Aptamers are single-stranded synthetic oligonucleotides that are able to capture their target molecule with high affinity and specificity. Therefore, they can be thought of as nucleic acid-based alternative to antibodies, which have several advantages over their amino acid-based counterparts. Consequently, aptamers can be used in different applications based on molecular recognition including affinity separations. This review will summarize the state-of-the-art in aptamer-based affinity separations; will discuss the current limitations and will highlight possible future prospects. The first part will point out the advantages of aptamers in downstream processing. Here, the properties of aptamers will be discussed along with their implications on downstream processing from a user's point of view. In the second part, a brief summary of the literature is given with focus on aptamer-based separation of proteins. Finally, some drawbacks of aptamers will be illustrated and possibilities to overcome these limitations will be suggested. New technologies in the fields of aptamer selection and synthesis are expected to further promote the use of aptamers as affinity ligands in downstream processing.