Rapid testing devices with cellulose membranebased lateral flow assay (LFA) could play a significant role due to their immense advantageous features to prevent and control infectious diseases like COVID-19. Compared to lab-based testing, it still lags behind due to its lower sensitivity. Herein, to increase the LFA sensitivity, barriers were designed and developed in such a way that there is a gradual increase and decrease of sample flow along with conjugated gold nanoparticles and the flow rate distribution is controlled across the thickness of the cellulose membrane-based LFA device. Finally, the barriers enhance reaction time, resulting in improved sensitivity. The two barriers, one between the conjugate pad and test zone and the other between the test and control zones, improve the detection limit from 2.0 (control or unmodified LFA device) to 0.5 nM, indicating 4-fold higher sensitivity. No additional chemicals or materials are added. ORF1ab nucleic acid of COVID-19 was used as the model target to demonstrate the concept. The stability analysis also supports the potential use of the same concept in other cellulose membrane-based platforms including point-of-care, do-it-yourselfbased microfluidics paper-based analytical devices and environment monitoring purposes.