Product Description
This Human APOA4 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of APOA4 protein (Cat: 16082-H08H) from the overexpression lysate was verified.
Expression Host
HEK293 Cells
Sequence Information
A DNA sequence encoding the human APOA4 (NP_000473.2) (Met1-Ser396) was expressed with a polyhistidine tag at the C-terminus.
Molecule Mass
The recombinant human APOA4 consists 387 amino acids and predicts a molecular mass of 44.8 kDa.
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.
Apolipoprotein is genetically associated with the risk of Alzheimer's disease (AD). The APOA1, APOC3, and APOA4 genes are closely linked and located on human chromosome 11. There was a decreased trend for levels of APOA1, APOC3, and APOA4 in AD patients. CONCLUSION: Low levels of APOA1, APOC3, and APOA4 are associated with risk of AD. APOA1, APOC3, and APOA4 should be developed as combined drugs for the therapy of AD. SNP(single nucleotide polymorphisms)in APOA1and APOA4 genes influences atherogenic characteristics of LDL particles in response to diet.