## 4 Discussion  
In this study, we found that previously developed collagen peptides that support VWF capture and platelet adhesion and aggregation do not adsorb to PDMS in a manner that supports platelet adhesion under flow. Rather, our data suggest that PDMS absorbs collagen peptides. Blocking the microfluidic patterning device with Pluronics F-127 or BSA did not show an increase in platelet adhesion in our flow assays, indicating that blocking the surface with common surfactants used in microfluidics does not effectively disrupt the absorption. However, the absorption of collagen peptides to a PDMS patterning device can be blocked by functionalizing with FOTS. Alternatively, FOTS functionalization of the assay surface (glass or PDMS) provides adsorption that is faster than absorption by the PDMS patterning device. This finding has ramifications, both for using PDMS for micropatterning collagen peptides and for using it as a material for microfluidic flow assays. The absorption of collagen peptides by PDMS is supported by several lines of evidence. First, platelets do not adhere to PDMS surfaces incubated at a concentration of collagen peptides that support adhesion to glass. Second, there was no observable adsorption of peptides on PDMS at the resolution provided by AFM. Third, incubation of collagen peptide solutions in a monolithic PDMS channel results in significant depletion. Finally, significant penetration of a fluorescently labeled collage peptide was observed in untreated PDMS. Untreated PDMS absorbs small, hydrophobic molecules, such as some fluorophores and sex hormones [28; 29]. Molecules with a logarithmic octanol-water partition coefficient (log P) > 2.62, such as rhodamine 6G and diazepam, are strongly absorbed into PDMS [30]. Similarly, the peptide angiotensin II (~1 kDa) is readily absorbed into native PDMS, which can be attenuated by OEGylation [31]. We are unaware of any prior work showing the absorption into untreated PDMS of triple helix peptides in the molecular weight range (~10 kDa) of those used in this study. It is unknown what properties of peptides might influence their absorption into PDMS, although our data shows that the three peptides used here absorb to different degrees. Whether these observations extends to other peptides, and what the molecular weight cut off is for absorption, requires further research. The absorption of collagen peptides in PDMS was blocked when the PDMS was functionalized with a fluorinated alkyl silane. This conclusion is based on observations that incubation of collagen peptides on FOTS-PDMS supported platelet adhesion under flow, showed significant topological changes suggesting adsorption, demonstrated negligible depletion of collagen peptide solutions, and showed negligible absorption into the channel wall as visualized with a fluorescently labeled peptide. Whether FOTS blocks absorption by steric hindrance, by promoting hydrophobic interactions, or by some other mechanism requires further investigation. Nevertheless, in practice this simple modification allows PDMS to be used for either micropatterning collagen peptides to glass slides or as a substrate for adsorption itself. The rate of adsorption of collagen peptides appears to be different for hydrophilic and hydrophobic surfaces. This inference is based on differences observed between untreated and FOTS-functionalized glass and silicon. These findings are supported by molecular dynamics simulations showing that adsorption of a collagen triple helix was more favorable for a hydrophobic surface compared to a hydrophilic surface due to stabilization of the triple helix structure [32]. For untreated glass or silicon, if PDMS was used for patterning collagen peptides, there was little adsorption observed by AFM and negligible amounts of platelet adhesion under flow. For FOTS-glass or FOTS-silicon, significant changes to the surface topology were observe by AFM, and platelet adhesion was supported under flow.