An Intact ACTH LC-MS/MS Assay as an Arbiter of Clinically Discordant Immunoassay Results
Bibliographic record
Abstract
BACKGROUND: Measurement of plasma adrenocorticotropic hormone (ACTH) is key in the differential diagnosis of hypothalamic-pituitary-adrenal disorders. Two-site sandwich immunoassays dominate clinical testing of ACTH in North America; however, discordant results between manufacturers have been repeatedly reported. To resolve the discrepancy, we developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for the intended measurand, biologically active intact ACTH (iACTH). METHODS: ). Epitope assignment of the Roche Elecsys antibodies was performed by MALDI-TOF mass spectrometry. A method comparison between Roche Elecsys and Siemens Immulite ACTH immunoassays was performed and clinically concordant/discordant results identified. In a subset of these samples, the iACTH concentration was determined using the LC-MS/MS method. RESULTS: The lower limit of the measuring interval of the iACTH LC-MS/MS assay was 9 pg/mL (2 pmol/L). The assay was linear from 9 to 1938 pg/mL (2 to 427 pmol/L). Epitope mapping revealed that the Roche capture and detection antibodies bound residues 9-12 and 36-39 of ACTH, respectively. The iACTH LC-MS/MS analysis demonstrated that for discordant results between 2 immunoassays studied, only the Roche results were highly positively correlated with the iACTH concentration. CONCLUSIONS: Immunoprecipitation of biologically active ACTH molecules followed by LC-MS/MS analysis enabled selective detection of iACTH and relevant biologically active fragments in plasma. Applied to the investigation of clinically discrepant results, this method can act as an arbiter of the concentration of iACTH present.
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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 | 0.007 | 0.011 |
| Meta-epidemiology (narrow) | 0.002 | 0.000 |
| Meta-epidemiology (broad) | 0.001 | 0.000 |
| Bibliometrics | 0.003 | 0.001 |
| Science and technology studies | 0.001 | 0.001 |
| Scholarly communication | 0.002 | 0.000 |
| Open science | 0.001 | 0.001 |
| Research integrity | 0.002 | 0.001 |
| Insufficient payload (model declined to judge) | 0.001 | 0.001 |
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".