Neurite‐Promoting Factor in Conditioned Medium from RN22 Schwannoma Cultures: Bioassay, Fractionation, and Properties

Abstract

On polyornithine (PORN) substrata dissociated 8 day chick embryo ciliary ganglionic neurons will survive if the culture medium is supplemented with ciliary neuronotropic factor [CNF]. Neuritic growth will not occur unless the substratum is derivatized with a PORN-bindable neurite promoting factor (PNPF). Soluble PNPF can be assayed by a convenient in vitro system and obtained in relatively large amounts from serum-free media conditioned over RN22 [rat] Schwannoma cultures. PNPF can be concentrated by using Amicon XM100 ultrafiltration and separated from nearly all of the non-active protein by using ion-exchange chromatography. The partially purified PNPF can be concentrated using XM100 and is heat- and protease-sensitive. In some cases a concentration-dependent interference with the expression of PNPF activity in the bioassay was noted. Graphical methods are proposed to permit the simultaneous determination of PNPF and the extent of such interference. Different treatments that affected the interference property did not always affect PNPF activity in a reciprocal manner, leaving open the possibility that the interference with PNPF activity results from reversible alteration of the PNPF molecule, or that there exists a separate interfering agent.