Paper-based bipolar electrochemical (P-BPEs) assays have attracted a lot of attention due to their low preparation cost, self-pumping property of paper, being portable. The dominant reporting method in the introduced P-BPEs is electrochemiluminescence (ECL). ECL limits the infield application of this type of paper-based sensor as they need expensive photon counting compartment. Therefore, the developing of a colorimetric method instead of ECL can be a promising feature. In the present study, the electrodeposition of Prussian Blue (PB) on the paper fibers has been introduced which has been used directly as a colorimetric method for the reporting cell of P-BPE. This disposable P-BPE biosensor is used for determination of glucose. The closed bipolar electrochemical cell is fabricated on a small part of paper using a laser printing-based process for paper hydrophobization. The bipolar and driving electrodes are provided by pressing the writing pencil HB on the paper. The mechanism of sensing of glucose is oxidation of the analyte in the sensing cell using glucose oxidase followed by reduction of the produced H₂O₂ by application of an external potential (10.0 V). This causes the oxidation of K₄Fe(CN)₆ in the presence of Fe(II) ions and subsequent formation of PB particles in the reporting cell. The intensity of the blue color in the reporting cell is used as a visual and colorimetric signal that can be digitally read using a scanner of digital camera. The parameters affecting the performance of the device were optimized using experimental design and chemometrics modeling. The P-BPE represents a very wide response range that extends from 0.1 mmol.L^-1 to 4.0 mol.L^-1 in the case of hydrogen peroxide, and from 0.1 to 50 mmol.L^-1 in the case of glucose. The limit of detections for hydrogen peroxide and glucose are 4.9 μmol.L^-1 and 70 μmol.L^-1 respectively.