-The application of channels with multiple thicknesses is a major area of interest within fields of tissue engineering and microfluidics. 3D printing facilitates cost-effective fabrication of PDMS-based microfluidics using 3D printed molds. In this work, the limitations and the accuracy of using 3D printed templates for microfluidic applications with commercial SLA and FDM 3D printers, are presented. A 3D microfluidic cell culturing device that contains multiple thicknesses is proposed and the accuracy of printed parts in three dimensions is demonstrated. The reusable molds can be printed in less than two hours, with the average cost of 0.35 US$, which lead to fast and cheap fabrication compared to conventional fabrication methods which require clean room facilities. The surface roughness of these 3D printed molds are 0.25 and 1.12 for flexible and clear resins.