<p>The fabrication process described herein offers a straightforward and cost-effective alternative to traditional methods, thereby eliminating the need for complex instrumentation. This approach addresses the increasing demand for environmentally sustainable laboratory solutions by employing biodegradable materials to fabricate paper-based analytical devices. This study presents a straightforward, instrument-free screen-printing technique for fabricating biodegradable paper-based microtiter plates. The devices are constructed from a patterned cellulose substrate coated with a hydrophobic polycaprolactone (PCL) polymer. This coating is precisely configured to define discrete hydrophilic wells, establish lateral hydrophobic barriers, and form a continuous hydrophobic backing layer, thereby achieving effective spatial isolation of the hydrophilic regions. Clear and well-defined interfaces were observed between hydrophilic and hydrophobic zones. The method demonstrated a minimum channel width of 513 ± 16&#xa0;μm for delivering aqueous samples. The process has high reliability in fabrication, with a strong linear correlation (R² = 0.99) observed between the set and actual widths. The fabricated paper microtiter plates were evaluated through colorimetric assays for Bovine Serum Albumin, glucose, and phosphate, utilizing a standard microplate reader and a flatbed scanner. The results exhibited a strong correlation (R² = 0.98) between the two measurement platforms and demonstrated good agreement with absorbance values obtained using commercial well plates. These findings underscore the effectiveness of the proposed method for fabricating paper microtiter plates, which offer a cost-effective and disposable alternative to conventional well plates, offering comparable analytical performances.</p>