QUANTITATIVE PCR FOR DETECTION OF FEMTOGRAM QUANTITIES OF INTERLEUKIN-8 MESSENGER-RNA EXPRESSION

A quantitative polymerase chain reaction (PCR) assay for mRNA expressi on of interleukin-8 (IL-8), a neutrophil chemotactant and activator, w as developed to examine the expression of this cytokine by colonic muc osa. A synthetic IL-8 RNA deleted in size of native IL-8 mRNA was used as an external control. The synthetic IL-8 RNA was mixed with total R NA from cells and converted to cDNA and amplified by PCR simultaneousl y. The lower limit of sensitivity for the assay was found to be more t han 1 femtogram of IL-8 mRNA. The assay determined IL-8 mRNA expressio n when the RNA was isolated from either human histiocytic lymphoma cel l line U937 cells or human colonic mucosa obtained from colitis patien ts and healthy controls. The development of the rapid and sensitive as say should provide a means to more fully evaluate the role of this cyt okine in diverse disease states with small scale.