Ray Optics Model for Optical Trapping of Biconcave Red Blood Cells.

Red blood cells (RBCs) or erythrocytes are essential for oxygenating the peripherical tissue in the human body. Impairment of their physical properties may lead to severe diseases. Optical tweezers have in experiments been shown to be a powerful tool for assessing the biochemical and biophysical properties of RBCs. Despite this success there has been little theoretical work investigating of the stability of erythrocytes in optical tweezers. In this paper we report a numerical study of the trapping of RBCs in the healthy, native biconcave disk conformation in optical tweezers using the ray optics approximation. We study trapping using both single- and dual-beam optical tweezers and show that the complex biconcave shape of the RBC is a significant factor in determining the optical forces and torques on the cell, and ultimately the equilibrium configuration of the RBC within the trap. We also numerically demonstrate how the addition of a third or even fourth trapping laser beam can be used to control the cell orientation in the optical trap. The present investigation sheds light on the trapping mechanism of healthy erythrocytes and can be exploited by experimentalist to envisage new experiments.

FLUIDOT: A Modular Microfluidic Platform for Single-Cell Study and Retrieval, with Applications in Drug Tolerance Screening and Antibody Mining.

Advancements in lab-on-a-chip technologies have revolutionized the single-cell analysis field. However, an accessible platform for in-depth screening and specific retrieval of single cells, which moreover enables studying diverse cell types and performing various downstream analyses, is still lacking. As a solution, FLUIDOT is introduced, a versatile microfluidic platform incorporating customizable microwells, optical tweezers and an interchangeable cell-retrieval system. Thanks to its smart microfluidic design, FLUIDOT is straightforward to fabricate and operate, rendering the technology widely accessible. The performance of FLUIDOT is validated and its versatility is subsequently demonstrated in two applications. First, drug tolerance in yeast cells is studied, resulting in the discovery of two treatment-tolerant populations. Second, B cells from convalescent COVID-19 patients are screened, leading to the discovery of highly affine, in vitro neutralizing monoclonal antibodies against SARS-CoV-2. Owing to its performance, flexibility, and accessibility, it is foreseen that FLUIDOT will enable phenotypic and genotypic analysis of diverse cell samples and thus elucidate unexplored biological questions.

Supported Natural Membranes on Microspheres for Protein-Protein Interaction Studies.

Multiple biological and pathological processes, such as signaling, cell-cell communication, and infection by various viruses, occur at the plasma membrane. The eukaryotic plasma membrane is made up of thousands of different lipids, membrane proteins, and glycolipids, and its composition is dynamic and constantly changing. Due to the central importance of membranes on the one hand and their complexity on the other, membrane model systems are instrumental for interrogating membrane-related biological processes. Here, we develop a new tool for protein-membrane interaction studies. Our method is based on natural membranes obtained from extracellular vesicles. We form membrane bilayers supported on polystyrene microspheres that can be trapped and manipulated using optical tweezers. This method allows working with membrane proteins of interest within a background of native membrane components where their correct orientation is preserved. We demonstrate our method's applicability by successfully measuring the interaction forces between the Spike protein of SARS-CoV-2 and its human receptor, ACE2. We further show that these interactions are blocked by the addition of an antibody against the receptor binding domain of the Spike protein. Our approach is versatile and broadly applicable for various membrane biology and biophysics questions.

Red blood cells under externally induced stressors probed by micro-Raman spectroscopy

Optical techniques are becoming increasingly popular for the analysis of body fluids, particularly so after the onset of the COVID-19 pandemic. Raman spectroscopy has found special significance among them due to its ability to perform label-free investigations of biological solutions with high sensitivity and specificity. The integration of Raman spectroscopy with optical tweezers-Raman Tweezers-has been explored in conducting biochemical investigations on individual red blood cells. We present in this chapter an evaluation of various stress agents, such as intravenous fluids, certain chemicals, and metal nanoparticles, on live, human red blood cells using the Raman Tweezers technique. The technique found efficacy in monitoring hemoglobin deoxygenation, heme aggregation, heme degradation, and membrane damage in red blood cells under the influence of exogenous agents. © 2022 Elsevier Inc. All rights reserved.

Programmed -1 ribosomal frameshifting from the perspective of the conformational dynamics of mRNA and ribosomes.

Programmed -1 ribosomal frameshifting (-1 PRF) is a translation mechanism that regulates the relative expression level of two proteins encoded on the same messenger RNA (mRNA). This regulation is commonly used by viruses such as coronaviruses and retroviruses but rarely by host human cells, and for this reason, it has long been considered as a therapeutic target for antiviral drug development. Understanding the molecular mechanism of -1 PRF is one step toward this goal. Minus-one PRF occurs with a certain efficiency when translating ribosomes encounter the specialized mRNA signal consisting of the frameshifting site and a downstream stimulatory structure, which impedes translocation of the ribosome. The impeded ribosome can still undergo profound conformational changes to proceed with translocation; however, some of these changes may be unique and essential to frameshifting. In addition, most stimulatory structures exhibit conformational dynamics and sufficient mechanical strength, which, when under the action of ribosomes, may in turn further promote -1 PRF efficiency. In this review, we discuss how the dynamic features of ribosomes and mRNA stimulatory structures may influence the occurrence of -1 PRF and propose a hypothetical frameshifting model that recapitulates the role of conformational dynamics.