We have developed an efficient and inexpensive method of reverse transfection from the solid phase to suppress genes with siRNA. The method enabled the realization of (i) a high efficiency of transfection; (ii) transfection of various types of cell; (iii) a high efficiency of gene knockdown by siRNA; (iv) a low toxicity to cells; and (v) a long-term stabilization (more than 210 d) of attached transfection mixture including siRNA in multiple wells. Although array-based reverse transfection has advantages in terms of miniaturization, the method has the advantage of enabling the inclusion of various soluble factors, such as humoral factors, drugs and ligands that affect gene expression, because the liquid phase is partitioned within the individual wells of each microtiter plate. Our method of reverse transfection with siRNA in multiple wells is a powerful and high-throughput tool for the analysis of signaling pathways.