In this work, the pore size-dependent PPL diffusion and protein distribution in SBA-15 support have been investigated in detail by confocal laser scanning microscopy (CLSM) and X-ray diffraction (XRD) observations in combination with hindered diffusion simulation, intraparticle diffusion analysis, and apparent kinetics calculation. The CLSM observations indicate porcine pancreatic lipase (PPL) diffuses partly into the pores with a diameter of 5.6 nm and much deeper into the pores with a diameter of 8.0 or 9.7 nm. PPL distribution along the pore length has been simulated by hindered diffusion model and the result coincides well with CLSM observations. Besides pore diffusion, boundary resistance is revealed by the intraparticle diffusion analysis of adsorption data. The populations of PPL adsorbed inside the pores or on the external surface are estimated. A higher PPL uptake is found in the 8.0 nm than 9.7 rim pores, indicative of the existence of an optimal pore size to match the protein dimension for maximum adsorption capacity. The so-called "confinement" of PPL in the mesoporous supports, revealed by the XRD observation and intraparticle diffusion analysis above, is further confirmed by apparent kinetics calculation.