Human HSP70 Baculovirus-Insect cells Overexpression Lysate: Product Information
This Human HSP70 overexpression lysate was created in Baculovirus-Insect cells and intented for use as a Western blot (WB) positive control. Purification of HSP70 protein (Cat: 11660-H07B) from the overexpression lysate was verified.
A DNA sequence encoding the human HSPA1A (P08107) (Ala 2-Asp 641) was expressed, with a polyhistidine tag at the N-terminus.
The recombinant human HSPA1A consists of 658 amino acids and predicts a molecular mass of 72.2 kDa. It migrates as an approximately 60 kDa band in SDS-PAGE under reducing conditions.
Human HSP70 Baculovirus-Insect cells 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 HSP70 Baculovirus-Insect cells Overexpression Lysate: Alternative Names
Human HEL-S-103 Overexpression Lysate; Human HSP70-1 Overexpression Lysate; Human HSP70-1A Overexpression Lysate; Human HSP70I Overexpression Lysate; Human HSP72 Overexpression Lysate; Human HSPA1 Overexpression Lysate
HSP70 Background Information
HSPA1A is a member of the Hsp7 protein family. The 7 kilodalton heat shock proteins (Hsp7s) are a family of ubiquitously expressed heat shock proteins. HSP are abundant and conserved proteins present in all cells. Upon temperature shock or other stress stimuli, HSP are synthesized intracellularly, which may protect cells from protein denaturation or from death. Extracellularly, HSP can serve a cytokine function to initiate both innate and adaptive immunity through activation of APC. HSP serves also a chaperone function and facilitates presentation of antigen peptide to T cells. Molecular chaperones of the Hsp7 family have diverse functions in cells. They assist the folding of newly synthesized and stress-denatured proteins, as well as the import of proteins into organelles, and the dissociation of aggregated proteins. The well-conserved Hsp7 chaperones are ATP dependent: binding and hydrolysis of ATP regulates their interactions with unfolded polypeptide substrates, and ATPase cycling is necessary for their function. All cellular functions of Hsp7 chaperones use the same mechanism of ATP-driven polypeptide binding and release.
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