Easy-to-Use Osmosis-Based Microfluidic Chip for Protein Crystallization: Application to a Monoclonal Antibody

Abstract

We developed a disposable microuidic chip which mimics the vapor diusion method for exploring protein crystallization conditions, but at the nanoliter scale. This device exploits the permeation of water through a thin poly(dimethylsiloxane) (PDMS) layer separating droplets stored in the chip and containing a mixture of proteins and precipitants, from an open microuidic reservoir. The water chemical activity xed by the reservoir makes it possible to modify the volume of the droplets in a controlled way (reduction or increase). Since PDMS is only permeable to water, the imposed water activity therefore increases or decreases the concentration of the solutes present in the droplet, and thus consequently the supersaturation of the solution. The specicity of our approach, in addition to the low volumes ensuring controlled mass transport conditions, is that the concentration of all the solutes is known at any time, thus allowing extraction of quantitative information on the crystallization process. We exploit these chips on a protein of therapeutic interest: the full-length monoclonal antibody anti-CD20. Our experiments allow us in particular to estimate both the solubility of this protein and the width of the metastable zone with Na 2 SO 4 and PEG400 as crystallizing agents. We also show that the ne tuning of the permeation rate makes it possible to perform crystallization/dissolution cycles to selectively dissolve small crystals and increase the mean size of the remaining anti-CD20 crystals.

Microfluidic Evaporation, Pervaporation, and Osmosis: From Passive Pumping to Solute Concentration

“…Devices similarly exploiting osmosis through a thin PDMS membrane but working without microvalves and pumps have been also developed by Luo et al and by Morais et al to explore the phase diagram of a full-length monoclonal antibody with its crystallizing agents . Hansen et al.…”

“…Devices similarly exploiting osmosis through a thin PDMS membrane but working without microvalves and pumps have been also developed by Luo et al 227 and by Morais et al to explore the phase diagram of a full-length monoclonal antibody with its crystallizing agents. 228 Hansen et al also developed a microfluidic device integrating both micromechanical valves and a thin PDMS membrane to add kinetic control by osmosis (or pervaporation) on the diffusive mixing of proteins and precipitants in microvolumes, the so-called free-interface diffusion 229 shown in Figure 33B. In this chip, water mass transport across a thin PDMS membrane from/to an open osmotic bath can be used to kinetically optimize protein crystallization.…”

Droplet-based microfluidic platform for viscosity measurement over extended concentration range

“…21 Protein crystallization is a costly and trial-and-error based process that is likewise greatly simplified by the use of micro-devices. 22,23…”

“…21 Protein crystallization is a costly and trial-anderror based process that is likewise greatly simplified by the use of micro-devices. 22,23 We designed a device able to combine some of the aforementioned techniques, offering a simple and reliable platform for investigating viscous features of various hydrophilic compounds.…”

A Three-Dimensional Liquid-Based Exchangeable Gradient Osmosis Chip for a Permeability Controllable Microfluidic Device

“…The osmosis mechanism has more demanding operating conditions than capillary and gravitational methods . Recently, a microfluidic chip that mimics the vapor diffusion method and nanochannels that use thermal osmosis with a temperature gradient have been reported to enhance the osmosis effect. , In this study, the pressure difference between the two channels through the cross-flow method was used to enhance the osmosis effect. In this paper, we introduce the liquid-based exchangeable gradient osmosis (LEGO) chip.…”