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Chitin Probes >
Rhodamine-conjugated Chitin-binding Probe |
 | | | | Rhodamine-conjugated Chitin-binding Probe | | | |
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Prices are in US dollars and valid only for US orders.
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Description:
Chitin, a homopolymer of β, 1-4 linked N-acetylglucosamine is the second most abundant polymer in nature. It is widely found among invertebrates including arthropods, nematodes and fungi, where it offers structural support and/or a protective barrier.
The chitin-binding probe is a recombinant fusion protein that binds specifically to chitin. The fusion protein comprises a small (5 kDa) chitin-binding domain (CBD) derived from the C-terminal region of chitinase A1 of Bacillus circulans WL-12 (1) fused to the C-terminus of maltose-binding protein (43 kDa) from E. coli (2). The fusion protein is labeled using tetramethylrhodamine isothiocyanate (TRITC) following standard methods. Labeled proteins are purified from free dye using a gel filtration column.
The CBD from B. circulans has an extremely high affinity for chitin (1) and has been used successfully as an affinity tag to enable the purification of recombinant proteins by a single chitin column (3).
The fluorescent chitin-binding probes may be used to specifically detect chitin in situ using standard fluorescence microscopy.
Recommended Working Dilution: Probe should be used at a 1:500 dilution in immunostaining buffers such as phosphate-or Tris-buffered saline. Sample may be prepared as in standard antibody staining protocols or other methods that allow the exposure of chitin (5). It is recommended to perform overnight staining at room temperature is recommended, although shorter incubation may be possible in certain samples. Standard fluorescence microscopy techniques are recommended for sample handling and image acquisition.
Source:
The CBD was cloned from the chitinase A1 from B. circulans WL-12 and fused to maltose-binding protein (MBP) (4). The fusion protein was overexpressed in E. coli at NEB.
Enzyme Properties
Molecular Weight:
Calculated: 48 kDa
Reaction & Storage Conditions
Concentration:
2 mg/ml
Storage Conditions:
50% Glycerol
50 µl phosphate-buffered saline
Storage Temperature:
-20°C
Quality Control for Current Lot
Quality control values for a specific lot can be found on the datacard which accompanies each product.
Quality Assurance Statement:
Purified to > 95% homogeneity as determined by SDS-PAGE. The degree of labeling is determined by calculating the moles of dye per mole of protein. The labeled fusion protein contains about 2-4 dye molecules per protein molecule.
References
- Watanabe, T. et al. (1994) J. Bacteriol., 176, 4465-4472.
- Guan, C. et al. (1998) Gene, 67, 21-30.
- Chong, S. et al. (1997) Gene, 192, 271-281.
- Xu, M., New England Biolabs, Inc., unpublished observations.
- Zhang, Y. et al. (2005) Dev. Biol., 285, 330-339.
Companion Products
Fluorescein-conjugated Chitin-binding Probe
TransPass™ P Protein Transfection Reagent
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