti V., Challuau D., Raoul C., Scamps F., Cuisinier F, & Hilaire C. (2023). The Secretome of Human Dental Pulp Stem Cells and Its Components GDF15 and HB-EGF Protect Amyotrophic Lateral Sclerosis Motoneurons against Death. Biomedicines, 11(8), 2152.

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Immunostaining of Neuronal Cultures

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Neuronal cultures were fixed for 15 min in 4% paraformaldehyde in phosphate-buffered saline (PBS), and incubated for 20 min in blocking buffer (10% donkey serum in 0.1% Triton-X100 in PBS). They were then incubated overnight at 4°C with the primary antibodies in the blocking buffer diluted to 1∶10. Cell cultures were then incubated for 1 h at room temperature with secondary antibody in the blocking buffer 1∶10, followed by mounting in Mowiol. For neurite analysis, images were obtained with a PL-Neofluar Zeiss 10×0.3 or 20×0.8 objectives mounted on an upright Zeiss microscope equipped with a CCD camera AxioCam MRm, using AxioVision software and quantified with ImageJ (V1.47a). Primary antibodies used were mouse anti-βIII-tubulin (Sigma Aldrich; 1∶400), rabbit anti-βIII-tubulin (Sigma-Aldrich; 1∶400), rabbit anti-laminin (Sigma-Aldrich; 1∶100), mouse anti Map2a/b (Sigma-Aldrich; 1∶1000), chicken anti-GFP (Abcam, Cambridge, MA, USA; 1∶3000), and mouse anti SMI312 (Abcam; 1∶1000). Secondary antibodies were Alexa Fluor-488, Alexa Fluor-594 or Alexa Fluor-647 (Life Technologies, 1∶500).

Benzina O., Cloitre T., Martin M., Raoul C., Gergely C, & Scamps F. (2014). Morphology and Intrinsic Excitability of Regenerating Sensory and Motor Neurons Grown on a Line Micropattern. PLoS ONE, 9(10), e110687.

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Chromatin Immunoprecipitation Assay for KDM2B

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Neuro-2a were grown in MEM containing 10% FBS for 24 h and incubated with 1% formaldehyde for 10 min at 37°C. Cells were collected, lysed, and sonicated during 10 min at medium intensity in the ultrasonic processor GEX-600 (Sonics & Materials) and treated for ChIP as recommended by the manufacturer (Upstate). We used a rabbit anti-KDM2B (Millipore) and the unrelated rabbit anti-βIII tubulin (Sigma) antibodies. For PCR analysis, we used ChIPCK forward primer (5´-AGTTTTTGGCTTCCAGCAGA-3´) and ChIPCK reverse primer (5´-ACATTAGTCATGGTCACGCG-3´) [10 ]. PCR was performed using 5 μl of template DNA, 1.5 mM MgCl₂, and 20 pmol of each primer for 35 cycles at 94°C for 30 seg, 60°C for 30 seg, and 72°C for 30 seg.

Domizi P., Malizia F., Chazarreta-Cifre L., Diacovich L, & Banchio C. (2019). KDM2B regulates choline kinase expression and neuronal differentiation of neuroblastoma cells. PLoS ONE, 14(1), e0210207.

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