Protein crystallization has long been considered to be an important strategy that offers the high stability and potency for the therapeutic proteins. The difficulty of crystallizing macromolecules such as proteins, however, is well known. Because of their flexibility, glycoproteins are more difficult to crystallize than non-glycosylated proteins. For this reason, the crystallization process for glycoproteins remains a major obstacle in the industrial production of many important therapeutic proteins. In this study, we have developed crystallization process for glycoproteins using alpha-lactalbumin as a model protein. This model protein was microcrystallized using a pH-shift method in polyethylene glycol (PEG) solution as a mother liquor. This microcrystallization process produced uniform and micron-sized alpha-lactalbumin microcrystals. The morphology of these crystals was spherical, and the mean size was about 1-2 mu m when observed by scanning electron microscope (SEM). X-ray diffractometry (XRD) and differential scanning calorimetry (DSC) showed that the microcrystals had a greater degree of crystallinity than alpha-lactalbumin lyophilized amorphous powder. These results suggest that the pH-shift crystallization in PEG solution could be applied to other glycoproteins, thereby increasing the stability of the important glycoprotein therapeutic agents. (c) 2005 Elsevier B.V. All rights reserved.