Platelia aspergillus

Peripheral blood samples were collected using heparin anticoagulant blood collection tubes (Becton, Dickinson and Company, USA) then sera were isolated by centrifugation at 4000 rpm for 10 min. Serum GM antigen levels were measured using a commercial sandwich enzyme-linked immunosorbent assay (ELISA) kit (Platelia™ Aspergillus; Bio-Rad, CA). An optical density (OD) index value of ≥0.5 was interpreted as a positive result.

SARS‐CoV‐2 PCR was performed by in‐house PCR or by Cepheids GeneXpert Xpress SARS‐CoV‐2 PCR as described by Wolters et al.²⁴ Triazole susceptibility screening was done using VIPcheck™ (Mediaproducts BV). MICs of Aspergillus fumigatus isolates were determined with broth microdilution using CLSI standards.²⁵ Fungal PCR targeting the Cyp51A gene was done using AsperGenius™ (PathoNostics). 1‐3 β‐d‐glucan (BDG) testing was done using the Fungitell assay (Associates of Cape Cod Inc). Galactomannan (GM) testing was done using Platelia Aspergillus (Bio‐Rad) and/or Aspergillus lateral flow device (AspLFD, OLM Diagnostics).

The GM-ELISA (Platelia Aspergillus; Bio-Rad, Edmonds, WA) was performed on positive BAL samples, according to the manufacturer's recommendations.¹⁸ (link) Briefly, 100 μL of the Platelia treatment solution was added to 300 μL of the BAL specimen and heated at 100 °C for 3 min before undergoing centrifugation. The supernatant and the horseradish peroxidase-labeled monoclonal antibody (EBA-2) were incubated at 37 °C in antibody-precoated microplates. The plates were washed and then incubated with a substrate chromogen reaction solution. The reaction was stopped by the use of 100 μL of sulfuric acid and after 30 min, the optical density (OD value) of each well was read using a microplate reader at 450 nm. One well for the negative control, two wells for the cut-off control, and one well for the positive control were used. The results of Galactomannan ELISA assay are expressed as “galactomannan index” (GMI = OD sample/mean OD cut-off control), by comparison to the “cutoff” control. A GMI index of ≥1.0 in two consecutive samples was considered positive.