Microarrays containing synthetic RV-VP1 N terminal peptides,
recombinant RSV-G protein and allergens
Comprehensive lists of the chip components spotted on microarrays are
reported in Tables S1-2. Synthetic VP1 N terminal peptides representing
the three genetic RV species (RV-A: n=18; RV-B: n=9; RV-C: n=10) were
selected and produced as described in a previous study (8). The
recombinant G protein of the RSV A2 strain was expressed inEscherichia coli BL21(DE3) cells, purified and characterized as
described (19). Food allergens used for the calibration of the
microarrays were purchased from Sigma Aldrich (Saint Louis, MO, USA)
(Table S2). Glass slides containing six microarrays surrounded by an
Epoxy frame were obtained from Paul Marienfeld GmbH & Co. KG
(Lauda-Königshofen, Germany) and coated with an amine-reactive complex
organic polymer, MCP-2 (Lucidant Polymers, Sunnyvale, CA, USA).
Microarray components were used at a concentration of 1 or 2 mg/ml in a
phosphate buffer (75 mM Na2HPO4), pH =
8.4 and spotted in triplicates using a SciFlexArrayer S12 (Scienion AG,
Berlin, Germany). After spotting, printed slides were placed in a humid
chamber at 75% relative humidity and incubated at room temperature (RT)
overnight. Slides were then blocked with 50 mM ethanolamine in
PBS/Tween, pH=9, for 15min and subsequently covered with a plate sealer
(Candor, Wangen, Germany). Microarrays were vacuumed and stored at 4°C
until use.