An electrochemical paper-immunosensor and method for detecting aminoglycoside antibiotics is developed in the present invention. Single-walled carbon nanotubes are coated on the common filtration paper by dip-dry cycles. With antibody against neomycin adding to the coating solution, a high sensitive biosensor for specific detection of neomycin is prepared, satisfying to the rigid authority regulations. The sensor is not only sensitive but also rapid, comparing with the classic ELISA method, with LOD of 0.04 ng mL−1 and the whole detection process lasting less than 30 min. Another notable advantage of this invention is the versatility of the sensor, similar method is engaged to prepare the versatile sensor for other aminoglycoside antibiotics, replacement with relevant antibodies.