Bioconjugates represent an emerging class of medicines, which offer therapeutic opportunities overtaking those of the individual components. Many novel bioconjugates have been explored in order to address various emerging medical needs. The last decade has witnessed the exponential growth of new site-selective bioconjugation techniques, however very few methods have made the way into human clinical trials. Here we discuss various applications of site-selective conjugation in biomedicines, including half-life extension, antibody–drug conjugates, conjugate vaccines, bispecific antibodies and cell therapy. The review is intended to highlight both the progress and challenges, and identify a potential roadmap to address the gap.