Comparative evaluation of quantitative PCR methods

This chapter quantitatively evaluates interleukin 5 (IL-5) gene expressions in the bronchoalveolar lavage (BAL) cells of patients with asthma. For this purpose, a method has been used that was sensitive, accurate, rapid, and easy to execute. Quantitative polymerase chain reaction (PCR) following the transcription of messenger RNA (mRNA) to complementary DNA (cDNA) is a sensitive method for evaluating cytokine mRNA expression in cells. There are two general methods of quantitative PCR: competitive and noncompetitive. Considerable emphasis has been paid to the competitive method of quantitative PCR because this method is internally controlled and is thought to have the ability to reduce tube-to-tube variation of PCR products. In contrast to the competitive method, noncompetitive method is relatively simple and rapid. However, the accuracy of the noncompetitive method, relative to the competitive method (the gold standard method) is not known. The two methods of quantitative PCR utilizing three test molecules and two reference molecules are compared in the chapter. Experimental data indicate that the values obtained by the noncompetitive method of quantitating IL-5 cDNA levels are comparable to those of the competitive method of quantitative PCR.