Pengzhan Liu

Cryo–electron microscopy (cryo-EM) has revolutionized structural biology by enabling imaging of macromolecular complexes at near-atomic resolution, but its success depends critically on specimen quality. Standard plunge freezing often produces nonuniform particle distributions, resulting in poor particle statistics and reduced imaging quality, particularly for vesicular structures and low-concentration, labile macromolecular complexes. Here, we introduce acoustofluidic cryo-EM (ACE), a noncontact specimen-preparation method that integrates acoustofluidic manipulation into a conventional plunge-freezing workflow. ACE enables localized nanoscale particle manipulation by mechanically coupling acoustic excitation into the cryo-EM grid, where forced vibration generates localized acoustic streaming through viscous boundary-layer effects. Guided by this physical mechanism, validation using liposomes and apoferritin demonstrates that ACE increases particle density by up to fourfold while preserving structural integrity and improving imaging quality. This strategy provides a broadly applicable approach for enhancing cryo-EM specimen quality, improving imaging efficiency, and facilitating high-resolution structural analysis.

Mechanomodulation, the process of altering cellular behavior through applied mechanical forces, plays a critical role in physiological processes and has substantial implications for cancer therapy, immunology, and drug development. However, precise and efficient stimulation of nonadherent cells remains a major challenge, limiting the investigation of mechanotransduction pathways and the development of targeted therapeutics. Here, we developed an acoustofluidic platform named Suspension-cell Targeted Response to Excitation via Acoustofluidic Mechanomodulation (STREAM) to enable precise, high-throughput stimulation of suspension cells. STREAM accomplishes this using 101.14-megahertz high-frequency surface acoustic waves to deliver controlled mechanical stimulation at a throughput of 500,000 cells per minute. STREAM modulates intracellular calcium ion (Ca ²⁺ ) signaling by activating mechanosensitive ion channels, triggering mitochondrial membrane disruption and tunable K562 leukemia cell apoptosis rates from 5.15 to 47.1%. STREAM provides a scalable, precise tool for studying mechanotransduction in suspension cells, with broad applications in cancer research, immunotherapy, and high-throughput drug screening.