P050 Centrally-determined standardization of flow cytometry methods reduces inter-laboratory variation in a prospective multicenter study
Notice bibliographique
Résumé
Background: Flow cytometry (FC) of mucosal biopsy and peripheral blood samples from patients with inflammatory bowel disease aids in characterization of cellular and molecular factors involved in the pathologic immune response in these diseases. This technique has potential to facilitate early drug development and elucidate mechanisms of action of prospective therapies. Lack of standardized methods and variation in FC outcomes across laboratories hamper its use in multicenter clinical trials. We compared the variation in 3 FC strategies among international laboratories. Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from buffy coats from 3 healthy volunteers, cultured in a cocktail +/− phorbol 12-myristate 13-acetate and ionomycin at a central laboratory, and then fixed, frozen, and shipped on dry ice to 7 international laboratories. Permeabilization and staining of PBMCs was performed at each laboratory in triplicate using a common protocol and centrally-provided reagents. Gating was performed according to 3 strategies: local gating with a local strategy, local gating with a central strategy, and central gating. A range of cell populations, with high or low event numbers and in stimulated and unstimulated conditions was chosen for analyses. Mean cell proportion was calculated across triplicates and within donors, conditions and strategies. The coefficient of variation (CV) for each FC parameter was calculated across laboratories. Among-strategy comparisons were made using a two-way ANOVA, adjusting for donor. Results: Mean inter-laboratory CV ranged from 2.1%–74.1% depending on cell population and gating strategy (5.1%–74.1% for local gating with a local strategy, 10.9%–65.6% for local gating with a central strategy, and 2.1%–20.9% for central gating [Table 1]). For each FC parameter, mean-inter laboratory CV differed significantly across gating strategies and variability was consistently lower with central gating, which reduced mean inter-laboratory CV by 3%-67%, depending on cell population. Conclusions: Flow cytometry can be performed by multiple international laboratories with reasonable precision using a common protocol for permeabilization and staining, and centrally-performed gating. Central gating was the only strategy with mean CVs consistently lower than 25%; a proposed standard for pharmacodynamic and exploratory biomarker assays [1]. Our results suggest that gating is a major source of variability in FC. References: [1] O'Hara DM et al, (2011), Recommendations for the validation of flow cytometric testing during drug development: II assays, J Immunol Methods, 120
Récupéré en direct depuis OpenAlex et désinversé. Les résumés ne sont pas conservés dans cette base de données : les index inversés représentent 8,6 Go des 9,3 Go de texte de la base, et le serveur dispose de 13 Go libres.
Comment cette classification a été obtenuedéplier
Prédiction distillée sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Apprise à partir de 10 348 étiquettes directes de Codex et de 10 348 étiquettes directes de Gemma. Le mode candidate est l'union des têtes enseignantes seuillées; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont ni des étiquettes humaines ni des étiquettes directes de modèles de pointe.
Scores Codex et Gemma par catégorie
| Catégorie | Codex | Gemma |
|---|---|---|
| Métarecherche | 0,001 | 0,000 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,000 | 0,000 |
| Bibliométrie | 0,000 | 0,000 |
| Études des sciences et des technologies | 0,000 | 0,000 |
| Communication savante | 0,000 | 0,000 |
| Science ouverte | 0,000 | 0,000 |
| Intégrité de la recherche | 0,000 | 0,000 |
| Charge utile insuffisante (le modèle a refusé de juger) | 0,000 | 0,000 |
Scores machine (provisoires)
Les deux têtes enseignantes du modèle étudiant, lues sur ce travail. Un score ordonne la base pour la relecture; il n'affirme jamais une catégorie, et le statut de validation accompagne chaque rangée tel quel.
Scores de référence d'un modèle non mature (critères de maturité non atteints, 7 itérations). Un score ordonne; il n'affirme jamais une catégorie.
score_only:v0-immature-baseline · tel quel depuis la passe de notation : score_only signifie que le nombre peut ordonner les travaux, et qu'aucune étiquette de catégorie n'en découleClassification
machine, non validéePrédiction automatique; un appel candidat d’une seule tête enseignante, pas un consensus.
Le détail, modèle par modèle et score par score, se trouve en fin de page sous « Comment cette classification a été obtenue ».