This Human TSPAN1 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of TSPAN1 protein (Cat: 13073-H04H) from the overexpression lysate was verified.
A DNA sequence encoding the human TSPAN1 (NP_005718.2) (Tyr110-Asn211) was expressed with the Fc region of mouse IgG1 at the N-terminus.
The recombinant human TSPAN1 consists 338 amino acids and predicts a molecular mass of 38.4 kDa.
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.
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.
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 NET1 Overexpression Lysate;Human TM4C Overexpression Lysate;Human TM4SF Overexpression Lysate;Human TSPAN1 Overexpression Lysate
TSPAN1 belongs to the transmembrane 4 superfamily, also known as the tetraspanin family. Tetraspanins have four hydrophobic domains, intracellular N- and C-termini and two extracellular domains. Tetraspanins act as scaffolding proteins, anchoring multiple proteins to one area of the cell membrane. They also mediate signal transduction events that play a role in the regulation of cell development, activation, growth and motility. TSPAN1 interacts with human thiamine transporter-1 (hTHTR-1). HTHTR-1 contributes to intestinal thiamine uptake, and its function is regulated at both the transcriptional and posttranscriptional levels. TSPAN1 and hTHTR-1 colocalize in human intestinal epithelial HuTu-80 cells. Coexpression of TSPAN1 in these cells led to a significant decrease in the rate of degradation of hTHTR-1 compared with cells expressing the hTHTR-1 alone; in fact the half-life of the TSPAN1 protein was twice longer in the former cell type compared with the latter cell type.
Chen L. et al., 2010, J Korean Med Sci. 25 (10): 1438-42. Chen L. et al., 2010, Tumori. 96 (5): 744-50. Nabokina SM. et al., 2011, Am J Physiol Gastrointest Liver Physiol. 301 (5): G808-13.