CSIRO Materials Science and Engineering, Bayview Avenue, Clayton, VIC 3168, Australia.
Colloids Surf B Biointerfaces. 2013 Aug 1;108:313-21. doi: 10.1016/j.colsurfb.2013.03.018. Epub 2013 Mar 20.
We have developed a technique to create 50μm-deep microwells coated with a reactive and robust thin film, which withstands photolithographic processing, and allows for subsequent chemical functionalisation with biological cues (i.e. peptides). First, plasma polymerisation of 1-bromopropane was used to generate a bromine-functionalised thin film (BrPP) on a substrate of silicon wafer. Second, an epoxy functionalised polymer UV photoresist, SU-8, was deposited and developed to create 50μm-deep patterned microwells that display the BrPP coating at their base. Third, amino acids or peptides were selectively attached to the bottom of the microwells through bromine displacement by an amine or thiol nucleophile. Each surface functionalisation step was monitored by XPS, AFM, and contact angle measurements. These functionalities were then used as linkers to immobilise enzymes (e.g. HRP), which retain activity at the end of the process as shown by a biochemical activity assay. Peptide promoters of cell attachment were also immobilised and their functionality was evaluated using an L929 fibroblast adhesion assay. In conclusion, this work describes an innovative combination of plasma thin film deposition and photolithography to create 50μm-deep functionalised microwells for peptide display in biological applications.
我们开发了一种技术，可在深度为 50μm 的微井中形成一层具有反应性和坚固性的薄膜，该薄膜可耐受光刻处理，并可随后用生物信号（例如肽）进行化学官能化。首先，通过等离子体聚合 1-溴丙烷在硅晶片基底上生成溴功能化的薄膜（BrPP）。其次，沉积并开发了一种环氧基官能化的聚合物 UV 光致抗蚀剂 SU-8，以形成深度为 50μm 的图案化微井，其底部显示出 BrPP 涂层。第三，通过胺或硫醇亲核试剂取代溴，将氨基酸或肽选择性地连接到底部的微井中。通过 XPS、AFM 和接触角测量来监测每个表面功能化步骤。然后，将这些官能团用作固定化酶（例如 HRP）的连接物，如通过生化活性测定所示，该酶在该过程结束时仍保持活性。还固定化了促进细胞附着的肽，并使用 L929 成纤维细胞附着测定法评估其功能。总之，这项工作描述了等离子体薄膜沉积和光刻技术的创新组合，可用于在生物应用中显示肽的 50μm 深度功能化微井。
