Noncollinear acousto-optic tunable filter (AOTF) based on TeO2 is a type of good light splitting device with the electric tuning. Because of its advantages of compact size, high stability, fast tuning, and being easy to carry out, it has high practical application value in hyperspectral imaging field. In this study, a hyperspectral microscopic imaging system was built by combining noncollinear AOTF with optical inverted microscope. In the range of visible light, the hyperspectral imaging of cutaneous squamous cell carcinoma was studied, and the spectra and the corresponding microscopic images at a series of optical central wavelengths were got. The performance of the hyperspectral imaging system was tested. The results shown that the bandwidth of the diffracted light in the range of 110~180 MHz was only 1.28~2.84 nm, which indicated that the AOTF in this study had a high spectral resolution with more than 102 spectral channels, and it could meet the needs of hyperspectral microscopic imaging and accurate identification of biological tissue structure. The system used higher quality TeO2 crystal, higher quality double balsaming lens and optimized RF driver to effectively depress the sidelobe of the diffraction spectrum. The tuning relationship between the acoustic frequency and the diffracted optical wavelength, and the relationship between the spectral bandwidth and the acoustic frequency were analyzed. The experimental results were in good agreement with the related the theoretical calculation. The experimental results shown the high image quality of the system because no obvious image shift with the optical wavelength was observed. By comparing the microscopic images of the cutaneous squamous cell carcinoma with different diffraction central wavelengths, the images were also the clearest at 522.52 nm, and the details of the cutaneous squamous cell carcinoma could be distinguished obviously. The difference of the whole brightness and the transmission difference coefficient with the optical wavelength were studied, and the regulations were agreed with the intuitive observation. Through the analysis of the image edge extraction, the results shown that 497.87~551.29 nm can be used to observe and study the cutaneous squamous cell carcinoma with a bright whole field of vision, meanwhile, the results also shown that 509.69~527.59 nm was the best window for accurate identification and analysis of cutaneous squamous cell carcinoma. This study provided a new method for the simple, flexible and rapid detection and diagnosis of human cutaneous squamous cell carcinoma.