A method of measuring tissue microstructure parameter γ with optical fiber probe was presented. Based on spherical fractal model of Intralipid, the diffuse reflection data of the photon through Intralipid was obtained using Monte Carlo simulation, and the function relationship between diffuse reflectance and γ was established by non-linear fitting of the data. Research shows that the diffuse reflectance is linearly related to the dimensionless reduced scattering coefficient μs′ ( is selecting aperture) and its slope is a quadratic function of γ. In the experiment, the diffuse reflectance of different μs′ was obtained with Intralipid, the linear relationship between diffuse reflectance and γ was verified. The γ of 2.04, the fractal dimension of 2.93 and the anisotropy factor of 0.816 were obtained by the slope, which were in good agreement with the parameters in literature, verifying the feasibility of the proposed method.