Amplification of cytokine-specific ELISAs increases the sensitivity of detection to 5-20 picograms per milliliter.

The ability to detect a protein is always limited to the sensitivity of the assays available. Progress in improving the sensitivity of protein detection will allow a more complete understanding of biological systems. Of particular interest to the field of immunology is the ability to characterize an immune response based upon the pattern of cytokines that are released in response to antigen. A Th1 response is characterized by the presence of IL-2, IL-12, TNF and IFN-gamma, whereas a Th2 response is characterized by IL-4, IL-5, IL-6 and IL-10. Often, these cytokines are present in in vitro-derived culture supernatants at extremely low concentrations and are therefore very difficult to detect. Although a number of improvements have been made to the sensitivity of the relevant detection assays, the most successful assays involve the presence of the cells being cultured thereby limiting the number of tests per culture to one. Here we describe an enhanced ELISA protocol where the sensitivity is equivalent or better than corresponding cell-based assays. This protocol will permit the sensitive measurement of multiple cytokines per single culture supernatant.