Isolation of Exosome Nanoparticles from Human Cerebrospinal Fluid for Proteomic Analysis

Abstract

Exosomes are nanoscale (30−150 nm) biological vesicles that are actively released from living cells and circulating into all body fluids. Recently, exosomes in cerebrospinal fluid (CSF) have been recognized as promising biomarkers for central nervous system (CNS) neoplasms. In this study, we report a labelfree method that can be used to rapidly isolate exosomes from CSF for proteomic analysis. Compared to ultracentrifugation and polyethylene glycol-based precipitation, our method isolates exosomes from 2 mL of CSF within 10 min, which is 18 times and 72 times shorter, respectively; the yield was increased by 4.47 times and 2.09 times and the purity was increased by 4.54 times and 9.76 times, respectively. The proteomic analysis further revealed that the exosomes isolated by our method identified more exosomerelated proteins, which may reflect the physiological status of diseases for exosome-based diagnosis. Therefore, the effective isolation of pure exosomes from CSF samples for protein analysis will benefit the downstream analysis and clinical translation of exosomes, thus promoting the early diagnosis of CNS neoplasms.

Exosome Processing and Characterization Approaches for Research and Technology Development

“…Similarly, CSF comes into contact with the spinal cord and meninges and can reflect pathologies of these CNS elements. CSF is a primary sample type used for CNS disease diagnosis, [ 192 ] but CSF‐specific exosome isolation methods are needed to achieve sufficient purity. For example, confounding nonexosomal proteins must be removed during exosome isolation.…”

“…[ 188 ] Other such proteins include abundant CSF proteins such as albumin and immunoglobulin, which can obscure scarcer proteins that are important in exosome‐based diagnostics. [ 192 ] To address these issues, a novel microfluidic device was developed [ 192 ] that uses negative pressure oscillation to streamline exosome isolation, and enabled the identification of more exosome‐related proteins than traditional ultracentrifugation and PEG‐based precipitation methods. Novel methods such as this microfluidics approach that improve the isolation of CSF‐derived exosomes are critical for clinical sample analysis and neurodegenerative diagnostic research.…”

Exosomes in Alzheimer’s Disease: From Being Pathological Players to Potential Diagnostics and Therapeutics

“…EXOs in the CSF have recently been identified as prospective biomarkers for central nervous system (CNS) disease [ 126 ]. In addition to blood EXOs, a strong correlation was found between CSF exosomal biomarkers and AD diagnosis since Aβ 42 , T-tau, and P-T 181 -tau differed significantly in AD patients and those with MCI and controls [ 127 ].…”

Biological Significance of the Protein Changes Occurring in the Cerebrospinal Fluid of Alzheimer’s Disease Patients: Getting Clues from Proteomic Studies

“…The finding that exosomes isolated from human CSF or brain samples sequestered oligomeric Aβ in the brain has led to propose their protective role in AD pathogenesis [ 47 ]. Indeed, several proteins of our database such as CHL1, KNG1, or APOA1 were found to be constituents of human CSF exosomes [ 48 ].…”