Biological assays for interferons

Abstract

Interferons (IFN) are potent biologically active proteins synthesised and secreted by somatic cells of all mammalian species. They have been well characterised, especially those of human origin, with respect to structure, biological activities, and clinical therapeutic effects. While structural differences are known to exist among the IFN species that constitute the "IFN family" and despite the existence of different receptors for type I and type II IFN, all species have been shown to exert a similar spectrum of in vitro biological activities in responsive cells. Principal among the biological activities induced by IFN is antiviral activity, the activity used to originally define IFN. Antiviral activity of IFN is mediated via cell receptors and is dependent on the activation of signalling pathways, the expression of specific gene products, and the development of antiviral mechanisms. Sensitivity of cells to IFN-mediated antiviral activity is variable, and depends on a number of factors including cell type, expression of IFN receptors and downstream effector response elements, effectiveness of antiviral mechanisms, and the type of virus used to infect cells. Nevertheless, by the judicious use of sensitive cell lines in combination with appropriate cytopathic viruses, effective assays to measure the antiviral activity have been developed. Historically, "antiviral assays" (AVA) were the first type of biological assays that were developed to measure the relative activity or potency of IFN preparations. However, the subsequent discoveries of several other biological activities of IFN has opened the way to the development of assays based on one or other of these activities. The latter include inhibition of cell proliferation, regulation of functional cellular activities, regulation of cellular differentiation and immunomodulation. More recently, the cloning of IFN responsive genes has led to the development of "reporter gene assays". In this case, the promoter region of IFN responsive genes is linked with a heterologous reporter gene, for example, firefly luciferase or alkaline phosphatase, and transfected into an IFN-sensitive cell line. Stably transfected cell lines exposed to IFN increase expression of the reporter gene product in direct relation to the dose of IFN, the readout being a measure of this product's enzymic action. The current review aims to give a critical overview of the development, specificity, standardisation and present use of the various biological assay methods now available for the quantification of IFN activity.