Nucleic acid based lateral flow test (LFT) for COVID-19 diagnosis
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2021
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info:eu-repo/semantics/openAccess
Özet
Dünya Sağlık Örgütü (DSÖ) tarafından resmi adı SARS-CoV-2 (Şiddetli Akut Solunum Sendromu-Koronavirus-2) olarak belirlenen yeni koronavirüs, ilk kez 2019 yılı Aralık ayı sonlarında Çin'in Wuhan şehrindeki deniz ürünleri pazarında ortaya çıktığı düşünülmektedir. Virüs oldukça bulaşıcı ve özellikle ileri yaş gruplarında ölümcül olmaktadır. Viral teşhis için birçok çalışma geliştirilmeye devam etmektedir. En yaygın ve güvenilir olanı DSÖ tarafından önerilen RT PCR (Real Time Polymerase Chain Reaction) olup özellikle N geni bölgelerine yöneliktir. Bu bölge en korunaklı ve spesifik bölge olduğu için moleküler teşhiste önemli olmuştur. Bununla birlikte hızlı testler laboratuvar yüklerinin hafifletilmesinde, antikor gelişimini kontrol etmede önemli yere sahiptir. Hızlı testler arasında ise yatay akışlı testler en yaygın ve pratik kullanımı olandır. Uzun vadede viral bulaşıklık ihtimali hep olacaktır ve pratik kullanımlı teşhis testlerine ihtiyaç devam edecektir. Bu amaçla çalışmamızda SARS-CoV-2'ye özgü N geni bölgelerinin yatay akışlı hızlı test platformu ile teşhisi hedeflenmiştir. Yatay akışlı test platformu Altın nanoparçacıklara (AuNP) dayalı sandviç modeli ile hibridizasyon temeline dayalı olarak tasarlanmıştır. Tasarlanan LFT platformu N geninin korunmuş bölgelerine özgü dizayn edilmiştir. N genine tamamlayıcı olan DNA oligonükleotitleri ile AuNP konjuge edilmiştir. Bu konjugasyon ürününe hibridizasyon oluşturacak diziler nitroselüloz membran üzerine test ve kontrol çizgisi olarak çizilmiştir. Hedef sekansı içeren örnek, numune pedine uygulandığında kapiller akım prensibine dayanarak hedef sekans test ve kontrol çizgisinde bulunan diziler ile hibridizasyon yapar. AuNP'den kaynaklı bir kırmızı renk oluşumu çıplak gözle görülebilir. Araştırma bulgularına göre, tasarlanan LFT, SARS-CoV-2 N geni bölgelerini spesifik olarak tanımış ve moleküler teşhise alternatif olabileceği ortaya konmuştur.
The new coronavirus, official name was determined by the WHO as SARS-CoV-2, is thought to have first appeared in the seafood market in Wuhan, China, in late December 2019. The virus is highly contagious and fatal, especially older age groups. Many studies continue to be developed for viral diagnosis. The most common and reliable one is the RT PCR recommended by WHO, and it is especially aimed at the N gene regions. This region has been important in molecular diagnosis as it is the most conserved and specific region. However, rapid tests have an important place in relieving laboratory and controlling antibody development. Among rapid tests, LFTs are the most common and practical. In long time, there will always be the possibility of viral contamination and the need for practical diagnostic tests will continue. For this purpose, our study aimed to identify the N gene regions specific to SARS-CoV-2 with a LFT platform. The LFT platform is designed on the basis of hybridization with sandwich model based on Gold nanoparticles (AuNP). The designed LFT platform was designed specifically for the conserved regions of the N gene. AuNP was conjugated with DNA oligonucleotides complementary to N gene. Sequences that will hybridize to this conjugation product were drawn on the nitrocellulose membrane as test lines. The sample containing the target sequence hybridizes with the sequences in the test lines based on the capillary flow principle when applied to the sample pad. A red color formation from AuNP can be seen with the naked eye. According to the research findings, the designed LFT specifically recognized the SARS-CoV-2 N gene regions and it was revealed that it could be an alternative to molecular diagnosis.
The new coronavirus, official name was determined by the WHO as SARS-CoV-2, is thought to have first appeared in the seafood market in Wuhan, China, in late December 2019. The virus is highly contagious and fatal, especially older age groups. Many studies continue to be developed for viral diagnosis. The most common and reliable one is the RT PCR recommended by WHO, and it is especially aimed at the N gene regions. This region has been important in molecular diagnosis as it is the most conserved and specific region. However, rapid tests have an important place in relieving laboratory and controlling antibody development. Among rapid tests, LFTs are the most common and practical. In long time, there will always be the possibility of viral contamination and the need for practical diagnostic tests will continue. For this purpose, our study aimed to identify the N gene regions specific to SARS-CoV-2 with a LFT platform. The LFT platform is designed on the basis of hybridization with sandwich model based on Gold nanoparticles (AuNP). The designed LFT platform was designed specifically for the conserved regions of the N gene. AuNP was conjugated with DNA oligonucleotides complementary to N gene. Sequences that will hybridize to this conjugation product were drawn on the nitrocellulose membrane as test lines. The sample containing the target sequence hybridizes with the sequences in the test lines based on the capillary flow principle when applied to the sample pad. A red color formation from AuNP can be seen with the naked eye. According to the research findings, the designed LFT specifically recognized the SARS-CoV-2 N gene regions and it was revealed that it could be an alternative to molecular diagnosis.
Açıklama
Anahtar Kelimeler
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İnönü Üniversitesi Fen Bilimleri Enstitüsü
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Künye
GÜLTEKİN, E. (2021). Nucleic acid based lateral flow test (LFT) for COVID-19 diagnosis
Yüksek Lisans Tezi, İnönü Üniversitesi.
