Zehra Seda ÜNAL HALBUTOĞULLARI, Zeynep Ece UTKAN KORUN, Cansu SUBAŞI DEMİR, Kamil Can KILIÇ, Büşra ALPER, Yusufhan YAZIR

Optimization of Protein Quantification in Wharton Jelly-Derived Mesenchymal Stem Cell Exosomes

Aim: Exosomes are small intracellular membrane-based vesicles with different compositions that are involved in various biological and pathological processes. They are secreted by all cell types and can be found in most body fluids, including the blood, saliva, and urine. Exosomes are nanometer-sized microvesicles, approximately 30-200 nm in diameter, that contain DNA, mRNAs, non-coding RNAs, membrane proteins, and cytosolic proteins. Mesenchymal stem cells exert paracrine effects through exosomes. Recent studies have shown that exosomes have important potential as a new alternative to cellular therapies. In this study, we aimed to obtain exosomes from Wharton-Jelly derived mesenchymal stem cells (WJ-MSCs), characterize methods, quantify protein from these exosomes, and optimize this by comparing it with the amount of nanoparticles and determining the most effective lysis solution using different lysis chemicals. Methods: For this purpose, first, WJ-MSCs' exosomes were isolated and characterized, and changes in protein levels were determined after treatment with 1 billion/ml particle exosomes with commonly used chemicals such as RIPA buffer, Mammalian Protein Extraction Reagent (M-PER), Tris-TritonX, and Tris-SDS. Results: As a result, approximately 2.5 µg/ml of 1 billion/ml particulate exosomes were detected with the BCA kit. Concentration of protein in these exosomes, and the protein concentration increased 3-4 times as a result of disintegration of the bilayer membranes in the cell membrane structure of these exosomes with RIPA buffer. Conclusion: It is predicted that these data can be used comparatively in future studies, especially for the quantification of mesenchymal stem cell exosomes.

Anahtar Kelimeler:

Wharton Jelly, Exosome, Mesenchymal Stem Cell, Protein Quantification, Lysis Buffer

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