Whey protein concentrates of cows’ milk are used widely in dairy products including infant formulas. A major component is the potent allergen bovine β‐lactoglobulin (BLG). Two easy and sensitive substrate‐amplified indirect competitive ELISAs for BLG, one using a rabbit polyclonal antibody (PAb) K97 raised against heat‐treated BLG, and another using a monoclonal antibody (MAb) 61B4 specific to the epitope Thr125‐Lys135are described. Furthermore, a new whey protein test kit specific for BLG was tested. A new video‐based digital image processing system was used to read, evaluate and calculate data from the microtitre plates. The detection limit was 0.08 ng ml‐1for the PAb assay, which was sufficient for the use of this assay as a test method for ‘milk‐free’ products. The detection limit was 3.2 ng ml‐1for the MAb assay and 144 ng ml‐1’ for the commercially available kit. Thirteen commercially available infant formulas on the Danish market, including four hypoallergenic products and cows’ milk, were tested. The BLG content varied from 0.2 to 727 μg ml‐1, quantified with the PAb assay. The hypoallergenic infant formula with the lowest amount of BLG was Nutramigen containing about 0.1% residual BLG compared with the ordinary infant formula Nidina 1. The knowledge of residual BLG content in hypoallergenic infant formulas is important for a more qualified nutritional recommendation to parents with children genetically susceptible to allergies and for quality assurance of infant formulas claimed to be hypoallergenic.