Lb media

The BL21 (DE3) competent cell line was transformed with the human SPANXN4 gene in the pExp-GST plasmid. Overnight cultures from a single colony were prepared in LB media (Sigma) with ampicillin and incubated at 37°C. The pre-inoculum was inoculated into 4,000 mL LB media with ampicillin. The culture was grown at 37°C to OD₆₀₀ 0.6–0.8, induced with 0.5 mM IPTG, and incubated at 25°C for ∼20 h. Cells were harvested by centrifugation and resuspended in lysis buffer (phosphate buffer pH 7.4 with 300 mM NaCl, lysozyme 0.1 mg/mL, DNase I 0.01 mg/mL, 0.02 mM MgCl2, and protease inhibitor cocktail 1×). The cells were lysed by sonication (5 s ON and 10 s OFF) for 5 min, 40% amplitude. The lysate was centrifuged at 12,000 rpm for 30 min at 4°C, and the supernatant was purified using an Ni–NTA Sepharose column (GE). The supernatant was passed twice through the column after equilibration with wash buffer (PBS, pH 7.4, containing 10 mM imidazole). The column was washed with 50 column volumes of wash buffer, followed by elution with elution buffer (PBS, pH 7.4 with 500 mM imidazole). The fractions containing SPANXN4 protein were pooled, dialyzed against PBS, and subjected to gel filtration using the HiLoad 26/600 Superdex 200 pg FPLC column. The fractions containing the protein were pooled and quantified using a BCA assay.

1 L drinking water was spiked with RFP-expressing E. coli to 150 CFU/mL. The spiked water was filtered through a 0.22 µm filter (Corning Inc., NY), and the filter was then inoculated in 10 mL LB media (Sigma-Aldrich, MO) and incubated for 6 hr at 37°C. 20 µL of Dynabeads® MAX anti-E. coli O157 beads (Life Technologies, CA) were added to 1.5 mL of the incubation media and further incubated for 20 min on a rotating stage at room temperature (RT). Beads with captured bacteria were separated by magnet and resuspended in 400 µL of Phosphate Buffered Saline (PBS). The resuspended solution was co-encapsulated 500∶1 with green fluorescently labeled anti-E. coli antibody (FITC, ab30522, Abcam, MA) in droplet reactors inside the chip positioned on the ScanDrop robotic stage. After a further 1 hr of incubation at RT, images were taken from different locations on the chip. The objective position movements were controlled via PR-PR, and the generated images were automatically uploaded to Dropbox. Each experiment consisted of 4 repeats.

E. coli (Thermo Fisher) were grown overnight at 37 °C shaking culture in 1 mL ampicillin (Life Technologies)-supplemented LB media (Sigma) from cryogenically preserved aliquots. Once amplified, E. coli culture was washed thoroughly and assessed for colony-forming unit (CFU) count via duplicate measurement of suspension optical density (OD). With the exception of killing assays, all bacteria employed in co-culture experiments were irradiated in a trans-illuminator chamber (UVITEC), equipped with eight UV-C (250–280 nm) lamps.