Cf5 absorbent pad

FF120HP nitrocellulose membrane (Cytiva; #13549205) was cut to 2.5 cm using a craft paper cutter and assembled onto a backing adhesive (DCN Diagnostics; #MIBA-020) with a CF5 absorbent pad (Cytiva; #8115–2250) and a Standard 14 sample pad (Cytiva; # 8133–2250). The assembled, uncut membrane was stripped using a BioDot dispenser (BioDot; #XYZ30600124), with a flow rate of 1 μL/cm and a dispense volume of 30 μL per 30 cm card. Striped membranes were dried in a Robbins Scientific Micro Hybridization Incubator 2000 at 37 °C for 30 min, then stored overnight at 20 °C in a desiccator chamber (Totech; SuperDry Desiccant Cabinet; #SD-151–21) at 5% humidity to dry completely. The striped card was then cut into 3 mm strips using a KinBio ZQ2000 Guillotine Cutter and stored at 20 °C in sealed 50 mL conical tubes (USA Scientific; #5622–7261) with desiccant packs (Interteck Packaging; #IN1G51). The test lines for each set of strips contained mouse anti-SARS-CoV-2 antibodies (Bioss; #bsm-41411M) at 1 mg/mL in 1× PBS, and the control lines contained goat polyclonal anti-mouse antibodies (Arista; #ABGAM-0500) at 1 mg/mL in 1× PBS.

The architecture of a typical LFA strip and the process of making LFA strips in our lab are shown in Supplementary Fig. 1. FF80HP nitrocellulose membrane (Cytiva; #13549206) was cut to 2.5 cm using a craft paper cutter (Fiskars; #199080–1007) and assembled on a 30-cm backing adhesive card (DCN Diagnostics; #MIBA-020) along with a CF5 absorbent pad (Cytiva; #8115–2250). The assembled membrane was striped using a BioDot dispenser (BioDot; #XYZ30600124), at a flow rate of 1 μL/cm and a dispensed volume of 30 μL per 30 cm card. The test line contained goat polyclonal anti-αhCG antibodies (Arista; #ABACG-0500) diluted to a concentration of 1 mg/mL in 1 × PBS. The control line contained goat polyclonal anti-mouse antibodies (Arista; #ABGAM-0500) diluted to a concentration of 1 mg/mL in 1 × PBS. Striped membranes were dried in a Robbins Scientific Micro Hybridization Incubator 2000 at 37 °C for 30 min, then stored overnight at 20 °C in a desiccator chamber (Totech; SuperDry Desiccant Cabinet; #SD-151–21) at 5% humidity. The striped card was cut into 3 mm strips using a KinBio ZQ2000 Guillotine Cutter and stored at 20 °C in sealed 50 mL conical tubes (USA Scientific; #5622–7261) with desiccant packs (Interteck Packaging; #IN1G51).

LFA strips (Fig 1) consisting of Standard 14 sample pad, FF80HP nitrocellulose membrane and CF-5 absorbent pad (all from Cytiva) were assembled on an adhesive backing card (MIBA-020; DCN Diagnostics). Test line (TL) and control line (CL) were striped on the nitrocellulose membrane with 1 mg/mL polyclonal goat anti-biotin antibodies (ab 6643; Abcam) and 1 mg/mL polyclonal goat anti-mouse antibodies (ABGAM-0500; Arista Biologicals, Inc.), respectively, using a BioDot dispenser (XYZ30600124) at a rate of 1 μL/cm. The striped membrane was dried at 37°C for 30 min in an incubator (Robbins Scientific Micro Hybridization Incubator 2000) and then cut into 3 mm wide strips using a ZQ2000 Guillotine Cutter (Kinbio). LFA strips were stored dried (in plastic tubes with desiccant bags) until use.