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GenExpert in detection of genetic polymorphisms for FV G1691A and FII G20201A: two years experience in a high prevalence area

Genexpert nella rivelazione dei polimorfismi genici per FV G1961A e FII G20210A: due anni di esperienza in un’area a elevata prevalenza

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La Rivista Italiana della Medicina di Laboratorio - Italian Journal of Laboratory Medicine

Summary

Background.

FVG1691A (factor V Leiden, FVL) and FII G20210A (prothrombin G, GPRO) polymorphisms are the most common genetic causes of thrombophilia. For diagnosis of these polymorphisms, two years ago we adopted the GeneXpert instrument, after a preliminary study conducted on frozen plasma that had already been typed. We report our experience concerning the routine application of this analysis system.

Methods.

Between January 2011 and December 2012, 1106 patients were evaluated for detection of FVL and GPRO using the Cepheid GeneXpert system (Instrumentation Laboratory, Milan, Italy). The first 142 were also tested with the LightCycler system (Roche, Monza, MI, Italy).

Results.

Results obtained with the GeneXpert system showed full agreement with those obtained with the LightCycler system. Full agreement was also observed with previously characterized frozen samples. Among the 1,106 subjects examined, 235 were FVL heterozygous, 20 homozygous FVL, 37 GPRO heterozygous, 3 homozygous GPRO, 15 double heterozygous FVL/GPRO, and 796 genetically normal. In the 2-year period we observed only 37 invalid results with the need to repeat the test.

Conclusions.

The GeneXpert system is a fully automated analytical system. In less than 35 minutes, it is possible to perform a combined determination of FVL and GPRO from a subject’s anticoagulated whole blood. The design of the kit, based on the use of individual disposable cartridges, in which are contained all the necessary reagents for the extraction of nucleic acids, their amplification and the detection of the amplicons, eliminates waste and allows determinations to be performed on demand. On the other hand, the extremely simple manual makes the test accessible to those laboratories not specializing in molecular biology techniques. In our experience, the GeneXpert system has proven reliable with total concordance with the results obtained with the system already in use in our laboratory. We also observed only a small percentage of invalid results with a satisfactory ratio (1.03) of the number of tests performed to the number producing a result.

Riassunto

Premesse.

I polimorfismi FVG1691A (Fattore V di Leiden: FVL) e FII G20210A (protrombina G, GPro) sono le più comuni cause genetiche di trombofilia. Per la diagnosi di questi polimorfismi abbiamo adottato, due anni orsono, dopo uno studio preliminare condotto su plasmi congelati già tipizzati, la strumentazione GeneXpert. Riportiamo quindi la nostra esperienza relativa all’applicazione in routine di tale analizzatore.

Metodi.

Tra gennaio 2011 e dicembre 2012 abbiamo valutato 1106 pazienti per la ricerca del FVL e della GPro utilizzando il sistema Cepheid GeneXpert (Instrumentation Laboratory, Milano, Italia). I primi 142 sono stati testati in duplicato con il sistema LightCycler (Roche, Monza, MI, Italia).

Risultati.

I risultati ottenuti con il sistema GeneXpert hanno dimostrato una piena concordanza con quelli ottenuti con l’analizzatore LightCycler; inoltre, ugualmente totale è stata la concordanza nei campioni congelati già tipizzati. Tra i 1106 soggetti esaminati abbiamo osservato 235 FVL eterozigoti, 20 FVL omozigoti, 37 GPro eterozigoti, 3 GPro omozigoti, 15 soggetti con doppia eterozigosi FVL/GPro, 796 soggetti normali. Nei due anni considerati abbiamo osservato solamente 37 risultati non validi con necessità di ripetere il test.

Conclusioni.

Il sistema GeneXpert è un sistema analitico completamente automatico che, in meno di 35 minuti, è in grado, partendo dal sangue intero anticoagulato del soggetto da esaminare, di effettuare la ricerca contemporanea del FVL e della GPro. La progettazione del kit, basato sull’utilizzo di singole cartucce a perdere, in cui sono contenuti tutti i reagenti necessari per l’estrazione degli acidi nucleici, la loro amplificazione, la rilevazione degli amplificati, annulla gli sprechi e permette di eseguire le determinazioni on demand. D’altro canto, l’estrema semplificazione della manualità necessaria rende accessibili questi test anche a Laboratori non specializzati in tecniche di Biologia Molecolare. Nella nostra esperienza il sistema GeneXpert si è dimostrato affidabile con una totale concordanza con i risultati ottenuti con il sistema precedentemente in uso presso il nostro Laboratorio. Inoltre, abbiamo osservato una ridotta percentuale di risultati non validi con una ratio fra test utilizzati e risultati prodotti assai soddisfacente (1.03).

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Authors and Affiliations

  1. Servizio di Medicina di Laboratorio, Ospedale Madonna della Navicella, Strada Madonna Marina 500, 30015, Chioggia, VE, Italy

    Sara Valverde, Ernesto Trabuio, Francesco Antico, Francesca Marangon & Gianluca Gessoni

  2. Servizio Trasfusionale, Ospedale Madonna della Navicella, Chioggia, Italy

    Maria Monica Salvadego, Samanta Beggio, Maria Grazia Piva & Gianluca Gessoni

Authors
  1. Sara Valverde
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  2. Ernesto Trabuio
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  3. Francesco Antico
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  4. Francesca Marangon
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  5. Maria Monica Salvadego
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  6. Samanta Beggio
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  7. Maria Grazia Piva
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  8. Gianluca Gessoni
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Corresponding author

Correspondence to Gianluca Gessoni.

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Valverde, S., Trabuio, E., Antico, F. et al. GenExpert in detection of genetic polymorphisms for FV G1691A and FII G20201A: two years experience in a high prevalence area. Riv Ital Med Lab 9, 143–148 (2013). https://doi.org/10.1007/s13631-013-0005-3

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  • DOI: https://doi.org/10.1007/s13631-013-0005-3

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