Weak protein-protein interactions are thought to modulate the viscoelastic properties of concentrated antibody solutions. Predicting the viscoelastic behavior of concentrated antibodies from their dilute solution behavior is of significant interest and remains a challenge. Here, we show that the diffusion interaction parameter (k(D)), a component of the osmotic second virial coefficient (B(2)) that is amenable to high-throughput measurement in dilute solutions, correlates well with the viscosity of concentrated monoclonal antibody (mAb) solutions. We measured the k(D) of 29 different mAbs (IgG(1) and IgG(4)) in four different solvent conditions (low and high ion normality) and found a linear dependence between k(D) and the exponential coefficient that describes the viscosity concentration profiles (|R| ≥ 0.9). Through experimentally measured effective charge measurements, under low ion normality where the electroviscous effect can dominate, we show that the mAb solution viscosity is poorly correlated with the mAb net charge (|R| ≤ 0.6). With this large data set, our results provide compelling evidence in support of weak intermolecular interactions, in contrast to the notion that the electroviscous effect is important in governing the viscoelastic behavior of concentrated mAb solutions. Our approach is particularly applicable as a screening tool for selecting mAbs with desirable viscosity properties early during lead candidate selection.

The present work investigates the influence of electrostatic surface potential distribution of monoclonal antibodies (MAbs) on intermolecular interactions and viscosity. Electrostatic models suggest MAb-1 has a less uniform surface charge distribution than MAb-2. The patches of positive and negative potential on MAb-1 are predicted to favor intermolecular attraction, even in the presence of a small net positive charge. Consistent with this expectation, MAb-1 exhibits a negative second virial coefficient (B₂₂), an increase in static structure factor, S((q→0)), and a decrease in hydrodynamic interaction parameter, H((q→0)), with increase in MAb-1 concentration. Conversely, MAb-2 did not show such heterogeneous charge distribution as MAb-1 and hence favors intermolecular repulsion (positive B₂₂), lower static structure factor, S((q→0)), and repulsion induced increase in momentum transfer, H((q→0)), to result in lower viscosity of MAb-2. Charge swap mutants of MAb-1, M-5 and M-7, showed a decrease in charge asymmetry and concomitantly a loss in self-associating behavior and lower viscosity than MAb-1. However, replacement of charge residues in the sequence of MAb-2, M-10, did not invoke charge distribution to the same extent as MAb-1 and hence exhibited a similar viscosity and self-association profile as MAb-2.

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The rheological properties of macromolecular and colloidal suspensions are dependent on the thermodynamic and kinetic parameters that define viscous flow, and remain an active field of study with broad implications in cellular biophysics, soft-matter theory, and biopharmaceutical technology. Here we use static light scattering, small-angle X-ray scattering, and viscosity measurements as a function of protein concentration to semiquantitatively correlate the oligomeric state of an IgG1 antibody (mAb1) with its rheological behavior at solution pH 6.0 and varying ionic strength (modified by 0.01-0.1 M Na2SO4). Solution SAXS characterization of 100 mM Na2SO4 solutions confirmed that mAb1 forms reversible dimers with extended structures in dilute solutions. Light-scattering measurements over a wide range of concentrations (1-175 mg/mL) provide detailed information on the equilibrium thermodynamic mAb1 interactions and their modulation by modest increases of Na2SO4. Through the use of interacting hard sphere models to fit light-scattering data, we establish that protein cluster formations consisting of 2-9 mAb1 molecules also increase the viscosity of 175 mg/mL IgG solutions from 52 up to 450 cP. The analysis of dilute and semidilute mAb1 solution rheology correlates linearly with the thermodynamic equilibrium cluster size, consistent with the viscosity behavior of elongated oligomeric structures that are not significantly dendrimeric or in a state of globular collapse. Furthermore, SAXS- and rheology-based structural modeling illustrate that only a small set of anisotropic interactions between complementary surfaces are required to nucleate and propagate protein clusters.