A streamlined approach toward the rapid fabrication of streptavidin–biotin-based protein microarrays was investigated. First, using our engineered versatile plasmid (pBADcM-tBirA) and an optimal coexpression strategy for biotin ligase and biotin acceptor peptide (BAP) chimeric recombinant protein, an autogeneration system for biotinylated probes was developed. This system permitted an advantageous biotinylation of BAP chimeric recombinant proteins, providing a strategy for the high-throughput synthesis of biotinylated probes. Then, to bypass the conventional rate-limiting steps, we employed an on-chip purification process to immobilize the biotinylated probes with high-throughput recombinant lysates. The integration of the autogeneration of probes and on-chip purification not only contributed to the effective and reliable fabrication of the protein microarray, but also enabled simplification of the process and an automated throughput format. This labor- and cost-effective approach may facilitate the use of protein microarrays for diagnosis, pharmacology, proteomics, and other laboratory initiatives.

中文翻译：

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通过生物素化探针的自动生成和片上纯化快速制造蛋白质微阵列。

研究了一种快速制造基于链霉亲和素-生物素的蛋白质微阵列的简化方法。首先，使用我们设计的通用质粒 (pBADcM-tBirA) 和生物素连接酶和生物素受体肽 (BAP) 嵌合重组蛋白的最佳共表达策略，开发了生物素化探针的自动生成系统。该系统允许对 BAP 嵌合重组蛋白进行有利的生物素化，为生物素化探针的高通量合成提供了一种策略。然后，为了绕过传统的限速步骤，我们采用了片上纯化过程，用高通量重组裂解物固定生物素化探针。探针自动生成和芯片上纯化的集成不仅有助于蛋白质微阵列的有效和可靠制造，而且还能够简化过程和自动化吞吐量格式。这种劳动力和成本效益的方法可以促进蛋白质微阵列在诊断、药理学、蛋白质组学和其他实验室计划中的使用。