Human Granzyme B HEK293 Overexpression Lysate 产品信息
Product Description
This Human Granzyme B overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of Granzyme B protein (Cat: 10345-H08H) from the overexpression lysate was verified.
Expression Host
HEK293 Cells
Sequence Information
A DNA sequence encoding the proform of human Granzyme B (NP_004122.1) (Met 1-Tyr 247) was expressed with a C-terminal polyhistidine tag.
Molecule Mass
The recombinant human Granzyme B consists of 240 amino acids and has a calculated molecular mass of 27 kDa. As a result of glycosylation, the apparent molecular mass of rhGranzyme B is approximately 36 kDa in SDS-PAGE under reducing conditions.
Human Granzyme B HEK293 Overexpression Lysate Usage Guide
Preparation Method
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.
Lysis Buffer
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
Recommend Usage
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.
Sample Buffer
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.
Application
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.
Human Granzyme B HEK293 Overexpression Lysate Alternative Names
Human CCPI Overexpression Lysate;Human CGL-1 Overexpression Lysate;Human CGL1 Overexpression Lysate;Human CSP-B Overexpression Lysate;Human CSPB Overexpression Lysate;Human CTLA1 Overexpression Lysate;Human CTSGL1 Overexpression Lysate;Human HLP Overexpression Lysate;Human SECT Overexpression Lysate
Granzyme B Background Information
Granzyme B, also known as GZMB, is the most prominent member of the granzyme family of cell death-inducing serine proteases expressed in the granules of cytotoxic T lymphocytes (CTLs) and NK cells. Granzyme B enters the target cells depending on another membrane-binding granule protein, perforin, results in the activation of effector caspases and mitochondrial depolarization through caspase-dependent and -independent pathways, and consequently induces rapid cell apoptosis. Over 3 substrates of GZMB have been identified including the key substrate caspase-3, ICAD and Bid. GZMB is suggested to protect the host by lysing cells bearing on their surface 'nonself' antigens such as bacterial and viral infected-cells and tumor cells, and accordingly plays an essential role in immunosurveillance.
Full Name
granzyme B (granzyme 2, cytotoxic T-lymphocyte-associated serine esterase 1)
References
Schmid J., et al.,(1987), Induction of mRNA for a serine protease and a beta-thromboglobulin-like protein in mitogen-stimulated human leukocytes. J. Immunol. 139:250-256. Caputo A., et al., (1988), Structure and differential mechanisms of regulation of expression of a serine esterase gene in activated human T lymphocytes.J. Biol. Chem. 263:6363-6369. Trapani J.A., et al.,(1988), Molecular cloning of an inducible serine esterase gene from human cytotoxic lymphocytes.Proc. Natl. Acad. Sci. U.S.A. 85:6924-6928.
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