The invention discloses a rigid ceramic/agarose composite microsphere and a preparation method thereof. The composite microsphere has a uniform composite structure of a rigid ceramic skeleton and agarose gel. The preparation method comprises the following steps of: firstly, preparing ceramic slurries, and uniformly mixing nano-silica powder, zirconia powder or titanium dioxide powder with sodium alginate solution; secondly, preparing a porous ceramic microsphere, inverse suspension dispersing the ceramic slurries in an oil phase, dropping calcium chloride solution to for curing into microspheres, collecting the microspheres, drying, forming the microspheres to through high temperature sintering; thirdly, filling agarose, immersing the ceramic microspheres into agarose solution, and cooling and solidifying after preserving the temperature in a sterilizing pot for a period of time; and fourthly, grinding and screening: grinding a solide mixture of ceramic and the agarose in a mortar, obtaining the microspheres with the required particle size through screening in water, and removing the agarose on the surfaces of the microspheres. The microsphere has the advantages of good rigidity, good controllable density, good hydrophilicity, good sphericity and low cost. Shown by test experiments, the microsphere can be used as a matrix of biomacromolecule chromatographic separation medium.