Generation of Stable Cell Lines of APP695 (K595N, M596L) Gene Over-Expression via Lentiviral Over-Expression System
摘要
There are now various theories on the pathophysiology of Alzheimer’s disease (AD), the β-Amyloid (Aβ) cascade hypothesis has garnered a lot of interest. This text applied lentivirus-encapsulated plasmid to transfect APP695 (K595N, M596L) gene into SH-SY5Y cells and thereby obtain a stable cell line over-expressing Aβ. We used qPCR to monitor the mRNA expression of β-Amyloid Precursor Protein (APP) gene, β-Site APP Cleaving Enzyme 1 (BACE1) gene, we also used Western Blot to detect the APP protein, BACE1 protein, and Aβ1-42, respectively, in addition, the expression of Aβ1-42 in SH-SY5Y cell lysate and cell culture supernatant was measured by Enzyme-Linked Immunosorbent Assay (ELISA). The findings demonstrated APP gene over-expression following APP695 (K595N, M596L) gene transfection, and also indicated the successful establishment of the Swedish mutant cell model.