Background The clinical efficiency of laser treatments is limited by the low penetration of visible light used in certain procedures like photodynamic therapy (PDT). Second Harmonic Generation (SHG) PDT is an innovative technique to overcome this limitation that enables the use of Near Infrared (NIR) light instead of visible light. NIR frequency bands present an optical window for deeper penetration into biological tissue. In this research, we compare the penetration depths of 405 and 808 nm continuous wave (CW) lasers and 808 nm pulsed wave (PW) laser in two different modes (high and low frequency). Methods Increasing thicknesses of beef and chicken tissue samples were irradiated under CW and PW lasers to determine penetration depths. Results The 808 nm CW laser penetrates 2.3 and 2.4 times deeper than the 405 nm CW laser in beef and chicken samples, respectively. 808 nm PW (pulse frequency—500 Hz) penetrates deeper than CW laser at the same wavelength. Further, increasing the pulse frequency achieves higher penetration depths. High frequency 808 nm PW (pulse frequency—71.4 MHz) penetrates 7.4- and 6.0-times deeper than 405 nm CW laser in chicken and beef, respectively. Conclusions The results demonstrate the higher penetration depths of high frequency PW laser compared to low frequency PW laser, CW laser of the same wavelength and CW laser with half the wavelength. The results indicate that integrating SHG in the PDT process along with pulsed NIR light may allow the treatment of 6–7 times bigger tumours than conventional PDT using blue light.