Using a reference collection of blood samples of Red Knots (Calidris canutus) from Florida and Alaska whose sex was determined by dissection, we demonstrate that the molecular method of sexing based on the size of polymerase chain reaction (PCR) products from the CHD genes after digestion with Hae III accurately identifies gender in this species. In males, the 110 basepair PCR product is cleaved into two smaller fragments, whereas in females this fragment remains uncut. Molecular sexing of other samples of Red Knots from the Dutch Wadden Sea, southern Brazil, and Delaware Bay revealed that methods using a discriminant function on size or plumage dimorphism are only partly effective in correctly assigning sex of birds. Given the low cost and accuracy of molecular sexing on very small blood samples, we anticipate that it will find increasing use in studies of the evolutionary ecology of shorebirds and other avian species.