Human beta ngf duoset elisa kit

Solid phase sandwich ELISA was performed to detect neurotrophic factors in nerve conduits. The snap-frozen 2-week conduits were thawed on ice. The tissue inside conduit was then collected, soaked in 500 μl tissue lysis buffer (786–180, Mammalian Cell PE LB, G-Biosciences) supplemented with protease inhibitors (1 mM PMSF, 1 mM Na₃VO₄, and 10 μg/ml Leupeptin), and then homogenized with GentleMACS dissociator. The tissue lysate was centrifuged under 14000 × g for 15 min at 4 °C to remove debris, and the supernatant was collected for ELISA test. Human BDNF Quantikine ELISA kit (DBD00, R&D Systems), human CNTF Quantikine ELISA kit (DNT00, R&D Systems), and human beta-NGF DuoSet ELISA kit (DY256–05, R&D Systems) were used for the tests following manufacturer’s instruction. The amount of growth factor was normalized by total protein concentration of tissue lysate for comparison. Total protein concentration of tissue lysate was determined by DC™ Protein Assay (BioRad) prior to ELISA.