Materials that are resistant to nonspecific protein adsorption are critical in the biomedical community. Specifically, nonfouling implantable biomaterials are necessary to reduce the undesirable, but natural foreign body response. The focus of this investigation is to demonstrate that polyampholyte hydrogels prepared with equimolar quantities of positively charged [2-(acryloyloxy)ethyl] trimethylammonium chloride (TMA) and negatively charged 2-carboxyethyl acrylate (CAA) monomers are a viable solution to this problem. TMA/CAA hydrogels were prepared and their physical and chemical properties were characterized. The fouling resistance of the TMA/CAA hydrogels were assessed at varying cross-linker densities using enzyme-linked immunosorbant assays (ELISAs). The results clearly demonstrate that TMA/CAA hydrogels are resistant to nonspecific protein adsorption. A unique advantage of the fouling resistant TMA/CAA system is that bioactive proteins can be covalently attached to these materials using standard conjugation chemistry. This was demonstrated in this study through a combination of ELISA investigations and short-term cell adhesion assays. The multifunctional properties of the TMA/CAA polyampholyte hydrogels shown in this work clearly demonstrate the potential for these materials for use as tissue regeneration scaffolds for many biomedical applications.

Polyampholyte polymer systems are composed of varying mixtures of charged monomer subunits. These polymeric systems have gained increasing attention because it is possible to design the final material properties through careful selection of the charged monomer subunits and controlling the polymer architecture. Characteristics that have been manipulated include the hydration, mechanical properties, pH responsive swelling, temperature responsive swelling, resistance to nonspecific protein adsorption, and protein conjugation capability. This had led researchers to propose the use of polyampholyte polymers as biosensor platforms, fouling release membranes, drug delivery vehicles, and tissue engineering scaffolds. This review is focused on advances that have been made over the last 5 years to develop polyampholyte polymers for these biomedical applications. V C 2013 Wiley Periodicals, Inc. J. Appl.Polym. Sci. 2014, 131, 40069.

Label-free biosensor technologies have the potential to revolutionize environmental monitoring, medical diagnostics, and food safety evaluation processes due to their unique combinations of high-sensitivity signal transducers and high-specificity recognition elements. This enables their ability to perform real-time detection of deleterious compounds at extremely low concentrations. However, to further improve the biosensors' performance in complex environments, such as wastewater, blood, and urine, it is necessary to minimize nonspecific binding, which in turn will increase their specificity, and decrease the rate of false positives. In the present work, we illustrate the potential of combining emerging high-sensitivity optical signal transducers, such as whispering gallery mode (WGM) microcavities, with covalently bound poly(ethylene glycol) (PEG) coatings of varying thickness, as an effective treatment for the prevention of nonspecific protein adsorption onto the biosensor surface. We monitor the sensitivity of the coated biosensor, and investigate the effect of PEG chain length on minimizing nonspecific adsorption via protein adsorption studies. Experimental results confirm not only that PEG-functionalization reduces nonspecific protein adsorption to the surface of the sensor by as much as a factor of 4 compared to an initialized control surface, but also that chain length significantly impacts the nonfouling character of the microcavity surface. Surprisingly, it is the short chain PEG surfaces that experience the best improvement in specificity, unlike many other systems where longer PEG chains are preferred. The combination of WGM microcavities with PEG coatings tuned specifically to the device will significantly improve the overall performance of biosensor platforms, and enable their wider application in complex, real-world monitoring scenarios.

