Spatially Resolved Multiplex Detection of Cytokines in Parkinson's Disease Rat Brain via CRISPR/Cas Biosensing

Abstract

The ability to accurately quantify localized neuroinflammatory and neurodegenerative biomarkers in vivo is critical for enabling the early diagnosis of Parkinson’s disease (PD). Here, we demonstrate a strategy for localized neuroinflammation quantification using an implantable glass-fiber-based CRISPR/Cas immunosensor. When interfaced with a portable fluorescence detector, our platform enables the spatially resolved multiplex detection of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and alpha-synuclein (α-syn) in the PD rat brain. The sensor surface was modified with functionalized liquid metal nanoparticles to enable antibody immobilization and long-term in vivo operation. This method demonstrated high specificity, excellent linearity in artificial cerebrospinal fluid (R2 > 0.91), sustained surface stability over 28 days, good biocompatibility with PC12 cells, and selective antibacterial activity against Staphylococcus aureus. Following cannula implantation into the substantia nigra pars compacta (SNc) of a 6-hydroxydopamine-induced PD rat model, the functionalized glass fiber enabled repeated, minimally invasive biomarker sampling at 6, 12, and 24 h postimplantation. Systemic inflammation induced by lipopolysaccharide (LPS) triggered marked elevations in TNF-α, IL-6, and α-syn, peaking at 6 h post-LPS. This implantable fiber-based biosensing method offers a promising reliable approach for early PD diagnosis, disease progression tracking, and investigation of neuroimmune interactions in neurodegenerative diseases in a point-of-care scenario.