Selective protein quantification on continuous chromatography equipment with limited absorbance sensing: A partial least squares and statistical wavelength selection solution
Bibliographic record
Abstract
Abstract Real‐time selective protein quantification is an integral component of operating continuous chromatography processes. Partial least squares models fit with spectroscopic UV‐Vis absorbance data have demonstrated the ability to selectively quantify proteins. With standard continuous chromatography equipment that is only capable of measuring absorbance at a few user‐defined wavelengths, the problem of selecting appropriate wavelengths that maximize the measurement capability of the instrument remains unaddressed. Therefore, we propose a method for selecting wavelengths for continuous chromatography equipment. We illustrate our method using sets of protein mixtures composed of bovine serum albumin and lysozyme. The first step is to refine the raw wavelength set with a statistical t ‐test and an absorbance magnitude test. Then, the wavelengths within the refined spectroscopic range are ranked. Three existing techniques are evaluated – sequential forward search, variable importance to projection scores, and the least absolute shrinkage and selection operator. The best technique (in this case, sequential forward search) determines a subset of three wavelengths for further evaluation on the BioSMB PD. We use an exhaustive approach to determine the final wavelength set. We show that soft sensor models trained from the method's wavelength selections can quantify the two proteins more accurately than from the wavelength set of 230, 260 and 280 nm, by a factor of four. The method is shown to determine appropriate wavelengths for different path lengths and protein concentration ranges. Overall, we provide a tool that alleviates the analytical bottleneck for practitioners seeking to develop advanced monitoring and control methods on standard equipment.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.002 | 0.003 |
| Meta-epidemiology (narrow) | 0.001 | 0.000 |
| Meta-epidemiology (broad) | 0.001 | 0.001 |
| Bibliometrics | 0.001 | 0.001 |
| Science and technology studies | 0.000 | 0.001 |
| Scholarly communication | 0.001 | 0.001 |
| Open science | 0.001 | 0.001 |
| Research integrity | 0.001 | 0.001 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".