For this test, 18 MTPs of the type Microfluor 2 (96-wells, polystyrene, round well, flat bottom, black, Thermo Fisher Scientific, Waltham, MA, USA) were prepared using a sciFLEXARRAYER S12 microarray printer installed with a piezo dispense capillary (PDC) size 60 with coating type 3 (Scienion AG, Berlin, Germany. The printing procedure was performed under strict humidity (55–65%) and temperature control (18–20 °C).
The design of the array (shown inFigure 1 ) consisted of three anti-human antibodies against CD63 (Bio-rad Laboratories Inc., Hercules, CA, USA), CD9, and CD81 (Ancell Corporation, Stillwater, MN, USA) at a concentration of 100 µg/mL, biotinylated goat anti-mouse IgG (Novus Biologicals, Centennial, CO, USA) in two concentrations (10 µg/mL and 50 µg/mL) as positive controls, and buffer as the negative control. The antibodies and controls were prepared in three different spotting buffers: 5% glycerol in PBS (denoted “gly”), 50 mM trehalose in PBS (denoted “tre”), and 1 × sciSPOT Protein D1 buffer (2 × concentrate, denoted “D1”) in PBS (Scienion AG, Berlin, Germany), which were all included in the same print design. When the printing was finalized, the MTPs were incubated in the controlled climate of the printer for 10 h to allow adsorption of the antibodies.
The design of the array (shown in