(343a) Impact of Cell Media Environment on the Salt Response of Surface-Bound Elastin-like Polymer

The Surface Enhanced Electrochemical Diagnostic Sensors (SEEDS) Laboratory is dedicated to creating innovative real-time electrochemical sensors with a wide range of applications, particularly in biomanufacturing. We are currently designing an elastin-like polymer (ELP)-based electrochemical biosensor for on-line monitoring of protein production in a bioreactor; therefore, it is essential to consider the impact environmental factors could have on our system. During fermentation, the protein product (analyte) is commonly secreted into the surrounding cell medium, meaning that our real-time biosensor will have to detect its analyte in a complex solution. Consequently, this complex solution could affect the biosensor surface stability, interfere with the analyte binding, or alter the detection signal in other ways.

Due to the critical role of the ELP component in our biosensor design, this study investigates how cell medium affects the salt-responsive behavior of an ELP, specifically I90, using electrochemical impedance spectroscopy (EIS). ELPs are highly flexible protein polymers that vary in the number of monomeric repeats [VPGXG]_n and identity of the guest residue ‘X’. Modifying these parameters affects the ELP hydrophobicity and, consequently, their stimuli-responsive behavior. I90, the ELP studied, consists of ninety repeats with isoleucine as the guest residue. It also includes an N-terminal cysteine (Cys) residue, which facilitates attachment to a gold surface. EIS measurements were conducted at five different salt concentrations in the presence of a redox mediator pair, ferri-/ferrocyanide (FFCN).

EIS results revealed that I90 responded along a salt gradient, where salt was added to PBS with FFCN, with a reversible behavior. However, in cell medium containing FFCN, the salt response was no longer reversible. The I90-modified surface only responded in the direction from low to high salt concentrations, but not in reverse. We attribute this behavior change to the presence of bovine serum albumin (BSA), a common additive in cell media, which readily binds to gold through its reduced cysteine. We hypothesize that BSA, due to its high concentration (2%), quickly displaced I90 during the salt measurements. Evidence supporting this hypothesis comes from the salt response of a BSA-modified gold surface, which exhibited the same behavior as the I90-modified surface when exposed to the BSA-containing cell medium.

Preliminary experiments with L-Cys as an antifouling agent showed promising results, where the self-assembled monolayer (SAM) of L-Cys successfully repelled BSA. Furthermore, we expect that this antifouling agent will not only repel BSA but also protect I90 from displacement during the salt measurements in cell medium.