A sensitive new bioassay for tumor necrosis factor

• Published in: Journal of immunological methods Vol. 175; no. 2; pp. 181 - 187
• Main Authors: Shahan, T.A., Siegel, P.D., Sorenson, W.G., Kuschner, W.G., Lewis, D.M.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 14.10.1994; Elsevier
• Subjects: Analysis of the immune response. Humoral and cellular immunity; Animals; Bioassay; Biological and medical sciences; Biological Assay - methods; Bronchoalveolar Lavage Fluid - cytology; Cell Line; Clone Cells; Colorimetric metabolic indicator; Coloring Agents; Cytotoxicity Tests, Immunologic; Fluorometric metabolic indicator; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Gentian Violet; Immunobiology; Lipopolysaccharides; Lymphokines, interleukins ( function, expression); Macrophages, Alveolar - immunology; Mice; Neutral Red; Oxazines; Oxidation-Reduction; Rats; Rats, Inbred Strains; Regulatory factors and their cellular receptors; Sensitivity and Specificity; Tumor necrosis factor; Tumor Necrosis Factor-alpha - analysis; Xanthenes; RFU; Bioassay; Colorimetric metabolic indicator; IMDM-15; LPS; IMDM; HI-HS; Fluorometric metabolic indicator; SD; TNF-α; Tumor necrosis factor; HI-FBS; rmTNF-α; Biological fluid; Cytokine; Colorimetry; Method; Fluorescent compound; Detection; Quantitative analysis
• ISSN: 0022-1759; 1872-7905
• DOI: 10.1016/0022-1759(94)90361-1

Tumor necrosis factor is an important cytokine involved in inflammation and assay of this cytokine in biological fluids may be important in the understanding of several disease processes. This report describes an improved TNF bioassay employing a newly isolated subclone of the cell line NCTC-clone 929 as well as a novel fluorescence indicator system for detecting of viability of the target cells. The limit of detection for the TNF hypersensitive cell line with this fluorescence viability assay 68 ± 2.5 fg/ml, which is approximately 3 × more sensitive than the parental clone and approximately 10 × more sensitive than that reported by Branch et al. (1991) using the neutral red indicator system. The hypersensitivity of the clone gradually declined over a 45-day period and at regular intervals new cells were cultivated from frozen stocks. Two different serum sources, bovine fetal serum and horse serum, and four different serum concentrations (5, 10, 15, 20%) were evaluated to optimize sensitivity. No difference was found between serum sources but sensitivity was significantly reduced if < 15% serum was used.