Human magnetic cytokine panel

Participant blood samples were collected within 24 hours of emergency department presentation in EDTA-containing vacutainers (BD Biosciences, San Jose, CA), transported on ice, spun down at 2500g for 10 min at 4°C, and stored at −80°C until further analysis. Cell-free plasma was analyzed using a human magnetic cytokine panel providing simultaneous measurement of 35 cytokines (Thermo Fisher Scientific, Waltham, MA). The assay was performed according to the manufacturer’s instructions with each subject sample performed in duplicate and then analyzed on a Luminex FLEXMAP 3D instrument.

Blood was collected into EDTA-anticoagulated tubes and plasma was frozen at −80°C until analysis. BAL samples were kept on ice and prepared within 4 hours of collection. BAL samples were centrifuged at 300 × g for 15 minutes at 4°C and the collected supernatant was frozen at −80°C.
Plasma and BAL samples were analyzed using a human magnetic cytokine panel providing parallel measurement of 35 cytokines and chemokines (Thermo Fisher Scientific). The assay was performed according to the manufacturer’s instructions with the following modification: the samples were fixed with 100 μL of 1% paraformaldehyde at room temperature for 60 minutes on a shaker at 700 RPM and washed once before the final resuspension. The samples were analyzed on a Luminex FLEXMAP 3D instrument.