Phenylmethylsulfonyl fluoride pmsf protease inhibitor

Serum was collected after primary immunisation to determine the level of J8‐specific systemic antibodies on days 20, 40 and 60. Blood collection via the tail artery was allowed to clot for 30 min at 37°C. Serum was collected after centrifugation for 10 min at 1000 g, heat‐inactivated for 10 min at 56°C and stored at –20°C.
Mice were administered 50 μL of a 0.1% solution of pilocarpine to induce salivation. Saliva was then collected in tubes containing 2 μL of 50 mmol L⁻¹ phenylmethylsulfonyl fluoride (PMSF) protease inhibitor (Sigma‐Aldrich, St. Louis, MO, USA). Centrifugation for 10 min at 13 000 g was undertaken to remove particulate matter and samples stored at −80°C.

Serum was collected on days 20, 40, and 60 after primary immunization to determine the level of J8-specific systemic antibodies. Blood was collected from mice via the tail artery and allowed to clot for at least 30 min at 37 °C. Serum was collected after centrifugation for 10 min at 1000 g and stored at −20 °C.
Mice were administered 50 μL of a 0.1% solution of pilocarpine to induce salivation. Saliva was then collected in eppendorf tubes containing 2 μL of 50 mmol/L phenylmethylsulfonyl fluoride (PMSF) protease inhibitor (Sigma Aldrich). Particulate matter was separated by centrifugation for 10 min at 13,000 g and samples were stored at −80 °C.
Six to 10 freshly voided fecal pellets were collected from individual mice, frozen and then lyophilized. The dry weight of fecal solids was determined before they were resuspended by vortexing in 5% nonfat dry milk, 50 mmol/L EDTA (Sigma Aldrich), 0.1 mg/mL soyabean trypsin inhibitor (Sigma Aldrich), and 2 mmol/L PMSF (20 μL/mg of dry weight). Solid matter was separated by centrifugation for 10 min at 15,000 g. The supernatants were stored at −80 °C.