Abstract
Stereolithography (SLA) and 3D printing in general have become a major manufacturing method for producing microfluidic devices. Consumer-grade printers combine resolutions around 50 µm, large printing volumes, and high printing speed (<2 h for a full chip), and their prices are continuously dropping. However, concerns remain concerning the cytocompatibility of commercial SLA resins, which can release, after 3D printing, a variety of chemicals, which can interfere with biological processes and/or compromise cell viability [1]. Several strategies have been reported to limit the resin toxicity, e.g., using coatings [2], post-curing [3], autoclaving [4].
In previous work, we proposed a generic post-treatment to address the fact that 3D-printed molds inhibit PDMS curing, using UV exposure and a thermal step [5]. We showed that this treatment eliminated volatile fragments of photo-initiators and promoted molecular recombination, preventing thereby undesired leaching at a later stage. Here, we evaluated if similar treatments reduce the cytotoxicity of six SLA resins, while following ISO standard 10993-12:2012. Small bricks of 5x5x3 mm3 were printed using a Formlabs-3 or DWS-29J+ printer and, respectively, Formlabs Clear, BioMed and Elastic resins and DWS DS3000 (transparent, biocompatible grade), DL260 (composite with ceramic) and GL4000 (flexible) resins. Bricks were exposed to UV for 1-2 h (FormCure, 405 nm, 5.5 mW/cm²) and to 120°C for 2-48 h. Following the same ISO standard, bricks were incubated in culture medium at a surface-to-volume ratio of 3 cm²/ml for 24 h. To examine cytocompatibility of the materials after treatment, these conditioned mediums were added to various cell types.
First, human monocytic cells THP-1 were differentiated into macrophages using phorbol 12-myristate-13-acetate (PMA), next incubated with conditioned RPMI-1640 medium for 24 h, and their metabolic activity measured with a PrestoBlue assay. For all resins except DS3000, untreated materials led to a significant decrease in metabolic activity. For all resins, a minimal duration for the treatment restored the metabolic activity, possibly above 100% (unconditioned medium) (Fig. 1). We hypothesized that activities above 100% are due to further differentiation into M1 macrophages, which are known to exhibit higher metabolism. We are now testing this hypothesis.
Next, germ cells (bull and boar semen, and bull oocytes) were used, following the same procedure, yet other media. The impact of two resins (Formlabs Clear and BioMed) with (1 h UV, 4 h at 120°C) or without treatment was evaluated in terms of semen motility (CASA measurement) and oocyte maturation. The sperm motility for medium conditioned with treated resins was slightly closer to the control, compared to untreated resins (Fig. 2). Oocyte maturation (oocyte reaching metaphase II stage; Fig. 3) was almost fully inhibited without any treatment of the 3D printed resins, and remained lower after treatment, this inhibition being more marked for the Clear (35%) than for the BioMed resin (70%).
Altogether, we have proposed a generic post-treatment to improve the SLA-resin cytocompatibility. Currently, we are analyzing the leachates from the resins that compromise cell viability and function and developing an oviduct-on-chip using the BioMed resin to study the process of fertilization.
In previous work, we proposed a generic post-treatment to address the fact that 3D-printed molds inhibit PDMS curing, using UV exposure and a thermal step [5]. We showed that this treatment eliminated volatile fragments of photo-initiators and promoted molecular recombination, preventing thereby undesired leaching at a later stage. Here, we evaluated if similar treatments reduce the cytotoxicity of six SLA resins, while following ISO standard 10993-12:2012. Small bricks of 5x5x3 mm3 were printed using a Formlabs-3 or DWS-29J+ printer and, respectively, Formlabs Clear, BioMed and Elastic resins and DWS DS3000 (transparent, biocompatible grade), DL260 (composite with ceramic) and GL4000 (flexible) resins. Bricks were exposed to UV for 1-2 h (FormCure, 405 nm, 5.5 mW/cm²) and to 120°C for 2-48 h. Following the same ISO standard, bricks were incubated in culture medium at a surface-to-volume ratio of 3 cm²/ml for 24 h. To examine cytocompatibility of the materials after treatment, these conditioned mediums were added to various cell types.
First, human monocytic cells THP-1 were differentiated into macrophages using phorbol 12-myristate-13-acetate (PMA), next incubated with conditioned RPMI-1640 medium for 24 h, and their metabolic activity measured with a PrestoBlue assay. For all resins except DS3000, untreated materials led to a significant decrease in metabolic activity. For all resins, a minimal duration for the treatment restored the metabolic activity, possibly above 100% (unconditioned medium) (Fig. 1). We hypothesized that activities above 100% are due to further differentiation into M1 macrophages, which are known to exhibit higher metabolism. We are now testing this hypothesis.
Next, germ cells (bull and boar semen, and bull oocytes) were used, following the same procedure, yet other media. The impact of two resins (Formlabs Clear and BioMed) with (1 h UV, 4 h at 120°C) or without treatment was evaluated in terms of semen motility (CASA measurement) and oocyte maturation. The sperm motility for medium conditioned with treated resins was slightly closer to the control, compared to untreated resins (Fig. 2). Oocyte maturation (oocyte reaching metaphase II stage; Fig. 3) was almost fully inhibited without any treatment of the 3D printed resins, and remained lower after treatment, this inhibition being more marked for the Clear (35%) than for the BioMed resin (70%).
Altogether, we have proposed a generic post-treatment to improve the SLA-resin cytocompatibility. Currently, we are analyzing the leachates from the resins that compromise cell viability and function and developing an oviduct-on-chip using the BioMed resin to study the process of fertilization.
Original language | English |
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Publication status | Published - 20 Oct 2023 |
Event | 27th International Conference on Miniaturized Systems for Chemistry and Life Sciences, µTAS 2023 - Katowice, Poland Duration: 15 Oct 2023 → 19 Oct 2023 Conference number: 27 |
Conference
Conference | 27th International Conference on Miniaturized Systems for Chemistry and Life Sciences, µTAS 2023 |
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Abbreviated title | MicroTAS 2023 |
Country/Territory | Poland |
City | Katowice |
Period | 15/10/23 → 19/10/23 |