The beta-gal interferon assay: a new, precise and sensitive method.
Cells of a human glioblastoma line were stably transfected with a glial fibrillary acidic protein (GFAP) promoter sequence/lacZ reporter gene. Following this modification, they produced Escherichia coli beta-galactosidase constitutively in amounts that could be measured through their conversion of an added fluorophore into a product readily estimated by fluorimetry. Human interferons (IFN) selectively and in a dose-dependent manner reduce the formation of beta-galactosidase in this system. We have used it as the basis for a novel assay that is sensitive (4-40 pg/ml), precise, completed in 30 h, and applicable to both type I and type II human IFNs. Statistical analysis showed interassay relative standard deviations ranging from 5% to 11%, and most individual assays revealed potencies with limits of error within 85%-115%. Neither partially trypsin-digested IFN nor the other cytokines and mitogens we tested reacted in this system, except for tumor necrosis factor-alpha (TNF-alpha). The high selectivity was further shown by the loss of response to IFN in the presence of the appropriate specific anti-IFN or anti-IFN-gamma receptor antibodies.
Division of Biotechnology, Medical Products Agency, Uppsala, Sweden. ulf.hammerling@eurona.se
U
U
Hammerling
E
E
Bongcam-Rudloff
N
N
Setterblad
R
R
Kroon
A K
AK
Rehnström
E
E
Viitanen
G
G
Andersson
L
L
Sjödin
D001681:Biological Assay; D005470:Fluorometry; D017930:Genes, Reporter; D006801:Humans; D007372:Interferons / Q000494:pharmacology*; D010957:Plasmids / Q000235:genetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured; D001616:beta-Galactosidase / Q000096:biosynthesis*
D007372:Interferons; D001616:beta-Galactosidase