Validacija metoda genotipizacije polimorfizama u genima za faktor zgrušavanja V, protrombin, inhibitor aktivatora plazminogena-1 i metilentetrahidrofolat reduktazu

master's thesis

• Title (english): Validation of Methods for Genotyping Polymorphisms in Factor V, Prothrombin, Plasminogen Activator Inhibitor-1, and Methylenetetrahydrofolate Reductase Genes
• Author: Janko Bubnjar
• Mentor: Sandra Šupraha Goreta (mentor)
• Mentor: Mario Štefanović (sumentor)
• Committee member: Sandra Šupraha-Goreta (predsjednik povjerenstva)
• Committee member: Ivana Ćelap (član povjerenstva)
• Committee member: Mario Štefanović (član povjerenstva)
• Granter: University of Zagreb; Faculty of Pharmacy and Biochemistry; (Department of biochemistry and molecular biology); Zagreb
• Defense date and country: 2023-06-30, Croatia
• Scientific / art field, discipline and subdiscipline: BIOMEDICINE AND HEALTHCARE; Pharmacy; Medical Biochemistry

Abstract

Venski tromboembolizam je potencijalno po život opasno stanje čija etiologija uključuje brojne stečene i
nasljedne rizične čimbenike. Nasljedni rizični čimbenici uključuju genske polimorfizme faktor V Leiden (F5
G1619A) i protrombin G20210A (F2 G20210A). Katkad se u rizične faktore za venski tromboembolizam
ubrajaju i genski polimorfizmi PAI-1 5G/4G i MTHFR C677T iako se oni ne mogu jednoznačno povezati s
povećanim rizikom venskog tromboembolizma. U Kliničkom zavodu za kemiju Kliničkog bolničkog centra
Sestre milosrdnice došlo je do promjene analitičkog sustava za genotipizaciju navedenih polimorfizama te je
uveden LightCycler® 480 (Roche Diagnostics GmbH, Švicarska). Cilj ovog rada bio je validirati analitičke
karakteristike mjernih postupaka genotipizacije F5 G1619A, F2 G20210A, PAI-1 5G/4G i MTHFR C677T
metodom lančane reakcije polimeraze uz analizu krivulje taljenja na sustavu LightCycler® 480 prije
korištenja u rutinskom radu. Ispitani su analitička nepreciznost, analitička osjetljivost, analitička specifičnost,
analitička točnost, usporedivost drugom metodom i granica detekcije. Svi ispitani parametri u potpunosti su
zadovoljili unaprijed definirane analitičke kriterije kvalitete i stoga se ispitani mjerni postupci mogu prihvatiti
za rutinski rad.

Abstract (english)

Venous thromboembolism is a potentially life-threatening condition whose etiology includes numerous
acquired and hereditary risk factors. Hereditary risk factors include gene polymorphisms factor V Leiden
(F5 G1619A) and prothrombin G20210A (F2 G20210A). Gene polymorphisms PAI-1 5G/4G and MTHFR
C677T are also sometimes considered a risk factor for venous thromboembolism although those can not
be consistently associated with increased risk of venous thromboembolism. In the Clinical Chemistry
Department of the Sestre milosrdnice University Hospital Center, the analytical system being used for
genotyping mentioned polymorphisms was changed by introducing LightCycler® 480 (Roche Diagnostics
GmbH, Switzerland). This study aimed to evaluate analytical characteristics of genotyping
polymorphisms F5 G1619A, F2 G20210A, PAI-1 5G/4G, and MTHFR C677T by real-time polymerase
chain reaction with melting curve analysis on LightCycler® 480 before putting it into routine operation.
Analytical imprecision, analytical sensitivity, analytical specificity, analytical accuracy, repeatability
using another method, and limit of detection were examined. All examined parameters fully met the predefined analytical quality criteria and therefore the evaluated methods can be accepted for routine work.

• Keywords: trombofilija; genotipizacija; validacija
• Keywords (english): thrombophilia; genotyping; validation
• Language: croatian
• URN:NBN: urn:nbn:hr:163:822136
• Study programme: Title: Medical biochemistry; Study programme type: university; Study level: integrated undergraduate and graduate; Academic / professional title: magistar/magistra medicinske biokemije (magistar/magistra medicinske biokemije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2023-11-21 15:12:44