Singh Anula Divyash, Patnam Sreekanth, Manocha Anisha, Bashyam Leena, Rengan Aravind Kumar, Sasidhar Manda Venkata
Apollo Hospitals Educational and Research Foundation (AHERF), Hyderabad, India.
Department of Biomedical Engineering, Indian Institute of Technology Hyderabad (IITH), Kandi, Hyderabad, India.
MethodsX. 2023 Aug 6;11:102310. doi: 10.1016/j.mex.2023.102310. eCollection 2023 Dec.
Urine is a highly advantageous biological specimen for biomarker research and is a non-invasive source. Most of the urinary biomarkers are non-specific, volatile and need extensive validation before clinical adoption. Extracellular vesicles are secreted by almost all cells and are involved in homoeostasis, intercellular communication, and cellular processes in healthy and pathophysiological states. Urinary extracellular vesicles (UEVs) are released from the urogenital system and mirror the molecular processes of physiological and pathological states of their source cells. Therefore, UEVs serve as a valuable source of biomarkers for the non-invasive diagnosis of various pathologies. They hold a promising source of multiplex biomarkers suitable for prognosis, diagnosis, and therapy monitoring. UEVs are easily accessible, non-invasive, and suited for longitudinal sampling. Although various techniques are available for isolating UEVs, there is yet to be a consensus on a standard and ideal protocol. We have optimized an efficient, reliable, and easily adoptable polyethylene glycol (PEG) based UEV isolation technique following MISEV guidelines. The method is suitable for various downstream applications of UEVs. This could be a cost-effective, consistent, and accessible procedure for many clinical labs and is most suited for longitudinal analysis. Adopting the protocol will pave the way for establishing UEVs as the ideal biomarker source. •Urine can be collected non-invasively and repeatedly, hence a very useful specimen for biomarker discovery. Urinary EVs (UEVs), derived from urine, offer a stable diagnostic tool, but standardised isolation and analysis approaches are warranted.•To have enough UEVs for any study, large volumes of urine sample are necessary, which limits different isolation methods by cost, yield, and time.•The protocol developed could help researchers by offering a cost-effective and dependable UEV isolation method and may lay the foundation for UEVs adoption in clinical space.
尿液是生物标志物研究中极具优势的生物样本，且是一种非侵入性来源。大多数尿液生物标志物是非特异性的、易挥发的，在临床应用前需要进行广泛验证。细胞外囊泡由几乎所有细胞分泌，参与健康和病理生理状态下的体内平衡、细胞间通讯及细胞过程。尿细胞外囊泡（UEVs）从泌尿生殖系统释放，反映其来源细胞生理和病理状态的分子过程。因此，UEVs是用于各种疾病非侵入性诊断的有价值生物标志物来源。它们是适用于预后、诊断和治疗监测的多重生物标志物的有前景来源。UEVs易于获取、非侵入性且适合纵向采样。尽管有多种技术可用于分离UEVs，但尚未就标准和理想方案达成共识。我们遵循MISEV指南优化了一种基于聚乙二醇（PEG）的高效、可靠且易于采用的UEV分离技术。该方法适用于UEVs的各种下游应用。这可能是许多临床实验室具有成本效益、一致且可及的程序，最适合纵向分析。采用该方案将为将UEVs确立为理想生物标志物来源铺平道路。•尿液可以非侵入性地反复收集，因此是生物标志物发现的非常有用的样本。源自尿液的尿细胞外囊泡（UEVs）提供了一种稳定的诊断工具，但需要标准化的分离和分析方法。•为了在任何研究中获得足够的UEVs，需要大量尿液样本，这在成本、产量和时间方面限制了不同的分离方法。•所开发的方案可为研究人员提供一种经济高效且可靠的UEV分离方法，可能为UEVs在临床领域的应用奠定基础。
