There are relatively limited studies on the protein of honey samples mainly because of the low amount of protein in honey (0.1-0.5 %), the difficulty in extracting honey protein from the sugar-rich environment, and the hindrance of protein characterization by conventional approaches. Several protein extraction methods such as mechanical (ultrafiltration and ultracentrifugation) and chemical (precipitation) techniques have been applied to different types of honey samples. Most of these studies reported the quantity and molecular size of honey protein from gel electrophoresis, but were unable to identify and characterize the protein. This limitation might be due to the low capacity of analytical equipment in those days. Although different precipitants have also been used, not all them are compatible with mass spectrometric methods during downstream analysis. As a result, the sample preparation step is essential in order to confidently characterize the low and varied amount of honey protein. Nowadays, honey protein is getting attention from researchers because of its potential activity in pharmacological applications. Therefore, honey protein extraction and determination by mass spectrometry are critically reviewed in order to stimulate further honey protein research.