Physicochemical Study of Viral Nanoparticles at the Air/Water Interface Jose F. Torres-Salgado, † Mauricio Comas-Garcia, ‡ Maria V. Villagrana-Escareñ o, † Ana L. Dura ́ n-Meza, † Jaime Ruiz-García,* ,† and Ruben D. Cadena-Nava* ,§ † Instituto de Física, Universidad Autó noma de San Luis Potosí, A ́ lvaro Obregó n 64, 78000 San Luis Potosí, San Luis Potosí, Me ́ xico ‡ HIV Dynamics and Replication Program, National Cancer Institute, Frederick, Maryland 21702, United States § Centro de Nanociencias y Nanotecnología, Universidad Nacional Autó noma de Me ́ xico, Carretera Tijuana-Ensenada Km. 107, 22860 Ensenada, Baja California, Me ́ xico *S Supporting Information ABSTRACT: The assembly of most single-stranded RNA (ssRNA) viruses into icosahedral nucleocapsids is a sponta- neous process driven by protein−protein and RNA−protein interactions. The precise nature of these interactions results in the assembly of extremely monodisperse and structurally indistinguishable nucleocapsids. In this work, by using a ssRNA plant virus (cowpea chlorotic mottle virus [CCMV]) as a charged nanoparticle we show that the diffusion of these nanoparticles from the bulk solution to the air/water interface is an irreversible adsorption process. By using the Langmuir technique, we measured the diffusion and adsorption of viral nucleocapsids at the air/water interface at different pH conditions. The pH changes, and therefore in the net surface charge of the virions, have a great influence in the diffusion rate from the bulk solution to the air/water interface. Moreover, assembly of mesoscopic and microscopic viral aggregates at this interface depends on the net surface charge of the virions and the surface pressure. By using Brewster’s angle microscopy we characterized these structures at the interface. Most common structures observed were clusters of virions and soap-frothlike micron-size structures. Furthermore, the CCMV films were compressed to form monolayers and multilayers from moderate to high surface pressures, respectively. After transferring the films from the air/ water interface onto mica by using the Langmuir−Blodgett technique, their morphology was characterized by atomic force microscopy. These viral monolayers showed closed-packing nano- and microscopic arrangements. ■ INTRODUCTION Langmuir monolayers have been widely studied since the early 1900s; 1 most of the studies where done with lipids such as fatty acids, phospholipids, and cholesterol. 2−6 Classical Langmuir monolayers are characterized by amphiphilic molecules where a hydrophilic “head” was attached to a hydrophobic “tail” and the interactions between molecules were relatively easy to study. 7−11 Until now, many more systems such as proteins, carbon nanotubes, 12−15 and fullerenes 16 have been studied at the air/water interface. However, larger biological samples have shown to be quite difficult to study mainly for the complexity of their interactions. The Langmuir technique allows the formation of monolayers or multilayers and the Langmuir− Blodgett technique allows transferring them onto solid substrates as an exact copy of the assembled monolayer at the air/liquid interface. 17−24 These techniques represent an alternative method of building monolayers and multilayers of nanoparticles, such as viruses, of the same surface charge without using polyelectrolytes or without chemical modification of viral cages. 25,26 The studies at the air/water interface have gone beyond the classical set of amphiphilic molecules. Pieranski showed that charged colloidal particles can be trapped irreversibly and assemble into two-dimensional (2D) colloidal crystals due to surface tension effects. 27,28 He proposed that 2D crystal formation is driven by repulsive interactions. Later, Ruiz-Garcia et al. showed that at low colloidal densities, they can also form 2D foamlike and cluster structures 29−31 as a result of additional attractive interactions. The thermodynamics of Langmuir monolayers can be described by the temperature, area, and surface pressure. The surface pressure, Π = γ* − γ, is defined as the difference between the surface tension of pure water, γ*, and the surface tension in the presence of a film or a monolayer, γ. The interaction between colloids can be compared to a dipole− dipole interaction because the part that is in the water dissociates the ions and the part that is in the air does not; therefore, the partial charge on each particle along with its Special Issue: William M. Gelbart Festschrift Received: January 20, 2016 Revised: February 29, 2016 Article pubs.acs.org/JPCB © XXXX American Chemical Society A DOI: 10.1021/acs.jpcb.6b00624 J. Phys. Chem. B XXXX, XXX, XXX−XXX