A POLYMERASE CHAIN REACTION-BASED METHOD FOR THE SEMIQUANTITATIVE STUDY OF INTERLEUKIN-8 MESSENGER-RNA IN HUMAN BASOPHIL LEUKOCYTES

We examined a potential method for quantitative analysis of cytokine e xpression patterns in purified human basophil leukocyte preparations. Basophil mRNA was reverse transcribed and cytokine cDNA levels determi ned by competitive polymerase chain reaction (PCR) with internal stand ard cDNAs constructed by site-directed mutagenesis. Co-reverse transcr ibed glyceraldehyde-3-phosphate-dehydrogenase (GAPDH) cDNA levels were used as an internal control to correct for unequal efficiencies in RN A isolation and reverse transcription. The method was subjected to a s tatistical validation giving the within series precision of the analys is. This method was used to examine interleukin-8 expression patterns in basophils from different donors. The results from this study reveal ed increased interleukin-8 mRNA levels after in vitro challenge with a variety of stimuli.