In-situ investigation of cellular level biomarkers using atomic force microscopy (AFM) based nano robot

Abstract

The rising requirements for superior, non-destructive analytical techniques capable of maintaining resolution at the nanoscale, particularly for in-situ analysis of sensitive samples such as living cells and viruses, are pressing. Existing methodologies fall short in characterizing the intricate structure and mechano-sensing transmission within cells. The establishment of tools capable of concurrently assessing diverse cellular features like protein recognition, mechanical biomarkers, force regulation, and morphology is a pivotal advancement in our comprehension of intercellular processes.

Atomic force microscopy (AFM), as a nanoscale sensor, is capable of obtaining comprehensive parametric information with sub-nanometer spatial resolution. The AFM architecture involves a force-sensing cantilever probe that minimizes sample damage through physical contact facilitated by laser reflection. Considering its control over the tip-cantilever structure, the AFM probe can function as a self-sensing robotic end-effector. This study enhances the sensing performance of AFM through the integration of multiple nano-mechanical sensors, auxiliary systems, and force sensing and imaging algorithms while aiming to reduce the AFM inertia sensing deficiency, imaging artifacts and manipulation error.

The fusion of advanced AFM-based nano-robot systems and label-free mechanical biomarkers enables cellular-level exploration and offers a benign and stable approach for biological research. The combined influence of cytoskeleton architecture, trans-membrane proteins, and signaling pathways facilitate communication with the micro-environment and internal organelles, controlling cell morphology, proliferation, and migration. An auscultation system is employed to stimulate cells with mechanical vibration, permitting quantitative sensing of cellular viscoelasticity. This approach allows for comprehensive mechanical profiling of cellular structure with superior spatial-temporal resolution, in contrast to traditional micro-rheology experiments that only capture information near the membrane. The integration of conventional measurements with auscultation enables the decoupling of cellular internal structures through vibration signal modeling.

In the realm of pathological and clinical research, we have engineered a nano-robot system equipped with an incubator permitting dynamic, long-term evaluation of multi-parametric changes at a single cell level. In response to the Covid-19 pandemic, an autonomous robot equipped with 270 nm UVGI LEDs was developed, facilitating the evaluation of UVGI's disinfection effects on bacteria, viruses, and virus-attacked Vero cells. The nano-robot was utilized to investigate embryo and endometrium implantation, emphasizing the crucial role of the Cadherin-1 protein and p120-catenin gene in the process. In an extended study of embryo/human blastoid behavior, we integrated a micromanipulator-AFM nanorobot with closed-loop control via micro-mechanical sensors, enabling simultaneous manipulation and scanning of the embryo. This approach broadens the application of AFM, permitting precise measurements of large, non-adhesive objects, with significant implications for embryo research.