This study describes the use of smartphone-based detection and multivariate image analysis (MIA) to perform dengue diagnosis on laser printed microzones. Toner-based microplates containing 96 microzones were fabricated on a transparency film surface through a single laser printing step. Enzyme-linked-immunosorbent assay (ELISA) experiments were then carried out to detect both immunoglobulin M (IgM) and G (IgG) antibodies in serum samples collected from patients infected and not infected with dengue virus. Principal component analysis (PCA) of ELISA digital image data acquired with a smartphone revealed good discrimination between samples from infected and non-infected patients.