Genotyping of hepatitis B virus from dried and stored serum on filter paper

Abstract

AbstrakGenotipe virus hepatitis B (VHB) mempunyai hubungan yang erat dengan prognosis dan terapinya serta diperlukan untuk studi epidemiologi. Pemeriksaan ini hanya bisa dikerjakan di kota-kota besar saja karena kesulitan pengiriman sampel akibat masalah geografis maupun fasilitas. Tujuan penelitian ini adalah untuk mengetahui apakah genotipe VHB dapat ditentukan dari serum kering pada kertas saring dan membandingkan hasil tersebut dengan serum yang diambil langsung dari pasien hepatitis B kronik (HBK) dan hepatoma. Dua puluh tiga sampel dapat diambil dari pasien HBK dan konsentrasi DNA VHB di tentukan dengan Cobas Amplicor HBM (Roche Diagnostics GmBH, Germany) kemudian diteteskan pada kertas saring (3 x 1 cm). Setelah dikeringkan dalam kantong plastik, diletakkan dalam amplop tertutup dan disimpan selama 1 minggu dalam suhu kamar (27 -33 o C). Ekstraksi DNA dilakukan dari kertas saring tersebut setelah diinkubasi dan penentuan genotipe VHB dilakukan dengan PCR menggunakan primer-primer spesifik. Untuk perbandingan, telah didapatkan 20 sampel pasien HBK-HBe (+) dan 29 sampel pasien hepatoma yang tidak dikeringkan. Genotipe VHB dapat dideteksi pada 18/23 (78,2%) serum kering pada kertas saring sedangkan pada serum yang tidak dikeringkan, dari pasien HBK-HBe(+) 20/20 (100%) sampel terdeteksi dan dari pasien hepatoma 24/29 (82,7%) sampel. Proporsi genotipe yang terdeteksi sesuai dengan proporsi genotipe yang pernah dilaporkan di Indonesia. Kesimpulan penelitian ini adalah genotipe VHB dapat dideteksi dari serum kering pada kertas saring yang disimpan selama 1 minggu. (Med J Indones 2005; 14: 215-9)Abstract HBV genotype has a close association with prognosis and therapy as well as for epidemiology study. However, this examination can be done only in large cities that are not practical to send serum sample due to geographical burden and facilities. The aim of this study is to know whether HBV genotype can be determined from dried and stored serum on filter paper and compare the result with sera drawn directly from chronic hepatitis B (CHB) and hepatoma patients. Twenty-three serum samples were obtained from CHB patients. HBV DNA were quantitatively determined with Cobas Amplicor HBM (Roche Diagnostics GmBH, Germany) and dropped on to 3 x 1 cm filter papers. After allowed to dry in a plastic clip, it were put in a closed envelope then stored for 1 week in room condition (27 -33 o C). DNA extraction were done from the filter papers after a short incubation period and HBV genotypes were determined with PCR and specific primers. For comparison, samples and 29 hepatoma samples were drawn directly and not dried. HBV genotype were detected in 18/23 (78.2%) from dried serum samples on filter paper while in sera that were not stored, from CHB-HBe(+) samples, 20/20 (100%) could be determined while from hepatoma patients, 24/29 (82.7%) samples. The proportion of genotype were in line with other reported HBV genotype examination for Indonesia. It is concluded that detection of HBV genotype can be done from dried serum in filter p...

HBV carriage in children born from HIV‐seropositive mothers in Senegal: The need of birth‐dose HBV vaccination

“…In this study, DBS were used as blood support for HIV EID due to its usefulness that confer DNA stability on the filter paper allowing samples transportation from far healthcare centers to reference laboratories. For HBV diagnosis, its use is also increasingly described as a new tool for epidemiological research of HBV markers [Gani et al, ; Kania et al, ; Mohamed et al, ]. Studies had first evaluated the use of DBS by comparison to serum using a radioimmunoassay method [Villa et al, ].…”