sTNF-RII ELISA Kit

Additional Information

Assay Background

Tumor Necrosis Factor is a pleiotropic inflammatory cytokine. On almost all cells receptors for TNF have been detected. TNF-Receptors (TNF-R) were originally designated as TNF-binding proteins. These TNF-binding proteins, present in urine and plasma, were later found to be the extracellular part of the membrane bound cell signaling receptor for TNF-alpha and TNF-beta. Two types of TNF-Receptors (molecules belonging to the family of nerve growth receptor, CD27, CD40 etc) have been identified: TNF-RI (p55/p60, CD120a) and TNF-RII (p75/p80, CD120b). The molecules share substantial identity in their extracellular part although the intra-cellular parts are structurally unrelated. This difference is reflected by the difference in function of the two receptors.
Activation of various cell types leads to proteolytic cleavage of TNF-R resulting in soluble (extracellular part) TNF-R. Soluble TNF-R (sTNF-R) are considered to play a physiological role in TNF binding and inactivation, although also other functions such as stabilization of TNF in circulation have been mentioned. Detection of sTNF-R offers a means of investigating activation of the immune system both in vitro and in vivo. In plasma of healthy individuals approx. 1 – 4 ng/ml of sTNF-R of each type are present.

Assay Principle

The sTNF-RII ELISA is a ready-to-use solid-phase enzyme-linked immunosorbent assay based on the sandwich principle with a working time of 4½ hours. The efficient format of a plate with twelve disposable 8-well strips allows free choice of batch size for the assay. Samples and standards are captured by a solid bound specific antibody. Biotinylated tracer antibody will bind to captured sTNF-RII. Streptavidin-peroxidase conjugate will bind to the biotinylated tracer antibody. Streptavidin-peroxidase conjugate will react with the substrate, tetramethylbenzidine (TMB). The enzyme reaction is stopped by the addition of citric acid. The absorbance at 450 nm is measured with a spectrophotometer. A standard curve is obtained by plotting the absorbance (linear) versus the corresponding concentrations of the sTNF-RII standards (log). The sTNF-RII concentration of samples, which are run concurrently with the standards, can be determined from the standard curve.