Revision 6
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IP, IF-F, IF-IC, FC-FP

REACTIVITY:

H M R Mk Z

SENSITIVITY:

Endogenous

MW (kDa):

52

Source/Isotype:

Rabbit IgG

UniProt ID:

#P68363

Entrez-Gene Id:

10376

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:50
Immunofluorescence (Frozen) 1:800 - 1:1600
Immunofluorescence (Immunocytochemistry) 1:800 - 1:1600
Flow Cytometry (Fixed/Permeabilized) 1:200 - 1:800

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Specificity / Sensitivity

Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb detects endogenous levels of α-tubulin only when acetylated at Lys40. This amino acid is not conserved in β-tubulin.

Species Reactivity:

Human, Mouse, Rat, Monkey, Zebrafish

Species predicted to react based on 100% sequence homology

Xenopus

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic acetylpeptide corresponding to residues surrounding Lys40 of human α-tubulin.

Background

The cytoskeleton consists of three types of cytosolic fibers: microtubules, microfilaments (actin filaments), and intermediate filaments. Globular tubulin subunits comprise the microtubule building block, with α/β-tubulin heterodimers forming the tubulin subunit common to all eukaryotic cells. γ-tubulin is required to nucleate polymerization of tubulin subunits to form microtubule polymers. Many cell movements are mediated by microtubule action, including the beating of cilia and flagella, cytoplasmic transport of membrane vesicles, chromosome alignment during meiosis/mitosis, and nerve-cell axon migration. These movements result from competitive microtubule polymerization and depolymerization or through the actions of microtubule motor proteins (1).
The Elongator complex catalytic subunit (Elp3) acetylates α-tubulin at Lys40 while the histone deacetylase HDAC6 functions as a tubulin deacetylase. This post-transcriptional modification may be required for dynamic cell shape remodeling, cell motility, tubulin stability and terminal branching of cortical neurons (2,3).

  1. Westermann, S. and Weber, K. (2003) Nat Rev Mol Cell Biol 4, 938-47.
  2. Creppe, C. et al. (2009) Cell 136, 551-564.
  3. Hubbert, C. et al. (2002) Nature 417, 455-458.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation IF-F: Immunofluorescence (Frozen) IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized)

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Revision 6
#5335

Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb

Western Blotting Image 1: Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb Expand Image
Western blot analysis of extracts from HeLa cells, untreated or TSA-treated (400 nM for 16 hours), using Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb (upper) and α-Tubulin (11H10) Rabbit mAb #2125 (lower). The acetyl-specificity of the antibody was verified by blocking with an acetyl- or non-acetylpeptide.
Immunoprecipitation Image 1: Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb Expand Image
Immunoprecipitation of acetyl-α-tubulin (Lys40) from HeLa extracts treated with Trichostatin A (TSA) #9950 (1μM, 18 hours). Lane 1 is 10% input, lane 2 is Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb, and lane 3 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900. Western blot analysis was performed with Acetyl-α-Tubulin (Lys40) (6-11 B-1) Mouse mAb #12152. Anti-mouse IgG, HRP-linked Antibody #7076 was used as a secondary antibody.
Immunofluorescence Image 1: Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of rat testes using Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb #5335 (green) and Pan-Keratin (C11) Mouse mAb #4545 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Immunofluorescence Image 1: Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of MCF10a cells, serum-starved (24 hr; left) or treated with Cytochalasin D (200 nM, 16 hr; right) using Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb (green). Actin filaments were labeled with DyLight 554 Phalloidin #13054 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Immunofluorescence Image 2: Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of HeLa cells, untreated (left) or treated with Trichostatin A #9950 (right), using Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb (green). Actin filaments have been labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Flow Cytometry Image 1: Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb Expand Image
Flow cytometric analysis of HeLa cells, untreated (blue) or treated with Trichostatin A (TSA) #9950 (400 nM, 16 hr; green) using Acetyl-α-Tubulin (Lys40) (D20G3) XP® Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.