Prometheus Panta for protein stability profiling with particle sizing and molecular weight determination by NanoTemper Technologies

Features:

• High-resolution information about your protein when you measure thermal unfolding, particle sizing, self-association, and aggregation simultaneously, throughout the entire thermal ramp
• Flexible throughput, with several options for sample handling
• Straightforward experimental design, with minimal sample preparation requirements and intuitive software user interface for both running experiments and data analysis

Parameters that Prometheus Panta measures:

Dynamic Light Scattering (DLS)

SIZE ANALYSIS

T_size (from growth of r_H in cumulants fit): Temperature at which average particle size begins to increase due to unfolding

Average scattering intensity: Identifies if sample concentration is too high or low for proper size distribution analysis

CUMULANT OR SIZE DISTRIBUTION/REGULARIZATION ANALYSIS

r_H: Hydrodynamic radius shows size of particle in its solvated state

PDI: Polydispersity index represents distribution of size populations within a sample, and is used to find impurities or poorly folded protein preparations

SELF-INTERACTION ANALYSES

k_D: The self-association parameter is used as an indication of a protein’s propensity to aggregate at higher concentrations in a given buffer environment

D₀ (*derived from k_D analysis): Theoretical diffusion constant at concentration = 0

nanoDSF

THERMAL UNFOLDING

T_m (for 330 nm, 350 nm, and ratio): Melting temperature, the point at which 50% of a protein is unfolded; higher melting temperatures indicate greater thermal stability

T_onset/T_on (for ratio): Temperature at which unfolding begins; higher T_on values indicate increased thermal stability, and it is considered favorable to have a sharper slope at the unfolding inflection point, which occurs when the T_on is close in value to the T_m

E_a (*derived from thermal unfolding data): Activation energy of unfolding, a parameter that informs on the stability of a protein in a given buffer environment

Reversibility of unfolding: Point at which aggregated protein cannot be recovered and refolded without other chemical means

Backreflection

AGGREGATION

T_turbidity: Temperature at which large amorphous aggregates (>12.5 nm radius) begin to form in solution

SLS

SIZING INFORMATION

Molecular weight: Average molecular weight of all particles in solution

SELF-ASSOCIATION AND AGGREGATION PROPENSITY

T_scattering: Temperature at which scattering intensity begins to change, indicating increased size and formation of smaller aggregates

B₂₂: Second virial coefficient ,used to extrapolate self-association behavior at higher concentrations