Fabrication of Robust Hydrogel Coatings on Polydimethylsiloxane Substrates Using Micropillar Anchor Structures with Chemical Surface Modification

A durable hydrophilic and protein-resistant surface of polydimethylsiloxane (PDMS) based devices is desirable in many biomedical applications such as implantable and microfluidic devices. This paper describes a stable antifouling hydrogel coating on PDMS surfaces. The coating method combines chemical modification and surface microstructure fabrication of PDMS substrates. Three-(trimethoxysilyl)­propyl methacrylates containing CC groups were used to modify PDMS surfaces with micropillar array structures fabricated by a replica molding method. The micropillar structures increase the surface area of PDMS surfaces, which facilitates secure bonding with a hydrogel coating compared to flat PMDS surfaces. The adhesion properties of the hydrogel coating on PDMS substrates were characterized using bending, stretching and water immersion tests. Long-term hydrophilic stability (maintaining a contact angle of 55° for a month) and a low protein adsorption property (35 ng/cm² of adsorbed BSA-FITC) of the hydrogel coated PDMS were demonstrated. This coating method is suitable for PDMS modification with most crosslinkable polymers containing CC groups, which can be useful for improving the anti-biofouling performance of PDMS-based biomedical microdevices

Appendix B. A figure showing alkaline/acid phosphatase (APA) activity relative to leucine-aminopeptidase (LAMP) activity and net nitrogen uptake by periphyton from Stony Brook incubated in Stony Brook water and Stony Creek water receiving prior exposure to substrata from an abandoned mine drainage (AMD)-impacted stream in order to lower the P concentration of Stony Brook water.

A figure showing alkaline/acid phosphatase (APA) activity relative to leucine-aminopeptidase (LAMP) activity and net nitrogen uptake by periphyton from Stony Brook incubated in Stony Brook water and Stony Creek water receiving prior exposure to substrata from an abandoned mine drainage (AMD)-impacted stream in order to lower the P concentration of Stony Brook water

Appendix A. A figure showing phosphorus removal by communities from the three anthracite streams compared to removal by colonized substrata following autoclaving.

A figure showing phosphorus removal by communities from the three anthracite streams compared to removal by colonized substrata following autoclaving

DataSheet3.CSV

<p>Temperature dictates the performance of aquatic ectotherms. However, the physiological and biochemical processes that drive thermally-mediated life history patterns (and limits) remain poorly understood because they are rarely studied simultaneously. In our previous work, we have established life history outcomes (e.g., survivorship, development time, growth rates, and fitness) in mayflies (Neocloeon triangulifer) reared at static temperatures ranging from 14 to 30°C at 2°C intervals. In this study, we conducted biochemical measurements (RT-qPCR of select genes and targeted, quantitative metabolomic profiling) on N. triangulifer mature larvae reared at temperatures associated with excellent survival and fitness (22–24°C), compromised survival and fitness (28°C), and chronic lethality (30°C—larvae survived for a few weeks but failed to emerge to adulthood). Patterns of gene expression were similar to those observed in acute ramping experiments reported previously: larvae reared at 30°C resulted in significant upregulation in the thermally responsive gene HEAT SHOCK PROTEIN 90 (HSP90) but no significant changes in hypoxia responsive genes [EGGLAYING DEFECTIVE9 (EGL-9) and LACTATE DEHYDROGENASE (LDH)]. Additionally, primers for genes associated with energy: INSULIN RECEPTOR (IR), mechanistic TARGET OF RAPAMYCIN (mTOR), and TREHALOSE 6 PHOSPHATE SYNTHASE (T6PS) were developed for this study. IR and mTOR were significantly upregulated while T6PS showed trend of downregulation in larvae reared at 30°C. Metabolomic profiles revealed general depletion of lipids and acylcarnitines in larvae exposed to chronic thermal stress, suggesting that larvae were energetically challenged despite continuous access to food. For example, concentrations of lysoPhosphatidylcholine (lysoPC) a C20:3 decreased as fitness decreased with increasing temperature (2.3- and 2.4-fold at 28 and 30°C relative to controls). Tissue concentrations of the biogenic amine histamine increased 2.1- and 3.1-fold with increasing temperature, and were strongly and negatively correlated with performance. Thus, both histamine and lysoPC a C20:3 are potential biomarkers of thermal stress. Taken together, our results primarily associate energetic challenge with thermally mediated fitness reduction in N. triangulifer.</p

