Bioassay detection of mouse nerve growth factor (mNGF) in the brain of adult mice

Two pools of seven brains each from adult Swiss-Webster mice were homogenized, and supernatants were collected for bioassay. PC-12 cells were placed in a bioassay plate at time zero, at a concentration of 10(4) cells per well, and primed for 48 hours in a medium containing 50 ng/ml of mNGF. The PC-12 cell bioassay for neurite outgrowth was conducted after primed cells were exposed to an NGF-free medium for 24 hours. Suitable controls for serum toxicity and cell viability were established. The sensitivity of the bioassay approximates 100 pg NGF/ml. The results showed 80-100% neurite outgrowth in wells exposed to brain pool supernatant (BPS) alone, and control level outgrowth (3-8%) in wells containing BPS and specific anti-beta-NGF antibody. Therefore, the brains of Swiss-Webster adult mice contain an NGF-like substance which promotes neurite outgrowth in PC-12 cells. The substance probably is NGF itself, since the effect is blocked by specific NGF antiserum.