Atomic force microscopy (AFM) was applied to the (-101) faces of calcium oxalate monohydrate (COM) crystals grown from calcium oxalate (CaOx) solutions. Microstructures of many spiral hillocks with step height of 1 nm were observed on the faces. Then using AFM in situ, we analysed the re-growth process of the spiral steps on the face of COM seed-crystals in CaOx growth solutions that contained growth inhibitors of glycosaminoglycans and studied their inhibition mechanisms on COM crystals. The total morphology of the faces of COM seed crystals re-grown in the CaOx growth solutions was assessed by scanning electron microscopy (SEM). In the growth solution without glycosaminoglycans (control experiment) or with chondroitin sulphate (ChS), AFM images and SEM micrographs of the faces of the re-grown seed crystals showed two-dimensional (2D) nucleation although 2D nucleation was delayed in the presence of ChS. However, the addition of dermatan sulphate (DS) to the growth solution resulted in isotropic growth by a step flow mode and spiral mechanism. With regard to the main inhibition mechanisms of two glycosaminoglycans (ChS and DS) on COM crystals, it can be concluded from these results that ChS delays 2D nucleation and DS inhibits 2D nucleation.