Product Description
This Mouse 4-1BBL overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of 4-1BBL protein (Cat: 50067-M07H) from the overexpression lysate was verified.
Expression Host
HEK293 Cells
Sequence Information
A DNA sequence encoding the extracellular domain (Arg 104-Glu 309) of mouse TNFSF9 (NP_033430.1) precursor was expressed with a N-terminal polyhistidine tag.
Molecule Mass
The secreted recombinant mouse TNFSF9 consists of 222 amino acids and has a calculated molecular mass of 25 kDa. As a result of glycosylation, the recombinant protein migrates as an approximately 40-45 kDa protein in SDS-PAGE under reducing conditions.
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.
Mouse 4-1BB-L Overexpression Lysate;Mouse 4-1BBL Overexpression Lysate;Mouse AI848817 Overexpression Lysate;Mouse Cd137l Overexpression Lysate;Mouse Ly63l Overexpression Lysate
References
Cheung CT, (2007) Neutralizing anti-4-1BBL treatment improves cardiac function in viral myocarditis. Lab Invest. 87(7): 651-61. Wang C, et al. (2009) Immune regulation by 4-1BB and 4-1BBL: complexities and challenges. Immunol Rev. 229(1): 192-215. Habib-Agahi M, et al. (2009) 4-1BBL costimulation retrieves CD28 expression in activated T cells. Cell Immunol. 256(1-2): 39-46. 
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