Native bone tissue is composed of a matrix of collagen, non-collagenous proteins, and calcium phosphate minerals, which are primarily hydroxyapatite (HA). The SIBLING (small integrin-binding ligand, N-linked glycoprotein) family of proteins is the primary non-collagenous protein group found in mineralized tissues. In this work, the mineralization induction capabilities of three of the SIBLING members, bone sialoprotein (BSP), osteopontin (OPN), and the calcium binding subdomain of dentin sialophosphoprotein, dentin phosphoprotein (DPP), are directly compared on a biomimetic collagen substrate. A self-assembled, loosely aligned collagen fibril substrate was prepared and then 125I radiolabeled adsorption isotherms were developed for BSP, OPN, and DPP. The results showed that BSP exhibited the highest binding capacity for collagen at lower concentrations, followed by DPP and OPN. However, at the highest concentrations all three proteins had similar adsorption levels. The adsorption isotherms were then used to identify conditions that resulted in identical amounts of adsorbed protein. These substrates were prepared and placed in simulated body fluid for 5 hours, 10 hours, and 24 hours at 37°C. The resulting mineral morphology was assessed by atomic force microscopy and the composition was determined using photochemical assays. Mineralization was seen in the presence of all of the proteins. However, DPP was seen to be the only protein that formed individual mineral nodules similar to those seen in developing bone. This suggests that DPP plays a significant role in the biomineralization process and that the incorporation of DPP into tissue engineering constructs may facilitate the induction of biomimetic mineral formation.

Dentin sialophosphoprotein (DSPP) is a member of the SIBLING (small integrin binding N-linked glycoprotein) family of proteins commonly found in mineralized tissues. Dentin phosphoprotein (DPP) is a naturally occurring subdomain of DSPP that contains the cell binding RGD sequence. Previously, the orientation and conformation of other SIBLING family members specifically bound to collagen I have been investigated with respect to their cell adhesion properties. In this study, the orientation of DPP under similar circumstances is examined, and the results are discussed relative to the previous investigations. Radiolabeled adsorption isotherms were developed for DPP adsorbing to both tissue culture polystyrene (TCPS) and collagen coated TCPS. Then, a MC3T3-E1 cell adhesion assay was performed on TCPS and collagen coated TCPS in the presence of identical amounts of adsorbed DPP. It was discovered that there was a significant difference in the number of bound cells on the TCPS and collagen coated TCPS, with a preference for TCPS. Furthermore, a cell inhibition assay was conducted to confirm that the cell binding that occurred was due to specific integrin interactions with the RGD sequence of DPP. These results suggest that the orientation of DPP, rather than its conformation, dictates the accessibility of the cell binding RGD domains of DPP and that the RGD sequence in DPP is less accessible when DPP is specifically bound to collagen. The results obtained in this study are in stark contrast to previous studies with related SIBLING proteins, and suggest that DPP does not play a key role in cell binding to the collagen matrix of developing bone.

Bone tissue is comprised of collagen, non-collagenous proteins, and hydroxyapatite and the SIBLING (small integrin binding, N-linked glycoprotein) family of proteins is the primary group of non-collagenous proteins. By replicating the native interactions between collagen and the SIBLING proteins at the interface of an implant, it is believed that a bone scaffold will more easily integrate with the surrounding tissue. In this work, bone sialoprotein, osteopontin (OPN), dentin sialoprotein (DSP), dentin phosphoprotein (DPP), C-terminal fragment of dentin matrix protein 1 (DMP1-C), and proteoglycan versions of DSP (DSP-PG) and DMP1 (DMP1-PG) were tested individually to determine their roles in collagen fibrillogenesis and the prevention of denaturation. It was shown that DSP and DPP slowed down fibrillogenesis, while other SIBLINGs had limited impact. In addition, the denaturation time was faster in the presence of DSP and OPN, indicating a negative impact. The role of calcium ions in these processes was also investigated. The presence of calcium ions sped up fibrillogenesis in all scenarios tested, but it had a negative impact by reducing the extent. Calcium also sped up the denaturation in most cases, with the exception of DMP1-C and DSP where the opposite was seen. Calcium had a similar effect on the proteoglycan variants in the fibrillogenesis process, but had no impact on the denaturation process in the presence of these two. It is believed that incorporating DMP1-C or DSP on the surface of a bone implant may improve the collagen interactions with the implant, thereby facilitating improved osteointegration.

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