A mild, fast and flexible method for photoimmobilization of biomolecules based on the light-initiated thiol-ene reaction has been developed. After investigation and optimization of various surface materials, surface chemistries and reaction parameters, microstructures and microarrays of biotin, oligonucleotides, peptides, and MUC1 tandem repeat glycopeptides were prepared with this photoimmobilization method. Furthermore, MUC1 tandem repeat glycopeptide microarrays were successfully used to probe antibodies in mouse serum obtained from vaccinated mice. Dimensions of biomolecule microstructures were shown to be freely controllable through photolithographic techniques, and features down to 5 microm in size covering an area of up to 75x25 mm were created. Use of a confocal laser microscope with a UV laser as UV-light source enabled further reduction of biotin feature size opening access to nanostructured biochips.