AN ELISA FOR MEASURING TUMOR-NECROSIS-FACTOR-ALPHA IN RAT PLASMA

Tumour necrosis factor (TNF) is a cytokine with multiple biological ac tivities which plays a pivotal role in the response of the body to inf ection. TNF is secreted in the the monomeric form and associates to yi eld a biologically active oligomeric molecule. Bioactive TNF can be me asured in plasma using cytotoxic assays which employ the murine cell-l ines L929 or WEHI 164 clone 13. However, it has become clear that inac tive TNF also circulates in vivo, which is not detected by bioassay. T hese inactive forms may play an important role in the regulation of TN F activity. The rat is often used as a model for the study of acute in fection and its systemic effects. Our aim was to develop an ELISA for rat TNF which would provide a convenient and cost effective method of assay. The assay employs two commercially available antibodies raised against recombinant murine TNF (rMuTNF) which exhibit cross-reactivity with rat TNF. The lower limit of detection for this assay was determi ned to be 39.0 pg/ml rMuTNF. The inter and intra-assay coefficients of variation were < 12.0%. Specificity of the assay was shown by the hig h degree of parallelism obtained between rMuTNF and commercially avail able rRatTNF. The assay described measures rat TNF in both plasma and tissue culture supernatant. The measurement of plasma concentrations o f TNF using both the ELISA and bioassay may help elucidate more fully the biological importance of TNF.