Which casein micelle removal method is suitable for studies of human milk extracellular vesicles? A systematic comparison of four different treatments for casein depletion before extracellular vesicle isolation from human milk.

ABSTRACT: Aim: This study aimed to systematically compare four casein micelle removal methods on the particle and protein characteristics of the isolated human milk EVs. Methods: The defatted milk was treated with 1% sodium citrate, 20 mM ethylenediaminetetraacetic acid (EDTA), 1% acetic acid, or 1% chymosin/calcium chloride for 30 min at 4 °C to remove casein micelles. EV isolation was performed using qEV size exclusion chromatography. Milk turbidity at the optical density 350 nm and dot immunoblot with casein antibody were applied to monitor the qEV fractions. Particle analyses were performed using transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA). The enrichment of human milk EV markers, i.e., tetraspanins, Alix, lactadherin, butyrophilin, and xanthine

SUBMITTER: Cetinkaya H 

PROVIDER: S-EPMC11648521 | biostudies-literature | 2024

REPOSITORIES: biostudies-literature