Labeling of viruses can be used to reveal viral infection pathways and screen potential anti-viral drugs. Complex procedures, including virus cultivation, purification and labeling are involved in traditional virus labeling methods. And the manipulation of living virus brings risk to researcher health. In this work, we report a general method for site-specific labeling of the envelope virus in an integrated microfluidic device with simple procedures and high security. Site-specific labeling of virus was achieved by fusing the biotin acceptor peptide (AP-tag) and the biotin ligase enzyme (BirA enzyme) with the envelope protein GP64 of baculovirus. The AP-tag could be modified by BirA enzyme to introduce the biotin moiety onto the viral envelope. Western blots and fluorescence colocalization analysis proved that the baculoviruses were biotinylated and labeled with high efficiency. The integrated device incorporated several operation steps including cell seeding, cell culture, cell transfection, virus culture and virus labeling. Since virus biotinylation was achieved during the process of virus cultivation, the complex procedures of virus labeling were simplified in our device. Furthermore the whole process could be completed in the integrated microfluidic device, and direct contact between viruses and researchers could be eliminated in our method, which could greatly reduce the risk to researcher health during living virus labeling.