Background Quantitative measurement of blood flow in specific biotissue targets is crucial for photobiomodulation therapy (PBMT). PBMT has been applied on many clinical applications. However, the conclusive confirmation of its effects on microcirculation and laser wavelength dependency in superficial and deep tissues is lacking. Objective This study examines the microcirculatory response and laser wavelength dependency of red and near infrared (NIR) lasers in biotissue. Laser Doppler flowmetry (LDF) was used to assess the flux and velocity of erythrocytes in buccal tissue when exposed to 660 nm and 830 nm laser radiation. Discussion The microcirculatory response of subjects during and after laser radiation were analyzed. Each laser has an energy density of 73.17 J/cm2 . Both the 660 nm and 830 nm lasers demonstrated the capability to enhance microcirculation in both superficial and deep tissues. Furthermore, we observed that laser wavelengths at 660 nm and 830 nm respectively enhanced the microcirculatory response in superficial and deep tissues. Importantly, this cumulative effect persisted for at least 10 minutes. Conclusions The present results indicate that PBM can effectively improve local blood perfusion in specific target areas and peripheral biotissues. These findings have potential applications in enhancing wound healing and local microcirculation within the target areas.