DataSheet1.xlsx

<p>Temperature dictates the performance of aquatic ectotherms. However, the physiological and biochemical processes that drive thermally-mediated life history patterns (and limits) remain poorly understood because they are rarely studied simultaneously. In our previous work, we have established life history outcomes (e.g., survivorship, development time, growth rates, and fitness) in mayflies (Neocloeon triangulifer) reared at static temperatures ranging from 14 to 30°C at 2°C intervals. In this study, we conducted biochemical measurements (RT-qPCR of select genes and targeted, quantitative metabolomic profiling) on N. triangulifer mature larvae reared at temperatures associated with excellent survival and fitness (22–24°C), compromised survival and fitness (28°C), and chronic lethality (30°C—larvae survived for a few weeks but failed to emerge to adulthood). Patterns of gene expression were similar to those observed in acute ramping experiments reported previously: larvae reared at 30°C resulted in significant upregulation in the thermally responsive gene HEAT SHOCK PROTEIN 90 (HSP90) but no significant changes in hypoxia responsive genes [EGGLAYING DEFECTIVE9 (EGL-9) and LACTATE DEHYDROGENASE (LDH)]. Additionally, primers for genes associated with energy: INSULIN RECEPTOR (IR), mechanistic TARGET OF RAPAMYCIN (mTOR), and TREHALOSE 6 PHOSPHATE SYNTHASE (T6PS) were developed for this study. IR and mTOR were significantly upregulated while T6PS showed trend of downregulation in larvae reared at 30°C. Metabolomic profiles revealed general depletion of lipids and acylcarnitines in larvae exposed to chronic thermal stress, suggesting that larvae were energetically challenged despite continuous access to food. For example, concentrations of lysoPhosphatidylcholine (lysoPC) a C20:3 decreased as fitness decreased with increasing temperature (2.3- and 2.4-fold at 28 and 30°C relative to controls). Tissue concentrations of the biogenic amine histamine increased 2.1- and 3.1-fold with increasing temperature, and were strongly and negatively correlated with performance. Thus, both histamine and lysoPC a C20:3 are potential biomarkers of thermal stress. Taken together, our results primarily associate energetic challenge with thermally mediated fitness reduction in N. triangulifer.</p

DataSheet2.xlsx

<p>Temperature dictates the performance of aquatic ectotherms. However, the physiological and biochemical processes that drive thermally-mediated life history patterns (and limits) remain poorly understood because they are rarely studied simultaneously. In our previous work, we have established life history outcomes (e.g., survivorship, development time, growth rates, and fitness) in mayflies (Neocloeon triangulifer) reared at static temperatures ranging from 14 to 30°C at 2°C intervals. In this study, we conducted biochemical measurements (RT-qPCR of select genes and targeted, quantitative metabolomic profiling) on N. triangulifer mature larvae reared at temperatures associated with excellent survival and fitness (22–24°C), compromised survival and fitness (28°C), and chronic lethality (30°C—larvae survived for a few weeks but failed to emerge to adulthood). Patterns of gene expression were similar to those observed in acute ramping experiments reported previously: larvae reared at 30°C resulted in significant upregulation in the thermally responsive gene HEAT SHOCK PROTEIN 90 (HSP90) but no significant changes in hypoxia responsive genes [EGGLAYING DEFECTIVE9 (EGL-9) and LACTATE DEHYDROGENASE (LDH)]. Additionally, primers for genes associated with energy: INSULIN RECEPTOR (IR), mechanistic TARGET OF RAPAMYCIN (mTOR), and TREHALOSE 6 PHOSPHATE SYNTHASE (T6PS) were developed for this study. IR and mTOR were significantly upregulated while T6PS showed trend of downregulation in larvae reared at 30°C. Metabolomic profiles revealed general depletion of lipids and acylcarnitines in larvae exposed to chronic thermal stress, suggesting that larvae were energetically challenged despite continuous access to food. For example, concentrations of lysoPhosphatidylcholine (lysoPC) a C20:3 decreased as fitness decreased with increasing temperature (2.3- and 2.4-fold at 28 and 30°C relative to controls). Tissue concentrations of the biogenic amine histamine increased 2.1- and 3.1-fold with increasing temperature, and were strongly and negatively correlated with performance. Thus, both histamine and lysoPC a C20:3 are potential biomarkers of thermal stress. Taken together, our results primarily associate energetic challenge with thermally mediated fitness reduction in N. triangulifer.</p