Solvent correction for different applications

Solvent correction is only appropriate for work where the following conditions are met:

The ligand is a macromolecule immobilized at a high density on the surface (typically 7,000-8,000 RU or more, although solvent correction can sometimes be useful at lower ligand levels depending on the application)
The analyte is a small molecule giving low expected responses (R_max typically less than 50 RU, often as low as 10-20 RU)
Organic solvent that gives a high bulk refractive index contribution (usually DMSO) is included in the sample and running buffer
The bulk response is high in relation to the measured binding response and is subject to variations between sample solutions

Under these conditions, include solvent correction cycles in your assay. You can decide whether to apply solvent correction or not when you evaluate the data.

Solvent correction for small molecule screening and ranking

Screening and ranking applications for low molecular weight drug candidates involve analysis of a number (often a large number) of different analytes, usually diluted from stock solutions in DMSO. The exact DMSO concentration may vary slightly between samples, giving a variation in bulk refractive index contribution that may be of the same order of magnitude as the sample responses themselves. Solvent correction eliminates the bulk variations, allowing direct comparison of binding levels between different samples.

Small molecule screening data from Biacore A100 without (left) and with (right) solvent correction.

Always use solvent correction for screening and ranking of small molecules in DMSO-containing buffers.

Solvent correction for kinetics determination

Kinetic determinations are performed on carefully prepared dilution series of samples, so that the DMSO concentration should in principle not vary between samples. In practice, however, small pipetting errors may lead to random variations in DMSO concentration, while an error in preparation of a stock solution may propagate systematically through the dilution series.

Most kinetic evaluation procedures allow local variations in a bulk refractive index parameter which can take care of small variations in DMSO concentration, so that solvent correction has practically no effect on the reported kinetic constants. Evaluation is however visually clearer and may be more robust if solvent correction is used to eliminate the bulk variation.

Single-cycle kinetics sensorgrams with (green) and without (red) solvent correction

Evaluation of single-cycle kinetics without (left) and with (right) solvent correction. Bulk variations are assigned to the RI parameter, and solvent correction has no effect on the reported kinetic constants.

Include solvent correction cycles in kinetic assays.

Solvent correction for steady state affinity determination

Like kinetic determinations, affinity measurements are performed on carefully prepared sample dilution series. Unlike kinetics, however, steady state affinity is evaluated from response levels, not from the sensorgram shape. The measured response levels will be affected by any bulk contributions resulting from variations in DMSO concentration.

If all samples in a dilution series have identical bulk contributions, this will appear as an offset in the evaluation (the plot of R_eq against concentration will not pass through the origin) and the reported affinity constant will not be affected. Random or systematic variations in bulk contribution will however affect the evaluation of R_eq against concentration directly.

Evaluation of steady state affinity without (left) and with (right) solvent correction. In this example the bulk contribution was similar in all samples, so that solvent correction reduces the offset term but has little effect on the calculated affinity.

Include solvent correction cycles in affinity assays.

Solvent correction for concentration assays

Solvent correction is not relevant to concentration assays:

Calibrated concentration assays are normally evaluated using response levels placed shortly after the sample injection, where solvent correction does not apply.
Calibration-free concentration analysis (CFCA) is not suitable for analytes smaller than about 5,000 Da, and solvent correction is not appropriate for analytes of this size.

Do not use solvent correction with concentration measurements.

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