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. 2012 Jun 4;7(6):e37845. doi: 10.1371/journal.pone.0037845

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Seeger et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.

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(A) Specificity ELISA using bLmrCD_AviC, bMsbA_AviC and bAcrB_AviC as target proteins. Seven DARPins (α-LmrCD#1-5, DARPin_Act2 and DARPin_Act3) were found to be highly specific for bLmrCD_AviC. Many initial DARPin binder-hits promiscuously bound to bLmrCD_AviC, bMsbA_AviC and bAcrB_AviC as exemplified with the “unsp. DARPin” and were therefore not useful for further analysis. DARPins specific for bMsbA_AviC (DARPin_55) and bAcrB_AviC (110819) were used as a positive control. (B) ELISA analyzing binding of the LmrCD-specific DARPins shown in (A) to LmrC (bLmrC-GFP), LmrD (bLmrD-GFP) and the nucleotide binding domain of LmrD (bLmrD-NBD_AviN). Binding to LmrCD (bLmrCD_AviC) was confirmed as positive control.