An iBody-based lateral flow assay for semi-quantitative determination of His-tagged protein concentration.

The polyhistidine tag (His-tag) is one of the most commonly used epitope tags in protein engineering. While His-tagged proteins can be detected reliably using immunological methods such as ELISA and Western blot, these methods are costly and time-intensive, necessitating more facile solutions for preliminary qualitative determination and concentration estimation. To this end, we present a rapid test strip assay based on iBody antibody mimetics that target the His-tag. We compare this strategy to commercial antibody-based assays and discuss the advantages and caveats of lateral flow assay design. Our test strip detected a panel of His-tagged proteins with different tag attachment strategies with a visual detection limit of 1 μM and densitometric detection limit of 0.5 μM. Due to its chemical nature, the presented assay exhibits wide reagent compatibility in comparison to antibody-based assays.