Sensitive Detection of Immunoglobulin G Stability Using in Real-Time Isothermal Differential Scanning Fluorimetry: Determinants of Protein Stability for Antibody-Based Therapeutics
Why this work is in the frame
A frame that forgets how it found something cannot be audited. These are the routes that admitted this work.
Bibliographic record
Abstract
Protein instability is a major obstacle in the production and delivery of monoclonal antibody–based therapies for cancer. This study presents real-time isothermal differential scanning fluorimetry as an emerging method to evaluate the stability of human immunoglobulin G protein with high sensitivity. The stability of polyclonal human immunoglobulin G against urea-induced denaturation was assessed following: (1) oxidation by the free-radical generator 2,2-Azobis[2-amidinopropane]dihydrochloride and (2) in selected storage buffers. Significant differences in immunoglobulin G stability were detected by real-time isothermal differential scanning fluorimetry when the immunoglobulin G was stored in 1,4-Piperazinediethanesulfonic acid buffer compared to phosphate-buffered saline, with half-maximal rate of denaturation occurring at a higher urea concentration in 1,4-Piperazinediethanesulfonic acid than phosphate-buffered saline ( K nd;PIPES = 3.56 ± 0.09 M, K nd;PBS = 2.94 ± 0.08 M; P < .01), but differential scanning fluorimetry did not detect differences in unfolding temperature ( T m;PIPES = 70.5 ± 0.3°C, T m;PBS = 69.7 ± 0.2°C). The effects of 2,2-Azobis[2-amidinopropane]dihydrochloride-induced oxidation on immunoglobulin G stability were analyzed by real-time isothermal differential scanning fluorimetry; the oxidized protein showed greater sensitivity to urea ( K nd;CNTRL = 3.96 ± 0.19 M, K nd;AAPH = 3.49 ± 0.07 M; P < .05). Similarly, differential scanning fluorimetry indicated greater thermal sensitivity of oxidized immunoglobulin G ( T m;CNTRL = 70.5 ± 0.3°C, T m;AAPH = 62.9 ± 0.1°C; P < .001). However, a third method for assessing protein stability, pulse proteolysis, proved to be substantially less sensitive and did not detect significant effects of 2,2-Azobis[2-amidinopropane]dihydrochloride on the half-maximal concentration of urea needed to denature immunoglobulin G ( C m;CNTRL = 6.8 ± 0.1 M; C m;AAPH = 6.4 ± 0.7 M). Overall these results demonstrate the merit of using real-time isothermal differential scanning fluorimetry as a rapid and sensitive technique for the evaluation of protein stability in solution using a quantitative real-time thermocycler.
Fetched live from OpenAlex and de-inverted. Abstracts are not stored in this database: the inverted indexes are 8.6 GB of the frame’s 9.3 GB of text, and the host has 13 GB free.
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.001 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.001 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 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 it