pH-responsive hydrogels have garnered considerable attention in actuator manipulation, drug delivery, and tissue engineering because of their capacity to undergo structural or volumetric changes in response to acid/base alterations. The fabrication of microstructures of stimuli-responsive biomaterials is crucial in the development of biomedicine and tissue engineering. Bovine serum albumin (BSA) is commonly used in tissue engineering and drug delivery because of its non-toxic, biodegradable, and biocompatible properties. This study presents the macroscopic pH response of BSA-glycidyl methacrylate (BSA-GMA) hydrogels, the microscopic pH response of three-dimensional (3D) hydrogel microstructures polymerized by femtosecond laser direct writing, and cell viability studies. Femtosecond laser direct writing enables the creation of high-precision 3D structures of BSA-GMA hydrogels. The results indicate that the pH responsiveness of the BSA-GMA hydrogels increased with either increasing concentration or decreasing methacrylation degree of BSA-GMA. Unlike the BSA hydrogel, the photopolymerization of the BSA-GMA hydrogel by femtosecond laser direct writing does not deplete amino acid groups. Consequently, the 3D BSA-GMA hydrogel demonstrates a stronger pH response because it contains more amino and carboxyl groups. Furthermore, confocal fluorescence imaging and analysis of relative cell growth rates of chondrocytes on the BSA-GMA scaffolds indicate that the BSA-GMA hydrogel has good biocompatibility. These protein microstructures with controlled morphology and pH-responsive properties have potential applications in tissue engineering, biomedicine, and biosensors.