Elisa

Participants underwent a venipuncture after informed consent. In case of a history of ICH, venipuncture was done at least 3 months after the symptomatic hemorrhage. EDTA plasma was collected in polypropylene tubes, centrifuged, aliquoted, and stored in polypropylene tubes at − 80 °C for all patients, at all locations. For all plasma analyses, the technician who performed the analysis was blinded for the clinical diagnosis, and patient and control samples were randomly analyzed to avoid bias.
Aβ38, Aβ40, and Aβ42 levels were quantified in the plasma using ELISAs (Euroimmun, Lübeck, Germany). All measurements were performed in duplicate. The coefficient of variation (CV) was < 20% for all duplicate measures, except for 4 Aβ38 measurements with a CV of 21–31%.
For all ELISAs, five quality control samples were included on each plate to correct for any inconsistencies between plates. These controls consisted of pooled EDTA plasma samples that were stored in aliquots at − 80 °C. For each analysis, a fresh aliquot was used.

Serum aCL and aβ2GPI antibodies were measured by enzyme-linked immunosorbent assays (ELISAs) (EUROIMMUN, Lübeck, Germany). In brief, the peripheral blood sample was examined in three wells, two wells with antigen and one without, to subtract the background from the specific binding. According to the instructions of the ELISA kit, cutoff levels were calculated at the 99th percentile. IgG, IgM, and IgA subtypes of aCL and aβ2GPI >20 CU confirmed at the 12-week interval were considered positive.

IL-1β, IL-6, IL-10, IL-17A, TNF-α, TRAIL, and NF-L were measured using Simoa HD-1 digital enzyme-linked immunosorbent assay (ELISA) (Quanterix, Lexington, MA, USA).
CXCL13 and INF-α/β concentrations were determined using ELISAs (Euroimmun, Lübeck, Germany and PBL Assay Science, NJ, USA, respectively) in the Department of Clinical Immunology, Odense University Hospital, also accredited according to the ISO 15189 standard. Lower cutoff sensitivity of the CXCL13 assay was set to 10 pg/ml, according to the manufacturer’s instructions [16 (link)].

Donors included in the study (n = 95) were divided into 5 groups according to their clinical characteristics. Candidate donors underwent a blood donors’ history questionnaire and symptoms related to SARS-Co-V infection were also recorded (i.e., fever, fatigue, headache, cough, dyspnea, diarrhea, loss of smell, loss of taste, duration of symptoms). For the detection of anti-SARS-CoV-2 Abs, commercially available ELISAs (Euroimmun Medizinische Labordiagnostika AG, Lubeck, Germany), which detect plasma IgG and IgA antibodies against the recombinant spike protein (S1 domain) of SARS-CoV-2 were used according to the manufacturer’s instructions. Ratios < 0.8 were considered negative, ≥0.8 to <1.1 were considered borderline, and ≥1.1 were considered positive.