The label-free detection of human serum albumin (HSA) in aqueous buffer is demonstrated using a simple, monolithic, two-electrode electrochemical biosensor. In this device, both millimeter-scale electrodes are coated with a thin layer of a composite containing M13 virus particles and the electronically conductive polymer poly(3,4 ethylenedioxy thiophene) or PEDOT. These virus particles, engineered to selectively bind HSA, serve as receptors in this biosensor. The resistance component of the electrical impedance, Zre, measured between these two electrodes provides electrical transduction of HSA binding to the virus-PEDOT film. The analysis of sample volumes as small as 50 µL is made possible using a microfluidic cell. Upon exposure to HSA, virus-PEDOT films show a prompt increase in Zre within 5 s and a stable Zre signal within 15 min. HSA concentrations in the range from 100 nM to 5 µM are detectable. Sensor-to-sensor reproducibility of the HSA measurement is characterized by a coefficient-of-variance (COV) ranging from 2–8% across this entire concentration range. In addition, virus-PEDOT sensors successfully detected HSA in synthetic urine solutions.

Caveolin-1 is a target for academic and pharmaceutical research due to its many cellular roles and associated diseases. We report peptide WL47 (1), a small, high-affinity, selective disrupter of cavolin-1 oligomers. Developed and optimized though screening and analysis of synthetic peptide libraries, ligand 1 has 7500-fold improved affinity compared to its T20 parent ligand, and an 80% decrease in sequence length. Ligand 1 will permit targeted study of caveolin-1 function.

Protein engineering by directed evolution can alter proteins' structures, properties, and functions. However, membrane proteins, despite their importance to living organisms, remain relatively unexplored as targets for protein engineering and directed evolution. This gap in capabilities likely results from the tendency of membrane proteins to aggregate and fail to overexpress in bacteria cells. For example, the membrane protein caveolin-1 has been implicated in many cell signaling pathways and diseases, yet the full-length protein is too aggregation-prone for detailed mutagenesis, directed evolution, and biophysical characterization. Using a phage-displayed library of full-length caveolin-1 variants, directed evolution with alternating subtractive and functional selections isolated a full-length, soluble variant, termed cav, for expression in E. coli. Cav folds correctly and binds to its known protein ligands HIV gp41, the catalytic domain of cAMP-dependent protein kinase A, and the polymerase I and transcript release factor. As expected, cav does not bind off-target proteins. Cellular studies show that cav retains the parent protein's ability to localize at the cellular membrane. Unlike truncated versions of caveolin, cav forms large, oligomeric complexes consisting of approximately >50 monomeric units without requiring additional cellular components. Cav's secondary structure is a mixture of α-helices and β-strands. Isothermal titration calorimetry experiments reveal that cav binds to gp41 and PKA with low micromolar binding affinity (K). In addition to the insights into caveolin structure and function, the approach applied here could be generalized to other membrane proteins.