Apolipoprotein B Secretion Assay from Primary Hepatocytes.

ABSTRACT

Apolipoprotein B (APOB) is the primary structural protein of atherogenic lipoproteins, which drive atherogenesis and thereby lead to deadly cardiovascular diseases (CVDs). Plasma levels of APOB-containing lipoproteins are tightly modulated by LDL receptor-mediated endocytic trafficking and cargo receptor-initiated exocytic route; the latter is much less well understood. This protocol aims to present an uncomplicated yet effective method for detecting APOB/lipoprotein secretion. We perform primary mouse hepatocyte isolation and culture coupled with well-established techniques such as immunoblotting for highly sensitive, specific, and semi-quantitative analysis of the lipoprotein secretion process. Its inherent simplicity facilitates ease of operation, rendering it a valuable tool widely utilized to explore the intricate landscape of cellular lipid metabolism and unravel the mechanistic complexities underlying lipoprotein-related diseases. Key features • A pipeline for the isolation and subsequent culture of mouse primary hepatocytes. • A procedure for tracking the secretion of APOB-containing lipoproteins. • A rapid and sensitive assay for detecting the APOB level based on immunoblotting.