Mouse p selectin elisa kit

Selp^−/− mice (male, 10–19 weeks of age) were retro-orbitally injected with platelets from wild-type C57BL/6J mice at 1 × 10⁸ per mouse per day. This injection was administered for 3 consecutive days in conjunction with the last three doses of G-CSF (250 μg/kg/day). The platelet purification and transfer procedures followed the prior study’s methods [53 (link)]. After Ficoll–Paque PLUS purification, the mouse plasma underwent two rounds of centrifugation at room temperature, each at 9170 × g for 10 min, to eliminate any residual platelets. Subsequently, the supernatant underwent ultra-centrifugation at 157,000 × g for 1.5 h at 4 °C to segregate microparticles (in the pellet) from the microparticle-free supernatant [54 (link)]. The concentrations of P-selectin in microparticle and soluble fraction were determined using the mouse P-selectin ELISA kit (RayBiotech, Inc. USA).

C57BL/6 J mice (male, 10–12 weeks old) were treated with 55 μg/kg G-CSF administered by retro-orbital injection once daily on 2 consecutive days for the EPO immunoassay or 5 consecutive days for the soluble P-selectin immunoassay. Animals treated with an identical volume of saline served as negative controls. C57BL/6 J mice that were retro-orbitally injected with 1.5 mg/kg LT and untreated served as positive controls for the EPO immunoassay and the soluble P-selectin immunoassay, respectively. PB (serum for the EPO immunoassay and plasma for the soluble P-selectin immunoassay) was collected retro-orbitally at 22, 44, and 66 h after initial injection for the EPO immunoassay, and day 0 before initial injection and days 1–5 after initial injection for the soluble P-selectin immunoassay. The EPO and soluble P-selectin immunoassays were performed using an enzyme-linked immunosorbent assay (ELISA) in accordance with the manufacturers’ instructions (Quantikine® Mouse/Rat EPO immunoassay, R&D systems; Mouse P-selectin ELISA kit, RayBiotech, Inc., USA).

Human and mouse plasma were twofold and 40-fold diluted with PBS, respectively. The levels of human and mouse sP-sel were measured using a custom-made ProcartaPlex Immunoassay (Thermo Fisher Scientific, Waltham, MA, USA) and enzyme-linked immunosorbent assay (ELISA) in accordance with the manufacturer’s instructions (mouse P-selectin ELISA kit, RayBiotech, Inc., USA), respectively.