Peptide Self-Assembled Monolayers for Label-Free and Unamplified Surface Plasmon Resonance Biosensing in Crude Cell Lysate

Short peptides, composed of polar or ionic amino acids, derived with a short organic thiol, significantly reduce nonspecific adsorption of proteins in complex biological matrices such as serum and crude cell lysate, which have nonspecific protein concentrations of 76 and 30-60 mg/mL, respectively. Minimizing these nonspecific interactions has allowed rapid and direct quantification of beta-lactamase in a crude cell lysate using a surface plasmon resonance (SPR) biosensor. A library of short peptides with varying chain length and amino acid composition were synthesized using a solid-phase approach. A 3-mer-captopropionic acid (3-MPA) linker was covalently attached to the amino terminus of the peptides to subsequently form a monolayer on gold in the form of 3-MPA(AA)(n)-OH, where n is the length of the amino acid chain (n = 2-5). Leu, Phe, Ser, Asp, and His were selected to investigate the effect on nonspecific adsorption with different physicochemical properties of the sidechains; aliphatic, aromatic, polar, acid, and base. Advancing contact angles measured the hydrophobicity of each peptidic self-assembled monolayer (SAM) and showed that hydrophilicity of the gold surface improved as the chain length of the polar or ionic peptides increased, while aromatic and aliphatic peptides decreased the hydrophilicity as the chain length increased. The nonspecific adsorption of undiluted bovine serum on SPR sensors prepared with the library of 3-MPA-(AA)(n)-OH showed that the lowest nonspecific adsorption occurred with polar or ionic amino acids with a chain length of n = 5. We demonstrate that a monolayer composed of 3-MPA-(Ser)(5)-OH has significant advantages, including the following: (1) it minimizes nonspecific adsorption in undiluted bovine serum; (2) it provides a high surface concentration of immobilized antibodies; (3) it shows a great retention of activity for the antibodies; (4) it improves the response from beta-lactamase by similar to 1. order of magnitude, compared to previous experiments; and (5) it allows direct quantification of submicromolar beta-lactamase concentration in a crude cell lysate with a nonspecific protein concentration of 30-60 mg/mL The use of this peptide-based monolayer offers great advantages for quantitative SPR biosensing in complex biological media.