A durable and biocompatible ascorbic acid-based covalent coating method of polydimethylsiloxane for dynamic cell culture

Abstract

Polydimethylsiloxane (PDMS) is widely used in dynamic biological microfluidic applications. As a highly hydrophobic material, native PDMS does not support cell attachment and culture, especially in dynamic conditions. Previous covalent coating methods use glutaraldehyde (GA) which, however, is cytotoxic. This paper introduces a novel and simple method for binding collagen type I covalently on PDMS using ascorbic acid (AA) as a cross-linker instead of GA. We compare the novel method against physisorption and GA cross-linker-based methods. The coatings are characterized by immunostaining, contact angle measurement, atomic force microscopy and infrared spectroscopy, and evaluated in static and stretched human adipose stem cell (hASC) cultures up to 13 days. We found that AA can replace GA as a cross-linker in the covalent coating method and that the coating is durable after sonication and after 6 days of stretching. Furthermore, we show that hASCs attach and proliferate better on AA cross-linked samples compared with physisorbed or GA-based methods. Thus, in this paper, we provide a new PDMS coating method for studying cells, such as hASCs, in static and dynamic conditions. The proposed method is an important step in the development of PDMS-based devices in cell and tissue engineering applications.

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“…This approach mainly consists of the attachment and distribution of endothelial cells. Due to the hydrophobic nature of cured PDMS that does not allow a convenient surface for cell attachment, surface modifications are needed such as APTES and PLL coatings (Leivo et al, 2017). In our approach, APTES was used as a linker chemical to promote protein attachment.…”

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“…Furthermore, we performed stretching continuously for 7 days and did not see any problems with cell adhesion. In this study, cell adhesion was enhanced utilizing the covalent surface functionalization with ascorbic acid (Leivo et al 2017). Ascorbic acid cross-links the gelatin covalently to the membrane and therefore provides a stable base for the cells during long-term stretching.…”

“…Surface functionalization is needed to render the initially hydrophobic surface of PDMS to be suitable for the attachment of extracellular matrix (ECM) proteins and eventually cells. In these experiments, we used a modified version of the covalent functionalization protocol recently introduced by Leivo et al (2017). Briefly, a three-step covalent functionalization procedure was applied targeting the nitrogen-rich molecular sidechains of extracellular matrix proteins (such as gelatin) to bind permanently onto the substrate.…”

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“…There was also a small peak appeared at around 1661 cm −1 after the GA grafting, indicating the vibration of C=N after APETS and GA reaction [ 18 ]. Finally, after the collagen grafting, the collagen functional groups emerged around 3375 and 1636 cm −1 due to the association of O-H and N-H stretching in collagen, as it has been proven that the amide indication could be observed around 3350, 1700–1600 cm −1 [ 19 , 20 ]. While in the adsorption group ( Fig.…”