Enzyme immobilization method in packaged freestanding microfluidic channel is developed for biomolecule assembly of biosensor in this research. A new micro fluidic fabrication method is proposed to fabricate freestanding fluidic structure by MEMS technology. The SU-8 polymer channel is treated by plasma treatment to change hydrophobic surface to hydrophilic property which can automatically drive the liquid into the channel by capillary force. The electro click chemistry method is carried out in hydrophilic SU-8 micro channel by capillary effect. The results show that enzyme immobilization can be immobilized by electro click chemistry in completely packaged freestanding channel. The proposed enzyme immobilization method in micro channel is capable of minimizing biosensor device for healthcare application.