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All Journal Dental Journal (Majalah Kedokteran Gigi) Folia Medica Indonesiana
Yasan, Yanna Debby Restifanny
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Biochemistry, Genetics & Molecular Biology Dentistry Health Professions Medicine & Pharmacology Neuroscience

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Optimization of proteinase K incubation protocol duration during DNA extraction from oral squamous cell carcinoma FFPE samples Meizarini, Asti; Puteri, Astari; Yasan, Yanna Debby Restifanny; Hussaini, Haizal Mohd
Dental Journal (Majalah Kedokteran Gigi) Vol. 56 No. 4 (2023): December
Publisher : Faculty of Dental Medicine, Universitas Airlangga https://fkg.unair.ac.id/en

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.20473/j.djmkg.v56.i4.p233-237

Abstract

Background: Formalin-fixed paraffin-embedded (FFPE) specimen archives are a valuable source of sample material for molecular biological analysis. However, the DNA isolated from FFPE samples is usually low in concentration and fragmented. Thus, it is necessary to optimize the FFPE DNA extraction protocol to obtain the best results. Proteinase K incubation is undoubtedly crucial in DNA extraction procedures, but this step is often not well explained in the manufacturer's manual. Purpose: This study aimed to find the optimal duration for proteinase K incubation protocols to achieve the highest DNA yields. Methods: Fifteen paraffin blocks of Oral Squamous Cell Carcinoma (OSCC) specimens were obtained, and the cancerous areas were microdissected into smaller cuts for DNA extraction. The samples were randomly divided into three groups (n=5) and subjected to three different proteinase K incubation protocols: one-hour incubation at 56ºC as per the manufacturer's instructions (Group I), 24-hour incubation at 56ºC (Group II), and 48 hours at room temperature with an additional four hours at 56ºC (Group III). The extracted DNA was then quantified using a Nanodrop spectrophotometer. The recorded data were analyzed using ANOVA-LSD. Results: The highest DNA concentration was found in Group III (107.74 ± 41.92), which was significantly higher compared to Group II (59.46 ± 30.32) and Group I (6.46 ± 1.97) (p<0.05). Conclusion: In conclusion, modifying the duration of proteinase K incubation protocols can lead to different DNA yield results. In this study, the most optimized protocol for proteinase K incubation, resulting in the highest DNA yields, was 48 hours at room temperature with an additional four hours at 56ºC.
PRELIMINARY LABORATORY EVALUATION OF RAPID DIAGNOSTIC TEST PROTOTYPE TO DETECT HBsAg IN WHOLE BLOOD AND SERUM Qurnianingsih, Ema; Wungu, Citrawati Dyah Kencono; Amin, Mochammad; Setyawati, Dewi; Yasan, Yanna Debby Restifanny; Lukitasari, Lina; Lusida, Maria Inge; Soetjipto, Soetjipto
Folia Medica Indonesiana Vol. 61, No. 2
Publisher : Folia Medica Indonesiana

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Abstract

Chronic hepatitis B virus (HBV) remains a major global health issue,[-10pc]AU: Please provide ORCID ID for all authors.[-7pc]AU: Please provide affiliation’s for all authors. with over 250 million people affected worldwide. In Indonesia, HBV prevalence has declined from high to moderate endemicity, but challenges persist due to regional disparities, risk of mother-to-child transmission, and the predominance of B3 and C1 sub-genotype. Despite available preventive measures and diagnostic tools, approximately 90% of cases remain undiagnosed, particularly in low-resource settings, indicating the need for improved screening and control strategies. Therefore, this study aims to conduct preliminary performance tests of KODC hepatitis B surface antigen (HBsAg) (rapid diagnostic test (RDT) prototype) produced by Konimex on serum and whole blood samples. Performance was assessed through sensitivity, precision, specificity, and comparative testing against commercial RDT. The results obtained were visually interpreted by 2 independent readers. KODC HBsAg RDT demonstrated a detection limit of 50 IU/ml in serum and 400 IU/ml in whole blood. Precision testing showed high reproducibility (RSD <3.02%), with no cross-reactivity against other pathogens. Compared with a commercial RDT, it showed equivalent performance in serum and superior specificity in whole blood. In addition, predictive value analysis confirmed excellent accuracy, with Positive predictive value (PPV) and negative predictive value (NPV) >99% in both sample types. Therefore, KODC HBsAg RDT prototype showed comparable performance to commercial tests with good sensitivity, precision, and specificity, indicating its potential as a practical tool for HBV screening in resource-limited settings.
Co-Authors Amin, Mochammad Asti Meizarini Hussaini, Haizal Mohd Lukitasari, Lina Maria Inge Lusida Puteri, Astari Qurnianingsih, Ema Setyawati, Dewi SOETJIPTO . Wungu, Citrawati Dyah Kencono