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This report describes for the first time the development of paper-based enzymatic reactors (PERs) for the detection of glucose (Glu) in artificial serum sample using a 3D printed batch injection analysis (BIA) cell coupled with electrochemical detection. The fabrication of the PERs involved firstly the oxidation of the paper surface with a sodium periodate solution. The oxidized paper was then perforated with a paper punch to create microdisks and activated with a solution containing N -hydroxysuccinimide (NHS) and N -(3-dimethylaminopropyl)- N ′-ethylcarbodiimide hydrochloride (EDC). Glucose oxidase (GOx) enzyme was then covalently immobilized on paper surface to promote the enzymatic assay for the detection of Glu in serum sample. After the addition of Glu on the PER surface placed inside a plastic syringe, the analyte penetrated through the paper surface under vertical flow promoting the enzymatic assay. The reaction product (H 2 O 2 ) was collected with an electronic micropipette in a microtube and analyzed in the 3D BIA cell coupled with screen-printed electrodes (SPEs). The overall preparation time and the cost estimated per PER were 2.5 h and $0.02, respectively. Likewise the PERs, the use of a 3D printer allowed the fabrication of a BIA cell within 4 h at cost of $5. The coupling of SPE with the 3D printed cell exhibited great analytical performance including repeatability and reproducibility lower than 2% as well as high sampling rate (30 injections h −1 ) under low injection volume (10 μL). The limit of detection (LD) and linear range achieved with the proposed approach was 0.11 mmol L −1 and 1–10 mmol L −1 , respectively. Lastly, the glucose concentration level was successfully determined using the proposed method and the values found were not statistically different from the data achieved by a reference method at confidence level of 95%. [ABSTRACT FROM AUTHOR]