Cd45 phycoerythrin clone 30 f11

Yellow-green fluorescent carboxylate-modified microspheres (Invitrogen/Thermo Fisher, Waltham, MA) were opsonized by incubating with Roswell Park Memorial Institute-1640 cell culture medium containing 10% fetal bovine serum (FBS) at 37 °C for 30 mins. The opsonized microspheres were then injected intraperitoneally at 23 hours after sham or CLP surgery in a dose of 10⁸ microspheres/200 μl volume per mouse. After one hour, 5 ml of sterile normal saline was injected and mixed thoroughly by gentle massage. Four milliliters of peritoneal lavage fluid were collected and centrifuged. A faction of cells (4 × 10⁵) were stained with the following surface markers: CD45-Phycoerythrin (clone 30-F11, 1:1500 dilution, BD Biosciences, San Jose, CA, ), Ly6G-Brillian Violet 421 (clone 1A8, 1:100 dilution, BD Biosciences, San Jose, CA), F4/80-Alexa Fluro 647 (clone T45-2342, 1:100 dilution, BD Biosciences, San Jose, CA) at 4 °C for 30 min in the dark as described previously ^{12 (link)}. Cells were washed twice in 4 ml of ice cold phosphate-buffered saline (PBS) buffer and resuspended in PBS containing 5% FBS, and then flow cytometry analysis was performed using a BD LSR II flow cytometer. The phagocytic function in single cell was expressed as the mean fluorescence intensity of microsphere within the chosen cell population.