Abstract: Background and Objective: Proteomics has been developed remarkably and become an intensive field of research; especially protein glycosylation which is considered one of the most common post-translational modifications. Glycosylated proteins play vital roles in several biological and molecular pathways and it represents over 50% of human proteins. The aim of this study was to examine differentially expressed glycoprotein profiles of breast cancer and its adjacent tissues among breast cancer women in Malaysia. Materials and Methods: A total of 48 pairs of normal and cancerous tissues were collected from breast cancer women at two hospitals in Penang, Malaysia: Seberang Jaya Hospital (SJH) and Kepala Batas Hospital (KBH) from January 2010- January 2013. Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE) was employed to map protein expression profiles. Gel-Eluted Liquid Fractionation Entrapment Electrophoresis (GELFREE) system was used for separation and fractionation of protein extracts. The protein extracts were subjected to molecular weight fractionation via electrophoretic separation through SDS PAGE material in tubular environment, all in the liquid phase. The enrichment of GELFREE fractions was carried out using glycopeptide enrichment strategy. After that, they were fragmented by tandem mass spectrometry (LC-MS/MS) analysis using LC/MS LTQ-Orbitrap Fusion. The protein profiling was done using PEAKS Client (7.5) software. Results: In this study, 11 glycoproteins were detected only in tumour tissues, but not in normal ones. Extracted protein concentrations, number of proteins and glycopeptides were found to be higher in tumour tissue samples compared to normal tissue samples. Conclusion: The findings of this study provided useful insights into the glycoproteins identification and could establish fundamental insights for further differential glycoproteomics profiling for breast cancer biomarker discovery.