McSweeney Megan A, Patterson Alexandra T, Loeffler Kathryn, Cuellar Lelo de Larrea Regina, McNerney Monica P, Kane Ravi S, Styczynski Mark P
School of Chemical and Biomolecular Engineering, Georgia Institute of Technology, Atlanta, GA 30332, USA.
Sci Adv. 2025 Feb 21;11(8):eado6280. doi: 10.1126/sciadv.ado6280.
Conventional laboratory protein detection techniques are not suitable for point-of-care (POC) use because they require expensive equipment and laborious protocols, and existing POC assays suffer from long development timescales. Here, we describe a modular cell-free biosensing platform for generalizable protein detection that we call TLISA (T7 RNA polymerase-linked immunosensing assay), designed for extreme flexibility and equipment-free use. TLISA uses a split T7 RNA polymerase fused to affinity domains against a protein. The target antigen drives polymerase reassembly, inducing reporter expression. We characterize the platform and then demonstrate its modularity by using 16 affinity domains against four different antigens with minimal protocol optimization. We show that TLISA is suitable for POC use by sensing human biomarkers in serum and saliva with a colorimetric readout within 1 hour and by demonstrating functionality after lyophilization. Altogether, this technology has the potential to enable truly rapid, reconfigurable, modular, and equipment-free detection of diverse classes of proteins.
传统的实验室蛋白质检测技术不适合即时检测（POC），因为它们需要昂贵的设备和繁琐的操作流程，而且现有的即时检测方法开发时间长。在此，我们描述了一种用于通用蛋白质检测的模块化无细胞生物传感平台，我们称之为TLISA（T7 RNA聚合酶连接免疫传感测定），其设计具有极高的灵活性且无需设备。TLISA使用与针对蛋白质的亲和结构域融合的分裂T7 RNA聚合酶。目标抗原驱动聚合酶重新组装，诱导报告基因表达。我们对该平台进行了表征，然后通过使用针对四种不同抗原的16个亲和结构域，在最小化方案优化的情况下展示了其模块化。我们表明，TLISA适用于即时检测，可在1小时内通过比色读数检测血清和唾液中的人类生物标志物，并在冻干后仍能发挥功能。总之，这项技术有潜力实现对各类蛋白质的真正快速、可重构、模块化且无需设备的检测。
