= SEC light-scattering detection
{c}
<SEC light-scattering detection> adds information from <scattering of light> to the separation. Static scattering with a <concentration> readout can infer a <weight-average molar mass> for material in an eluting slice without the standard-family retention calibration. Dynamic scattering estimates a <diffusion coefficient> and hence a <hydrodynamic radius>. “Absolute” here means independent of a retention-to-<molar mass> standard curve; it still needs calibrated detectors, optical response parameters, and a valid sample model. Coeluting species produce a mixture-weighted result, rather than individually identified <molecules>.
A concrete manufacturer test used a Bio SEC-3 $300\,\mathrm{\mathring A}$ column, $7.8\times300\,\mathrm{mm}$ with $3\,\mathrm{\mu m}$ particles, an Agilent 1260 Infinity G5611A quaternary pump and G5667A autosampler, a $10\,\mathrm{mm}$ diode-array flow cell, and dual-angle/dynamic scattering detectors. A <buffer solution> at <pH> $7.4$ flowed at $0.75\,\mathrm{mL\,min^{-1}}$; column and scattering detector were at $30\,{}^\circ\mathrm C$. <Absorbance> at $280\,\mathrm{nm}$ supplied a concentration-sensitive readout. <Dilution> and loading comparisons tested response: seventeen ovalbumin injections spanning $2$–$960\,\mathrm{\mu g}$ gave a reported average $47{,}198\,\mathrm{g\,mol^{-1}}$ with $2\%$ relative standard deviation. The note reports poorer separation of thyroglobulin from aggregates and an elevated mass estimate, illustrating the coelution limit. Its $0.2\,\mathrm{\mu m}$ sample filtration also limits interpretation of larger <protein aggregation> products removed before injection.<Determination of Protein Molecular Weight and Size Using the Agilent 1260 Infinity Multi-Detector Bio-SEC Solution with Advanced Light Scattering Detection by Sonja Schneider>[Sonja Schneider, Determination of Protein Molecular Weight and Size Using the Agilent 1260 Infinity Multi-Detector Bio-SEC Solution with Advanced Light Scattering Detection, 5991-3955EN, printed pp. 2–4: apparatus, Table 1, Figure 2 and Table 3]{ref}
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