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Effect of Polymer Gel Elasticity on Complex Coacervate Phase Behavior.

ABSTRACT: Gels are key materials in biological systems such as tissues and may control biocondensate formation and structure. To further understand the effects of elastic environments on biomacromolecular assembly, we have investigated the phase behavior and radii of complex coacervate droplets in polyacrylamide (PAM) networks as a function of gel modulus. Poly-l-lysine (PLL) and sodium hyaluronate (HA) complex coacervate phases were prepared in PAM gels with moduli varying from 0.035 to 15.0 kPa. The size of the complex coacervate droplets is reported from bright-field microscopy and confocal fluorescence microscopy. Overall, the complex coacervate droplet volume decreases inversely with the modulus. Fluorescence microscopy is also used to determine the phase behavior and concentration of fluorescently tagged HA in the complex coacervate phases as a function of ionic strength (100-270 mM). We find that the critical ionic strength and complex coacervate stability are nonmonotonic as a function of the network modulus and that the local gel concentration can be used to control phase behavior and complex coacervate droplet size scale. By understanding how elastic environments influence simple electrostatic assembly, we can further understand how biomacromolecules exist in complex, crowded, and elastic cellular environments.

SUBMITTER: Wilcox KG

PROVIDER: S-EPMC11010254 | biostudies-literature | 2024 Apr

REPOSITORIES: biostudies-literature

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Effect of Polymer Gel Elasticity on Complex Coacervate Phase Behavior.

Wilcox Kathryn G KG Yamagami Kai R KR Roopnarine Brittany K BK Linscott Adam A Morozova Svetlana S

ACS polymers Au 20231226 2

Gels are key materials in biological systems such as tissues and may control biocondensate formation and structure. To further understand the effects of elastic environments on biomacromolecular assembly, we have investigated the phase behavior and radii of complex coacervate droplets in polyacrylamide (PAM) networks as a function of gel modulus. Poly-l-lysine (PLL) and sodium hyaluronate (HA) complex coacervate phases were prepared in PAM gels with moduli varying from 0.035 to 15.0 kPa. The siz ...[more]

PMID: 38618006

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Project description:Applications as diverse as drug delivery and immunoassays require hydrogels to house high concentration macromolecular solutions. Yet, thermodynamic partitioning acts to lower the equilibrium concentration of macromolecules in the hydrogel, as compared to the surrounding liquid phase. For immunoassays that utilize a target antigen immobilized in the hydrogel, partitioning hinders introduction of detection antibody into the gel and, consequently, reduces the in-gel concentration of detection antibody, adversely impacting assay sensitivity. Recently, we developed a single-cell targeted proteomic assay with polyacrylamide gel electrophoresis of single cell lysates followed by an in-gel immunoassay. In the present work, we overcome partitioning that both limits analytical sensitivity and increases consumption of costly detection antibody by performing the immunoassay step after dehydrating the antigen-containing polyacrylamide gel. Gels are rehydrated with a solution of detection antibody. We hypothesized that matching the volume of detection antibody solution with the hydrogel water volume fraction would ensure that, at equilibrium, the detection antibody mass resides in the gel and not in the liquid surrounding the gel. Using this approach, we observe (compared with antibody incubation of hydrated gels): (i) 4-11 fold higher concentration of antibody in the dehydrated gels and in the single-cell assay (ii) higher fluorescence immunoassay signal, with up to 5-fold increases in signal-to-noise-ratio and (iii) reduced detection antibody consumption. We also find that detection antibody signal may be less well-correlated with target protein levels (GFP) using this method, suggesting a trade-off between analytical sensitivity and variation in immunoprobe signal. Our volume-matching approach for introducing macromolecular solutions to hydrogels increases the local in-gel concentration of detection antibody without requiring modification of the hydrogel structure, and thus we anticipate broad applicability to hydrogel-based assays, diagnostics, and drug delivery.

Dataset Information

Effect of Polymer Gel Elasticity on Complex Coacervate Phase Behavior.

Publications

Effect of Polymer Gel Elasticity on Complex Coacervate Phase Behavior.

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