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Tubulin gamma chain (monoclonal antibodies)

AS20 4482  |  Clonality: Monoclonal  |  Host: Mouse |  Reactivity: Chlorella vulgaris, Nicotiana tabcum, Human, Protozoa

Tubulin gamma chain (monoclonal antibodies) in the group Antibodies for Plant/Algal  / Developmental Biology / Cytoskeleton at Agrisera AB (Antibodies for research) (AS20 4482)

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Product Information

Immunogen KLH-conjugated gamma-tubulin peptide EYHAATRPDYISWGTQ, amino acids 434-449. UniProt: P23258
The epitope was located in the aminoacid sequence PDYISW (aa441-446 in human), which is identical for gamma-tubulin 1 and gamma-tubulin 2.
Host Mouse
Clonality Monoclonal
Subclass/isotype IgG1
Purity Purified by Protein A affinity chromatography in PBS with 15 mM sodium azide.
Format Liquid
Quantity 100 µg
Storage Store at 4°C; Do not freeze.
Do not exceed exipry date is provided on the tube. 
Please, remember to spin tubes briefly prior to opening them to avoid any losses that might occur from lyophilized material adhering to the cap or sides of the tubes.

Tested applications Immunocytochemistry (ICC), Western blot (WB)
Recommended dilution 2-8 µg/ml (ICC), 1 µg/ml (WB)
Expected | apparent MW

51 kDa

Reactivity

Confirmed reactivity Chlorella vulgaris, Nicotiana tabacum, human, protozoa
Predicted reactivity Species of your interest not listed? Contact us
Not reactive in No confirmed exceptions from predicted reactivity are currently known.

Application examples

Application examples Western blot using monoclonal anti-tubulin gamma chain antibodies on plant and algal samples

Samples:
1 - 14 µg of Nicotiana tabacum, L. whole leaf extract of adult plant- control (untreated)
2 - 14 µg of Nicotiana tabacum, L. whole leaf extract of adult plant after treatment with 100 µM AgNP-PVP
3 - 14 µg of Nicotiana tabacum, L. whole leaf extract of adult plant after treatment with 100 µM AgNP-CTAB
4 - 14 µg of Nicotiana tabacum, L. whole leaf extract of adult plant after treatment with 100 µM AgNO3
5 - 14 µg of Nicotiana tabacum, L. whole leaf extract of adult plant after treatment with 100 µM AgNP-PVP+cys
6 - 14 µg of Nicotiana tabacum, L. whole leaf extract of adult plant after treatment with 100 µM AgNP-CTAB+cys
7 - 14 µg of Nicotiana tabacum, L. whole leaf extract of adult plant after treatment with 100 µM AgNO3+cys
8 - 14 µg of Nicotiana tabacum, L. extract of 4 weeks old seedlings- control (untreated)
9 – 14 µg of Chlorella vulgaris extract- control (untreated)

14 µg/well of total protein extracted freshly from 0.05 g of lyophilized whole leaf and root of adult tobacco plants following the Phenol extraction protocol1 with extraction buffer containing Trizma base (500 mM), Ethylenediaminetetraacetic acid (EDTA) (50 mM), sucrose (700 mM) and Potassium chloride (KCl) (100 Mm) with addition of phenylmethylsulfonyl fluoride (PMSF) (1mM) and 2% β-mercaptoethanol. After short incubation on 4°C with agitation, phenol was added. The phenol (supernatant) phase containing proteins, was collected after centrifugation and equal volume of extraction buffer was added. After centrifugation, supernatant phase was collected and 4 volumes of 0.1 M ammonium acetate (with 10% methanol) was added, and proteins were precipitated ON/-20°C. The next day, protein pellets were washed 3 times in ammonium acetate with rounds of centrifugations in between, and finally in aceton. Protein pellet was lastly resuspended in Isoelectric focusing buffer (IEF) containing 9 M urea, 4% CHAPS, 20 mM DTT, 1.2% Ampholytes pH 3 to 10. Protein concentrations was measured with modified Bradford method1 and denatured with Laemmli sample buffer2 at 95°C for 5 min. Total proteins were separated on 12 % SDS-PAGE and blotted 1h to nitrocellulose (pore size of 0.2 µm), using wet transfer. Blot was blocked with 2 % milk in PBS-T, 1h/RT. Blot was incubated in the primary antibody at a dilution of 1: 1 000 for 1h/RT with agitation in a solution of 2 % milk in PBS-T and then ON/4°C. The antibody solution was decanted and the blot was then washed 3 times for 10 min in 2% milk in PBS-T at RT with agitation. Blot was incubated in Agrisera matching secondary antibody (goat anti-Mouse IgG1 HRP conjugated, AS16 3715) diluted to 1:10 000 in 2% milk in PBS-T for 1h/RT with agitation. The blot was washed twice for 10 min in PBS-T developed for 5 min with AgriseraECLSuperBright (AS16 ECL-S). Exposure time was 12 min.

Courtesty of MSc, Karla Košpić, University of Zagreb, Faculty of Science Department of Biology, Croatia

Additional information

This antibody recognizes C-terminus (amino acids 434-449 in human) of gamma-tubulin, a 48 kDa structural constituent of cytoskeleton and microtubule organizing center (MTOC).
The epitope which this antibody is recognizing is conserved in Arabidopsis thaliana Tubulin gamma-1 chain, UniProt: P38557, Gene ID: At3g61650 and Tubulin gamma-2 chain, UniProt: P38558, Gene ID:  At5g05620

Recommended secondary antibody: goat anti-mouse IgG1 AS16 3715

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Background

Background

Tubulin gamma chain is essential for the control of microtubular network. It is found at microtubule organizing centers (MTOC) such as the spindle poles

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