Recombinant Mouse TSLP Protein, CF Summary
Product Specifications
Tyr20-Glu140, with a C-terminal 10-His tag
Analysis
17 kDa after deglycosylation, reducing conditions
Product Datasheets
Carrier Free
CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.
In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.
555-TSB
| Formulation | Supplied as a 0.2 μm filtered solution in PBS. |
| Shipping | The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below. |
| Stability & Storage: | Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
|
Assay Procedure
- Assay Buffer: 50 mM NaOAc, pH 4.5
- Recombinant F. meningosepticum Endo-beta -N-acetylglucosaminidase F3/Endo F3 (rFmEndo F3) (Catalog # 5548-GH)
- Substrate: Recombinant Mouse TSLP (rmTSLP) (Catalog # 555-TSB)
- 15% SDS-PAGE gel
- Reducing SDS-PAGE gel loading buffer (≥3X concentration)
- Silver Staining reagents
- BioRad GS-800 densitometer (or equivalent)
- Dilute rFmEndo F3 to 5, 2.5, 1.25, 0.625. 0.313, 0.156, 0.078 μg/mL in Assay Buffer.
- Dilute Substrate to 100 μg/mL in Assay Buffer.
- Combine 10 μL rFmEndo F3 at each dilution with 10 μL of 100 μg/mL Substrate. Include a control containing 10 μL Assay Buffer and 10 μL of 100 μg/mL Substrate.
- Incubate the reaction and control at 37 °C for 30 minutes.
- Add 10 μL of reducing gel buffer to each reaction. Boil sample at 100 °C for 3 to 5 minutes before loading to gel.
- Load all of the sample (30 μL) per lane on a 15% gel. As a staining development control load 25 ng BSA.
- Perform electrophoresis.
- Stain gel with silver stain, stop stain development when 25 ng BSA becomes visible.
- Analyze % deglycosylation by densitometry.
- Determine the DC50 for rFmEndoF3 by plotting % Substrate deglycosylated vs. amount with 4-PL fitting.
- rFmEndo F3: 50, 25, 12.5, 6.25, 3.13, 1.56, 0.78 ng
- rmTSLP: 1 μg
Reconstitution Calculator
Background: TSLP
Stromal Lymphopoietin (TSLP) was originally identified from the conditioned medium of a mouse thymic stromal cell line as a protein that promoted the development of B cells. The activity of mouse TSLP overlaps with, but is distinct from, that of mouse IL-7 (1). Mouse TSLP cDNA encodes a 140 amino acid (aa) residue precursor protein with a 19 aa signal sequence. Within the mature region, there are three potential N-glycosylation sites. The Sf 21 cell expressed recombinant mouseTSLP is likely to be glycosylated at all three sites, as three major glycoforms were visible on SDS-PAGE (Figure 1). Insect cells are known to express relatively simple and homogeneous N-glycans that mainly belong to the high mannose type (2). This recombinant protein was found to be an excellent substrate for N-specific glycosidases such as Endo F3 (Figure 1). The majority of the glycans on recombinant mouse TSLP can be readily removed by Endo F3. However, a small percentage of the glycans is somewhat resistant to Endo F3 digestion, possibly lacking core fucose, as it is known that core fucosylated N-glycans are strongly preferred substrates for Endo F3 digestion (3).
- Sims, J.E. et al. (2000) J. Exp. Med. 192:671.
- Staudacher, E. et al. (1992) Eur. J. Biochem. 207:987.
- Tarentino, A.L. and T.H. Jr. Plummer (1994) Glycobiology 4: 771.
Citation for Recombinant Mouse TSLP Protein, CF
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
1 Citation: Showing 1 - 1
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Intravenous gammaglobulin suppresses inflammation through a novel Th2 pathway.
Authors: Anthony RM, Kobayashi T, Wermeling F, Ravetch JV
Nature, 2011-06-19;475(7354):110-3.
Species: Mouse
Sample Types: In Vivo
Applications: In Vivo
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