Intercellular adhesion : II. The purification and properties of a horse serum protein that promotes neural retina...

When 8-day-old embryonic chicken neural retina is dissociated, the resulting single cells adhere to each other at a rapid rate at 37 °C, and slowly at 5 °C. However, the addition of horse serum substantially accelerates the rate of aggregation at 5 °C, although it shows no effect at 37 °C; the rate at 37 °C exceeded the maximum rate at 5 °C (i.e., in the presence of excess horse serum). The kinetics of the horse serum effect were investigated, and, based on these results, an assay was devised for the active component in the serum. The active protein, termed "neural retina aggregating protein" (NRP), was purified 48-fold from the serum, and at this stage appears close to homogeneity. The rate of aggregation of neural retina cells is significantly stimulated by adding 1 μg of purified NRP per ml. Purified NRP shows certain properties in common with horse macroglobulins (horse Immunoglobulin M, IgM), but there are also some important differences. For example, it cross-reacts with antibodies to horse IgM, it can be reduced to subunits with thiols, and it is of high molecular weight (about 1.6×10(6)). However, NRP has a higher sedimentation value than IgM (22.4 S compared to 19 S), and purified IgM does not increase the rate of aggregation of neural retina cells at 100-fold the concentration of NRP required for optimum activity. Inhibitory activity was not detected at any stage of the purification of NRP from horse serum. However, other sera (chick and calf) were shown to reduce the normal slow rate of aggregation at 5 °C and to inhibit the effect of NRP. Some preliminary studies indicate NRP may be specific for neural retina cells. For example, the rate of aggregation of 5-day-old embryonic limb bud cells is unaffected by concentrations of NRP that are optimal for neural retina cells.