Institute for Basic Science (IBS), Center for Nanomedicine, Seoul, 03722, Republic of Korea.
Graduate Program of Nano Biomedical Engineering (NanoBME), Advanced Science Institute Yonsei University, Seoul, 03722, Republic of Korea.
Adv Sci (Weinh). 2023 Apr;10(10):e2206872. doi: 10.1002/advs.202206872. Epub 2023 Feb 1.
CRISPR/Cas systems offer a powerful sensing mechanism to transduce sequence-specific information into amplified analytical signals. However, performing multiplexed CRISPR/Cas assays remains challenging and often requires complex approaches for multiplexed assays. Here, a hydrogel-based CRISPR/Cas12 system termed CLAMP (Cas-Loaded Annotated Micro-Particles) is described. The approach compartmentalizes the CRISPR/Cas reaction in spatially-encoded hydrogel microparticles (HMPs). Each HMP is identifiable by its face code and becomes fluorescent when target DNA is present. The assay is further streamlined by capturing HMPs inside a microfluidic device; the captured particles are then automatically recognized by a machine-learning algorithm. The CLAMP assay is fast, highly sensitive (attomolar detection limits with preamplification), and capable of multiplexing in a single-pot assay. As a proof-of-concept clinical application, CLAMP is applied to detect nucleic acid targets of human papillomavirus in cervical brushing samples.
CRISPR/Cas 系统提供了一种强大的感应机制，可将序列特异性信息转化为放大的分析信号。然而，进行多重 CRISPR/Cas 检测仍然具有挑战性，通常需要复杂的多重检测方法。在此，描述了一种基于水凝胶的 CRISPR/Cas12 系统，称为 CLAMP（Cas 加载的注释微颗粒）。该方法将 CRISPR/Cas 反应分隔在空间编码的水凝胶微颗粒（HMP）中。每个 HMP 都可以通过其面码识别，并且当存在靶 DNA 时会发出荧光。该检测方法通过在微流控装置内捕获 HMP 进一步简化；然后，通过机器学习算法自动识别捕获的颗粒。CLAMP 检测法快速、高度灵敏（预扩增时可达皮摩尔检测限），并且能够在单管检测中进行多重检测。作为概念验证临床应用，CLAMP 用于检测宫颈刷样中人类乳头瘤病毒的核酸靶标。
