To date, there is no valid analytical method available to determine the total whey protein (TWP) content in cheese. Therefore, the aim of this study was to develop HPLC methods for whey protein determination in mature cheese. For that purpose, foil-ripened whey protein-enriched model cheese and traditional Edam-type cheese were produced and analyzed. Suitable protein extraction methods and subsequent analytical methods to determine the acid-soluble whey protein (ASWP) and TWP content in cheese were developed. To characterize the influence of proteolysis on native and denatured whey proteins, the ASWP and TWP contents were determined throughout the ripening process. Both chromatographic methods showed that the individual whey proteins (α-lactalbumin and β-lactoglobulin) were not degraded during ripening. However, the analyzed ASWP content increased by up to 25% throughout ripening. Compared to traditional Edam-type cheese, the β-lactoglobulin content in whey protein-enriched cheese (containing 30% high-heat milk) increased by a factor of 3.5. To evaluate the chromatographic results, two different calculation models were used to estimate a reference value for the TWP content in the manufactured cheese. Further studies are required to optimize the quantification of TWP content in hard, semihard, soft, and cream cheeses.