A small, simple intravital microcirculation multi-spectral imaging system was constructed. Orthogonal polarization spectral (OPS) imaging and liquid crystal tunable filter (LCTF) were used for intravital microcirculation multi-spectral imaging. LCTF has excellent imaging quality and low-power loss, without moving parts and image shift. It can provide continuously, rapid and random-access selection of any wavelength in the visible to near-infrared range. An ear of nude mouse was adopted as experiment specimen. The multi-spectral images of ear microvessel of the nude mouse were acquired by this imaging system in two ways (detection polarization angle 90 degrees and 0 degrees). Compared with detection polarization angle 0 degrees way the images acquired by orthogonal polarization spectral imaging way(detection polarization angle 90 degrees) had better contrast and more details. It is concluded that the experiment has proved that the orthogonal polarization multi-spectral technique can enhance the effect of observation of intravital microcirculation.