Fig. 2. Analysis of neurite growth-inhibitory activity after separation of myelin proteins by anion exchange chromatography.A, The elution profile of the Q-Sepharose column is shown. A, Chaps-extract of spinal cord myelin was loaded onto a Q-Sepharose column and eluted by a step salt gradient as indicated (0.1–1 m NaCl). Activity of each fraction was tested in the bioassay. Three activity peaks (indicated byarrows) eluting at 150 (peak I), 400 (peak II), and 800 (peak III) mm NaCl could be determined. B, Dot-blot profile of active fractions to monitor distribution of myelin-associated neurite growth inhibitors. Active fractions of each peak were pooled, and aliquots of 5 μg each were dotted on membranes and probed with anti-MAG, IN-1 antibodies, and Alcian blue, respectively. Note that the presence of MAG correlates with peak I, the IN-1 antigen with peak II, and the enrichment of proteoglycans with peak III. C, Aliquots (2 μg/lane) of each activity pool were electrophoresed on 6% SDS-polyacrylamide gels and visualized by silver staining. Lane 1, Activity peak I; lane 2, activity peak II; lane 3, activity peak III, proteins appear as a mixture of a high-molecular weight smear;lane 4, activity peak III digested with chondroitinase ABC. Enzyme treatment causes the disappearance of the smear and increases the amount of components at 400, 320, 180, and 140 kDa. The mobility of the molecular weight marker is indicated by thearrows to the left. Laminin was used as a marker for 400 kDa.