Human Cystatin S HEK293 Overexpression Lysate: Product Information
This Human Cystatin S overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of Cystatin S protein (Cat: 11542-H08H) from the overexpression lysate was verified.
A DNA sequence encoding the human CST4 (NP_001890.1) (Met 1-Ala 141) was fused with a polyhistidine tag at the C-terminus.
The secreted recombinant human CST4 consists of 132 amino acids and migrates as an approximately 16 kDa band in SDS-PAGE under reducing conditions as predicted.
Human Cystatin S HEK293 Overexpression Lysate: Usage Guide
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
1. Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube.
2. Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Western Blot (WB) Optimal dilutions/concentrations should be determined by the end user.
Human Cystatin S HEK293 Overexpression Lysate: Alternative Names
Human CST4 Overexpression Lysate; Human MGC71923 Overexpression Lysate
Cystatin S Background Information
Cystatin-S, also known as Cystatin-4, Salivary acidic protein 1, Cystatin-SA-III and CST4, is a secreted protein which belongs to thecystatin family. Cystatin-4 / CST4 is expressed in submandibular and sublingual saliva but not in parotid saliva (at protein level). It is also expressed in saliva, tears, urine and seminal fluid. The cystatin superfamily encompasses proteins that contain multiple cystatin-like sequences. Some of the members are active cysteine protease inhibitors, while others have lost or perhaps never acquired this inhibitory activity. There are three inhibitory families in the superfamily, including the type 1 cystatins (stefins), type 2 cystatins and the kininogens. The type 2 cystatin proteins are a class of cysteine proteinase inhibitors found in a variety of human fluids and secretions. Cystatin-4 / CST4 strongly inhibits papain and ficin, partially inhibits stem bromelain and bovine cathepsin C, but does not inhibit porcine cathepsin B or clostripain. Papain is inhibited non-competitively. Cystatin-4 / CST4 is an S-type cystatin, based on its high level of expression in saliva, tears and seminal plasma. The specific role in these fluids is unclear but antibacterial and antiviral activity is present, consistent with a protective function.
Bobek L.A., et al.,(1991), Human salivary cystatin S. Cloning, sequence analysis, hybridization in situ and immunocytochemistry. Biochem. J. 278:627-635.
Saitoh E., et al., (1992), Characterization of two members (CST4 and CST5) of the cystatin gene family and molecular evolution of cystatin genes.Agents Actions 38:340-348.
Deloukas P., et al.,(2001), The DNA sequence and comparative analysis of human chromosome 20.Nature 414:865-871.
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