High‐Throughput and Dosage‐Controlled Intracellular Delivery of Large Cargos by an Acoustic‐Electric Micro‐Vortices Platform. Issue 1 (29th October 2021)

• Record Type: Journal Article
• Authors: Aghaamoo, Mohammad; Chen, Yu‐Hsi; Li, Xuan; Garg, Neha; Jiang, Ruoyu; Yun, Jeremy Tian‐Hao; Lee, Abraham Phillip
• Is Part Of: Advanced science. Volume 9:Issue 1(2022)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2021-10-29
• Subjects: CRISPR‐Cas9 -- intracellular delivery -- large cargo -- precise‐dose delivery; Science -- Periodicals; 505
• DOI: 10.1002/advs.202102021
• Languages: English
• ISSNs: 2198-3844

Abstract

Abstract: A high‐throughput non‐viral intracellular delivery platform is introduced for the transfection of large cargos with dosage‐control. This platform, termed Acoustic‐Electric Shear Orbiting Poration (AESOP), optimizes the delivery of intended cargo sizes with poration of the cell membranes via mechanical shear followed by the modulated expansion of these nanopores via electric field. Furthermore, AESOP utilizes acoustic microstreaming vortices wherein up to millions of cells are trapped and mixed uniformly with exogenous cargos, enabling the delivery of cargos into cells with targeted dosages. Intracellular delivery of a wide range of molecule sizes (<1 kDa to 2 MDa) with high efficiency (>90%), cell viability (>80%), and uniform dosages (<60% coefficient of variation (CV)) simultaneously into 1 million cells min −1 per single chip is demonstrated. AESOP is successfully applied to two gene editing applications that require the delivery of large plasmids: i) enhanced green fluorescent protein (eGFP) plasmid (6.1 kbp) transfection, and ii) clustered regularly interspaced short palindromic repeats (CRISPR)‐Cas9‐mediated gene knockout using a 9.3 kbp plasmid DNA encoding Cas9 protein and single guide RNA (sgRNA). Compared to alternative platforms, this platform offers dosage‐controlled intracellular delivery of large plasmids simultaneously to large populations of cells while maintaining cell viability at comparable delivery efficiencies. Abstract : A versatile multimodal acoustic‐electric micro‐vortices platform capable of efficient and precise‐dosage delivery of large cargos into both adherent and suspension cells is presented. With this platform, large‐plasmid (>9kbp) transfection for clustered regularly interspaced short palindromic repeats (CRISPR)‐Cas9 with uniform dosages (<60% coefficient of variation (CV)) at 1 million cells min −1 per single chip is demonstrated.