Catalog no e30 103

Total IgA levels were determined in gut rinse samples as previously described (Merino-Guzmán et al., 2017 (link)). Briefly, an intestinal section of 5 cm from Meckel’s diverticulum was taken and rinsed three times with 5 mL 0.9% saline; then the rinse was collected in a tube and centrifuged at 1864 × g at 4°C for 10 min. The supernatant was poured into a 96-microwell plate and stored at -20°C until tested. A commercial indirect ELISA set was used to quantify IgA according to the manufacturer’s instructions (Catalog No. E30-103, Bethyl Laboratories, Inc., Montgomery, TX, United States). 96-well plates (Catalog No. 439454, Nunc MaxiSorp, Thermo Fisher Scientific, Rochester, NY) were used, and samples were measured at 450 nm using an ELISA plate reader (Synergy HT, multi-mode microplate reader, BioTek Instruments, Inc., Winooski, VT, United States). The obtained chicken IgA concentration was multiplied by the dilution factor (1:100) to determine the amount of chicken IgA in the undiluted samples.

Total IgA levels were determined in 12 gut rinse samples per group as previously described (29 (link)). An intestinal section of 5 cm from Meckel's diverticulum to the ileocecal junction was taken and rinsed three times with 5 mL of 0.9% saline; then the rinse was collected in a tube and centrifuged at 1,864 × g at 4°C for 10 min. The supernatant was poured into a 96-microwell plate and stored at −20°C until tested. A commercial indirect ELISA set was used to quantify IgA according to the manufacturer's instructions (Catalog No. E30-103, Bethyl Laboratories Inc., Montgomery, TX 77356) in samples diluted 1:100. High protein-binding capacity 96-well plates (Catalog No. 439454, Nunc MaxiSorp, Thermo Fisher Scientific, Rochester, NY) were used, and samples were measured at 450 nm using an ELISA plate reader (Synergy HT, multi-mode microplate reader, BioTek Instruments, Inc., Winooski, VT, USA). Total intestinal IgA levels obtained were multiplied by the dilution factor (100) to determine the amount of chicken IgA in the undiluted samples.

Total IgA levels in experiments 2 and 3 were determined in 12 gut rinse samples each as previously described [33 (link)]. An intestinal section of 5 cm from the Meckel’s diverticulum to the ileocecal junction was taken and rinsed three times with 5 mL of 0.9% saline; then the rinse was collected in a tube and centrifuged at 1864× g at 4 °C for 10 min. The supernatant was poured into a 96-microwell plate and stored at −20 °C until tested. A commercial indirect ELISA set was used to quantify IgA according to the manufacturer’s instructions (Catalog No. E30-103, Bethyl Laboratories Inc., Montgomery, TX 77356, USA). 96-well plates (Catalog No. 439454, Nunc MaxiSorp, Thermo Fisher Scientific, Rochester, NY, USA) were used, and samples diluted 1:100 were measured at 450 nm using an ELISA plate reader (Synergy HT, multi-mode microplate reader, BioTek Instruments, Inc., Winooski, VT, USA). Total intestinal IgA levels obtained were multiplied by the dilution factor (100) to determine the amount of chicken IgA in the undiluted samples.