Xylose

AS07 267 | Clonality: Polyclonal | Host: Rabbit | Reactivity: higher plants and algae


product information
Background		This antibody specifically cross-reacts against xylose residues bound to the protein N-glycans in beta1,2. This residue is characterisitc of the plant protein N-glycans and is absent in protein N-glycans from animals. This residue is added in the Golgi apparatus.
Immunogen		xylose residues bound to the N-glycan in beta 1,2
Host		Rabbit
Clonality		Polyclonal
Clone
Purity		Affinity purified serum in PBS, pH 7.4
Format		Lyophilized in PBS pH 7.4
Quantity		50 µg
Reconstitution		For reconstitution add 50 µl of sterile water
Storage		Store lyophilized/reconstituted at -20°C; once reconstituted make aliquots to avoid repeated freeze-thaw cycles. 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		Western blot (WB)
Related products		AS07 268 \| anti-fucose rabbit antibody Plant and algal protein extraction buffer Secondary antibodies
Additional information		Beta (1,2) xylose is present exclusively in plant N-glycans so antibodies against this sugar moiety should not cross-react with any mammal glycoprotein. This antibody do not bind free D-xylose. This antibody does not seem to work in immunolocalization.

application information
Recommended dilution		1 µg/ml (ELISA), 2 µg/10 ml incubation buffer (WB)
Expected \| apparent MW		10-100 for various glycoproteins
Confirmed reactivity		Higher plants and algae
Predicted reactivity		Higher plants
Not reactive in		No confirmed exceptions from predicted reactivity are currently known.
Additional information		Negative control: Fetuin, a glycoprotein containing fucose linked in alpha 1.6 and no xylose, Sigma, product number F3385. Positive control: Type II - horseradish peroxidase which contains β1.2 Xylose and α1.3 fucose, Sigma, product number P8250
Selected references		Nakanishi et al. (2017). Protection of Human Colon Cells from Shiga Toxin by Plant-based Recombinant Secretory IgA. Sci Rep. 2017 Apr 3;7:45843. doi: 10.1038/srep45843. (ELISA) Hanania et al. (2017). Establishment of a tobacco BY2 cell line devoid of plant specific xylose and fucose as a platform for the production of biotherapeutic proteins. Plant Biotechnol J. 2017 Feb 3. doi: 10.1111/pbi.12702. Ebert et al. (2015). Identification and Characterization of a Golgi-Localized UDP-Xylose Transporter Family from Arabidopsis. Plant Cell. 2015 Mar 24. pii: tpc.114.133827. Mathieu-Rivet et al. (2013). Exploring the N-glycosylation pathway in Chlamydomonas reinhardtii unravels novel complex structures. Mol Cell Proteomics, Aug 2.

Application example

Total cell extract from Arabidopsis thaliana wild type (1) and cell extracts from different mutants defective in the complex N-glycan maturation pathway (2-5) (data not published yet).

Primary antibody has been used at 2 µg/10 ml of incubation buffer. Detection has been done using ECL.

western blot using anti-xylose antibodies

Dot blot reaction of anti-Fucose and anti-Xylose antibodies with various controls: Avidin (Fuc+/Xyl+), Fetuin (Fuc-/Xyl-), PLA2 (Fuc+/Xyl-) and Mur1-2 (Fuc-/Xyl+). 2 µl of each extract were spotted on a nitrocellulose membrane, placed on top of 2 WHATMAN filters (one soaked in TBS-T) and dried for 1.5 h at RT. The mem-brane was blocked for 30 min with 2% low-fat milk powder in TBS-T (0.1% TWEEN 20) and incubated with anti-Fucose(1) (AS07 268, 1:1000) or anti-Xylose(2) (AS07 267, 1:1000) for 30 min and then with secondary anti-rabbit(1:1000) antibody (ALP conjugated, recommended secondary antibody AS09 607). Membrane was washed with TBS-T 3 x 10 minutes before reaction development using alkaline phosphatase reagent BCIP®/NTB premixed solution (Sigma, Prod. No. B6404).

Please follow this link for a more detailed Dot-Blot protocol

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