Cryptosporidium parvum is a waterborne pathogen, yet no suitable surrogate has been established for quantifying its filtration removal in porous media. Carboxyl polystyrene microspheres with size, density, and shape similar to C. parvum were coated with biotin (free and containing amine, NH(2)) and glycoprotein. These biomolecules have isoelectric points similar to C. parvum (pH ≈ 2), and glycoprotein is a major type of surface protein that oocysts possess. Zeta potential (ζ) and filtration removal of particles in sand of two different grain sizes were examined. Compared to unmodified microspheres, modified microspheres achieved a superior match to the oocysts in ζ, concentration, mass recovery, and collision coefficient. They showed the same log reduction in concentration as oocysts, whereas results from unmodified microspheres deviated by 1 order of magnitude. Of the three types of modified microspheres, glycoprotein-coated microspheres best resembled oocyst concentration, despite having ζ similar to NH(2)-biotin-coated microspheres, suggesting that surface protein also played an important role in particle attachment on solid surfaces. With further validation in environmental conditions, the surrogates developed here could be a cost-effective new tool for assessing oocyst filtration in porous media, for example, to evaluate the performance of sand filters in water and wastewater treatment, water recycling through riverbank filtration, and aquifer recharge.