Background

Biomaterials play a pivotal role in the development of advanced medical devices, drug delivery systems, and regenerative therapies to enhance patient outcomes and quality of life [1–4]. However, current drawbacks include device-associated infection, adverse immune responses, and in-service degradation that can collectively reduce implant performance [5–7]. Rational design of bio-instructive materials remains unattainable due to our limited understanding of material–biological interactions [1]. As a result, screening is a commonly employed approach for discovering and optimising novel biomaterials with a desired biological function, such as pro- or anti-inflammatory properties. High-throughput screening (HTS) strategies have accelerated the development of biomaterials by enabling researchers to rapidly analyse a large number of samples or conditions in a systematic manner [8]. A commonly used HTS approach for biomaterials is the use of printed polymer microarrays, which have been employed to screen cell responses to materials, allowing reproducible control over cellular behaviour [9]. Key examples are screening for scalable synthetic cultureware for human pluripotent stem cells [10], polymers to modulate the foreign body response and wound healing [11,12], and a new class of bacteria-attachment-resistant materials [13]. In these studies, thousands of materials are robotically printed and in situ ultraviolet (UV)-polymerised on a single slide. This approach is used to discern specific biological interactions by culturing cells directly on the polymer array surfaces within a shared culture medium [14,15]. Despite the success of such polymer microarray platforms in biomaterials’ discovery, the secreted biochemical signals are an untapped resource in understanding cellular phenotypes. Paracrine signalling is also a worry requiring follow-up studies on individually scaled up polymers before potential hits can be verified. Understanding cell-surface interactions requires probing biochemical cues, in addition to biomechanical, topographical, and material chemistry/bio-interfacial cues. Hence, without knowledge of the secretome profile of the cells following their interaction with materials, the understanding of the functionality of the polymer is incomplete. Furthermore, polymer microarrays are limited to the assessment of cells with strong adhesion abilities. Notably, for cells such as dendritic and T cells, which possess weaker substrate adhesion tendencies, attributing their phenotypical changes to a particular polymer spot would be incredibly challenging [16–18].

In response to these limitations of the polymer microarray, we developed a novel platform that combines existing arraying technology such as contact and inkjet printing, multi-well microchambers, and high-throughput secretome profiling. The platform benefits from reusable superstructures (ProPlate^®) that are mountable on the printed glass substrates. This offers the provision of confined culture volumes, each designated for a distinct polymer condition with a surface area and volume compatible with cell culture for a few days. This separation allows us to closely study the mediators released by cells, helping us understand how these materials interact with cells and the mechanisms of the biomolecules involved. In addition, this system allows for high-throughput surface characterisation such as time-of-flight secondary ion mass spectrometry, x-ray photoelectron spectroscopy, and attenuated total reflectance–Fourier transform infrared spectroscopy (ATR-FTIR), ensuring the accurate identification and validation of controlling moieties [19]. Together, these analytical approaches have the potential to enhance the discovery of new biomaterials and improve our understanding of biomaterials–cells interface. Yet, there are constraints to the throughput and time-related aspects of this platform. In a single operational cycle, the platform demonstrates the capacity to fabricate ten slides, each accommodating 16 unique chemical compositions, resulting in the synthesis of 160 distinct polymer entities, thereby yielding 8,960 discrete polymer spots. By way of comparison, a polymer microarray system can generate 17,280 polymer deposition sites from 576 chemistries in triplicates across ten slides [20]. Nevertheless, this limited output can be ameliorated through the implementation of 64-well superstructures from GraceBio-Labs and the application of multiplexing methodologies as alternatives to conventional techniques such as enzyme-linked immunosorbent assay (ELISA). Here, we provide a step-by-step description of methodologies and troubleshooting aspects involved in developing this platform, including preparation of substrate, fabrication of multi-well chambers, and provision of confined area for each polymer condition, followed by relevant examples of the anticipated outcomes such as chemical characterisation and biological assessments of the secreted soluble mediators.