Commercial manufacture of wheat starch and vital wheat gluten involves physical separation of starch granules and gluten particles formed in a neutral aqueous system. The wet-separation of gluten proteins and starch from wheat flour is based on their water insolubility, density, and particle size. Upon wetting, gluten proteins in wheat endosperm aggregate and form particles that are larger in size but less dense than starch granules. Wheat starch and vital gluten are currently produced industrially through wet-milling of wheat flour principally by four processes; the Martin, Alfa-Laval/Raisio, Hydrocyclone, and the High-Pressure Disintegration (HD) processes. The industrial processes differ mainly in the forms of the flour–water mixtures presented to the fractionation equipment (centrifuge, hydrocyclone, or screen) or in the initial separation practice of starch and gluten fractions from flour. However, the processes essentially merge into one as the intermediate starch and gluten-particle streams are purified to give >98% pure starch and ∼80% protein vital gluten. Handling of a second-grade starch stream, flour water-soluble, and fibrous residues may differ at various processing steps. Small-scale tests are available to assess the wet-milling quality of flours to be wet-processed by the Martin, Batter, Alfa-Laval/Raisio, and the HD processes. Yet, there are no laboratory tests reported for the Hydrocyclone process. Given the development of numerous new wheat varieties each year and of high-amylose and waxy wheats, it is important that small-scale wet-milling tests to assess the wet-milling quality and suitability of flour samples for the industrial processes be available.