Semi-Quantitative Determination of Differential Gene Expression in Primary Tumors and Matched Metastases by RT-PCR: Comparison with Other Methods

Analysis of differential antigen expression in tumor and metastases vs normal tissue is frequently performed in oncological research. If suitable antibodies are available, its specificity is tested in Western blot, and the distribution of the antigen in the tissue is followed by immunohistochemistry. The information on the relative amount of the antigen in each tissue is obtained by quantitative evaluation of Western blots or enzyme-linked immunosorbent assay (ELISA) data. However, if no antibody is available for antigen of which the coding sequence is known, Northern blotting is the procedure of choice to quantify differences in gene expression at the transcriptional level. This is the second-best solution, since the mRNA level does not always correctly reflect the amount of the antigen at the protein level. Furthermore, the contamination with mRNAs from undesired cells and RNA losses caused by endogenous RNases may distort the completed picture. In fact, when tissue is investigated, total-RNA or whole-cell-lysates are always inadvertently contaminated with material from the nondesired cells.