A crucial step in the development of implanted medical devices, in vivo diagnostics, and microarrays is the effective prevention of nonspecific protein adsorption from real-world complex media such as blood plasma or serum. In this work, a zwitterionic poly(carboxybetaine acrylamide) (polyCBAA) biomimetic material was employed to create a unique biorecognition coating with an ultralow fouling background, enabling the sensitive and specific detection of proteins in blood plasma. Conditions for surface activation, protein immobilization, and surface deactivation of the carboxylate groups in the polyCBAA coating were determined. An antibody-functionalized polyCBAA surface platform was used to detect a target protein in blood plasma using a sensitive surface plasmon resonance (SPR) sensor. A selective protein was directly detected from 100% human blood plasma with extraordinary specificity and sensitivity. The total nonspecific protein adsorption on the functionalized polyCBAA surface was very low (<3 ng/cm (2) for undiluted blood plasma). Because of the significant reduction of nonspecific protein adsorption, it was possible to monitor the kinetics of antigen-antibody interactions in undiluted blood plasma. The functionalization effectiveness and detection characteristics using a cancer protein marker candidate of polyCBAA were compared with those of the conventional nonfouling oligo(ethylene glycol)-based surface chemistry.

Human blood serum and plasma pose significant challenges to blood-contacting devices and implanted materials because of their high nonspecific adsorption onto surfaces. In this work, we investigated nonspecific protein adsorption from single protein solutions and complex media such as undiluted human blood serum and plasma onto poly(carboxybetaine acrylamide) (polyCBAA)-grafted surfaces at different temperatures. The polyCBAA grafting was done via atom-transfer radical polymerization (ATRP) with varying film thicknesses. The objective is to create a surface that experiences "zero" protein adsorption from complex undiluted human blood serum and plasma. Results show that protein adsorption from undiluted human blood serum, plasma, and aged serum on the polyCBAA-grafted surface is undetectable at both 25 and 37 degrees C by a surface plasmon resonance (SPR) sensor. This was achieved with a film thickness of approximately 21 nm. Furthermore, it is demonstrated that the polyCBAA surfaces after antibody immobilization maintain undetectable protein adsorption from undiluted human blood serum. This is the first time that an effective nonfouling material suitable for applications in complex blood media has been demonstrated.

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Zwitterionic carboxybetaine (CB) has unique dual functionality for ligand immobilization on a nonfouling background. The properties of CB groups depend on their spacer groups between the positive quaternary amine groups and the negative carboxyl groups and environmental factors (e.g., ionic strengths and pH values). In this work, five polycarboxybetaines were prepared, including one polycarboxybetaine methacrylate (polyCBMA) and four polycarboxybetaine acrylamides (polyCBAAs) with different spacer groups. The polymers were grafted from a gold surface covered with initiators using surface-initiated atom transfer radical polymerization. Fibrinogen adsorption was measured as a function of ionic strengths and pH values using surface plasmon resonance sensors. The responsive protein adsorption on four polyCBAAs was mapped out. Results show that most of these surfaces exhibit high protein resistance in a wide range of ionic strengths and are more effective than zwitterionic self-assembled monolayers. Although protein adsorption tends to increase at low ionic strength and low pH value, it is still very low for polycarboxybetaines with a methylene, an ethylene, or a propylene spacer group but is more evident for polyCBAA with a longer spacer group (i.e., a pentene group). The response to ionic strengths and pH values can be attributed to the antipolyelectrolyte and protonation/deprotonation properties of polycarboxybetaines, respectively. Both of these properties are related to the spacer groups of CBs.

In this work, we investigate protein adsorption from single protein solutions and complex media such as 100% blood serum and plasma onto poly(sulfobetaine methacrylate) (polySBMA)-grafted surfaces via atom transfer radical polymerization (ATRP) at varying film thicknesses. It is interesting to observe that protein adsorption exhibits a minimum at a medium film thickness. Results show that the surface with 62 nm polySBMA brushes presents the best nonfouling character in 100% blood serum and plasma although all of these surfaces are highly resistant to nonspecific protein adsorption from single fibrinogen and lysozyme solutions. Surface resistance to 100% blood serum or plasma is necessary for many applications from blood-contacting devices to drug delivery. This work provides a new in vitro evaluation standard for the application of biomaterials in vivo.

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