Neurology 4 plex e assay kit

Blood samples collected in EDTA tubes were centrifuged at 3,500 rpm for 15 min and plasma was removed. Then, the plasma samples were frozen at −80°C and were freeze-thawed only once. EDTA plasma Aβ40, Aβ42, GFAP, NFL, and P-tau181 were quantified using an ultra-sensitive Simoa technology (Quanterix, MA, United States) on the automated Simoa HD-X platform (GBIO, Hangzhou, China), according to the manufacturer’s instruction. The Neurology 4-Plex E Assay Kit (Cat No:103670) and Ptau181 Advantage V2 Assay Kit (Cat No:103714) were purchased from Quanterix and used accordingly. Plasma samples were diluted at a 1:4 ratio for measurement. Calibrators, internal quality controls, and all samples were measured in duplicate. The mean coefficients of variation (CVs) of duplicate measurement for concentration were 2.83% (Aβ40), 3.31% (Aβ42), 4.48% (GFAP), 3.22% (NFL), and 5.81% (P-tau181). Few samples with intra-assay CVs larger than 20% were re-measured. The values were discarded if the variance was still >20% after being re-measured. The assays were performed using kits with the same lot number. Operators were unaware of the participants’ disease status.

Fasting venous blood of 4 ml whole blood (EDTA anticoagulant tube) was collected from all PD patients and healthy controls, centrifuged at 4 °C within 2 h (2000 rpm, 10 min), the upper plasma was collected and stored in the refrigerator at −80 °C to avoid repeated freezing and thawing. Plasma GFAP, NfL, Aβ40, Aβ42, and p-tau181 were measured for all participants at baseline, 1-year, and 2-years follow-up using an ultrasensitive single-molecule array (Simoa^TM; Quanterix, Billerica, MA, USA) on the automated Simoa^TM HD-X platform (GBIO, Hangzhou, China). The Neurology 4-Plex E Assay Kit (Cat No:103670) and p-tau181 Advantage V2 Assay Kit (Cat No:103714) were used (Quanterix). The plasma sample was randomized, blinded, and measured using a batch of reagents from the same lot.