Unchained Labs eBook - 13

Unleash your biologic * Protein characterization turned up to 11

at 4 °C, three of the molecules, mAb 1, 2, and
4, yielded measurements of the expected
size, with a diameter of 10-11 nm. Their polydispersity index (PDI) values were also below
0.25, which indicates a monodisperse population. In contrast, mAb 3 was measured to
have a diameter of 175 nm and a PDI of
almost 1.4, suggesting significant aggregate
content. Obtaining this information early
allows a researcher to eliminate this
particular formulation for this mAb from
further consideration.

Simultaneous kD and B22
measurements
After ranking and refining samples to a set of
thermally stable molecules or formulations,
Uncle can aid in reducing the risk of aggregation
when moving candidates to high concentration conditions. Weak, non-specific interactions play important roles in solubility and aggregation in a crowded protein solution. The
diffusion interaction parameter (kD) and the
second virial coefficient (B22) are both used
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for predicting the colloidal stability and
aggregation propensity of proteins. These
measurements may be used to confirm
stability and minimize the risk of the biologic
interacting with itself.
Negative k D and B22 values are interpreted to
mean that the protein favors self-association
over complete solvation. Positive values
indicate repulsive interactions between
protein molecules, or more favorable
protein-solvent interactions. Zero or nearzero values indicate neutral interactions.
Both parameters are obtained simultaneously
on Uncle with one sample set. Each experiment
involves preparing a sample (a specific
protein in a given formulation) across a
range of concentrations. For k D values,
Uncle software uses the hydrodynamic
radius measured by DLS for each sample
across the concentration series to calculate
diffusion coefficients. To obtain the B 22
values, the derived light-scattering intensity
of each sample is calibrated by the scattering
intensity of a known intensity reference.

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Unchained Labs eBook

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