Optimisation and comparison of orthogonal methods for separation and characterisation of extracellular vesicles to investigate how representative infant milk formula is of milk
| Autor: | Jessie Santoro, Anindya Mukhopadhya, Lorraine O'Driscoll, Zivile Useckaite, Barry Moran |
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| Rok vydání: | 2020 |
| Předmět: |
Imaging flow cytometry
food.ingredient Nanoparticle tracking analysis Hydrochloric acid Milk formula 01 natural sciences Extracellular vesicles Analytical Chemistry Acetic acid chemistry.chemical_compound Extracellular Vesicles 0404 agricultural biotechnology food Skimmed milk Animals Humans Chromatography Chemistry 010401 analytical chemistry Caseins Infant 04 agricultural and veterinary sciences General Medicine Flow Cytometry 040401 food science Casein micelles Infant Formula 0104 chemical sciences Milk Transmission electron microscopy Ultracentrifuge Ultracentrifugation Food Science |
| Zdroj: | Food chemistry. 353 |
| ISSN: | 1873-7072 |
| Popis: | Many infants are fed infant milk formula (IMF). However, IMF production from skim milk (SM) involves harsh treatment. So, we hypothesised that the quantity and/or quality of extracellular vesicles (EVs) in IMF may be reduced. Thus, firstly, we aimed to optimise separation of EVs from IMF and SM and, secondly, we aimed to compare the EV isolates from these two sources. Prior to EV isolation, abundant casein micelles of similar sizes to EVs were removed by treating milk samples with either acetic acid or hydrochloric acid. Samples progressed to differential ultracentrifugation (DUC) or gradient ultracentrifugation (GUC). EV characterisation included BCA, SDS-PAGE, nanoparticle tracking (NTA), electron microscopy (TEM), immunoblotting, and imaging flow cytometry (IFCM). Reduced EV concentrations were found in IMF. SM-derived EVs were intact, while IMF contained disrupted EV-like structures. EV biomarkers were more abundant with isolates from SM, indicating EV proteins in IMF are compromised. Altogether, a suitable method combining acid pre-treatment with GUC for EV separation from milk products was developed. EVs appear to be substantially compromised in IMF compared to SM. |
| Databáze: | OpenAIRE |
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