The isolation of native multiprotein complexes, particularly those involving virus–host or viral protein interactions, remains a fundamental approach for identifying functional interaction partners of viral proteins. The Strep/FLAG-TAP tag (commonly abbreviated as the SF tag) is a modified version of tandem affinity purification (TAP). Unlike traditional TAP purification, which relies on proteolytic cleavage steps, the SF tag simplifies the process by eliminating the need for a tobacco etch virus (TEV) protease cleavage site. This approach allows target proteins to be purified under mild conditions through either single-step or tandem purification. The SF tag typically consists of two StrepII tags and one FLAG tag, enhancing its utility for protein complex isolation. This streamlined design improves the efficiency of the purification process, yielding high-purity protein complexes and minimizing nonspecific elution. This method is particularly effective in eukaryotic systems, especially in mammalian cells, where the method results in excellent preservation of complex stability while reducing contamination. This chapter explores the mechanisms underlying tandem affinity purification using the SF tag, presenting a comprehensive guide for experimental procedures. Additionally, it explores innovations and modifications to the SF-tag system that extend its application beyond protein complex purification to include functional studies.