Analisis Perbandingan Kualitas Produk Amplikon Gen PMSA2 Antara Spesimen Spot Darah Kering dan Vena

Authors

  • Arsyam Mawardi Cenderawasih University
  • Hendra K. Maury Cenderawasih University
  • Yustinus Maladan Balitbangkes Papua

DOI:

https://doi.org/10.31957/jbp.949

Abstract

This study is aimed to analyze the comparative quality of PMSA2 gene amplicon product stability from two different specimen sources, spot specimens of dried blood and venous blood, as well as selecting the best storage method for specimens of blood samples. This research uses descriptive laboratory research methods. The research began with the process of sample preparation for dried blood spot and venous blood, each using Whatman 903 paper and vacuum tubes containing EDTA, isolating genomic DNA using KIT Zymo Research, amplification of PMSA-2 genes with PCR, detection of PCR products through electrophoresis, measurement of DNA concentration and absorbance, and data analysis. The results of this study are expected to be a source of information about the advantages of two specimen storage methods for clinical blood samples, as well as providing a clear description of the quality of each specimen storage method based on the quality of its amplicon products. The results showed that a total of ten medical samples of dried blood spot and ten venous blood were isolated from the genomic DNA of ten and nine, respectively. PMSA2 gene amplicons detected were seven in venous blood and six in dried blood spot. Venous blood specimens have sensitivity in detecting PMSA genes in samples with the highest value of 554 ng / μL and purity of 2,007 (WB7), and concentration of 550.2 and highest purity of 2,076 (WB10). Venous blood storage techniques using categorized vacuum tubes are effective in the detection of PMSA2 genes and have time efficiency in the process. From these results it was also concluded that the comparison analysis of amplicon products between venous blood specimens was better, more stable and efficient than dry blood spot specimens, thus recommending storage of venous blood specimens using vacuum tubes as the best storage method of blood sample specimens.

Downloads

Download data is not yet available.

Author Biographies

Arsyam Mawardi, Cenderawasih University

Department of Biology

Hendra K. Maury, Cenderawasih University

Department of Biology

References

Adda, C.G., MacRaild, C.A., Reiling, L., Wycherley, K., Boyle, M.J., dan Kienzle, V. 2012. Antigenic Characterization of an Intrinsically Unstructured Protein, Plasmodium falciparum Merozoite Surface Protein 2, Infection and Immunity, 80(12): 4177-4185.

Chambers A.G., Andrew J.P., Juncong Y., Alexander G.C., & Christoph H.B. 2013. Multiplexed Quantitation of Endogenous Proteins in Dried Blood Spots by Multiple Reaction Monitoring - Mass Spectrometry. The American Society for Biochemistry and Molecular Biology, 12 (3):781-791.

Corkill, G., R. Rapley. 2008. The Manipulation of Nucleic Acids: Basic Tools and Techniques. In: Molecular Biomethods Handbook Second Edition. Ed: Walker, J.M., Rapley, R. Humana Press, NJ, USA.

Deglon J., Aurelien T., Patrice M., & Christian S. 2012. Direct analysis of dried blood spots coupled with mass spectrometry:concepts and biomedical applications. Anal.Bioanal. Chem., 402:2485-2498.

Edwards R.L., Andrew J.C., Mark B., Paul G., Josephine B., & Helen J.C. 2011. Hemoglobin Variant Analysis via Direct Surface Sampling of Dried Blood Spots Coupled with High - Resolution Mass Spectrometry. Anal. Chem, 83:2265-2270.

Evans C., Mark A., Peter B., Jeffrey , Christopher A.J., Wenkui L., Steve L., Luca M., Timothy O., Philip T., Xiaomin W., Enaksha W., John W., Eric W., & Patricia Z. 2015. Implementing Dried Blood Spot Sampling for Clinical Pharmacokinetic Determinations: Considerations from the IQ Consortium Microsampling Working Group. The AAPS Journal, 17 (2): 292-300.

FDA (2001) Guidance for Industry Bioanalytical Method Validation.Food and Drug Administration United States of America: Department of Health and Human Services.

Khosravinia, H., H.N.N. Murthy, D.T. Parasad, & N. Pirany. 2007. Optimizing Factors Influencing DNA Extraction from Fresh Whole Avian Blood. African Journal of Biotechnology. 6(4): 481-486.

Liu G., Ji Q.C., Jemal M., Tymiak A.A. & Arnold M.E. 2011. Approach to evaluating dried blood spot sample stability during drying process and discovery of a treated card to maintain analyte stability by rapid on-card pH modification. Anal. Chem. 83: 9033.

Mawardi, A., dan M.L. Simonapendi. 2016. Uji efektivitas metode isolasi DNA genom kopi arabika (Coffea arabica L.) asal kabupaten Jayawijaya. Jurnal Biologi Papua. 8(1): 7-12.

Mawardi, A., L.E. Aisoi dan P.N. Lefaan. 2017. Kloning dan Analisis Bioinformatika Gen MSP1 Plasmodium falciparum Isolat Kota Jayapura. Jurnal Biologi Papua. 10(1): 1-10.

Mulyani, Y., A. Purwanto., I. Nurruhwati. 2011. Perbandingan Beberapa Metode Isolasi DNA untuk Deteksi Dini Koi Herpes Virus (KHV) Pada Ikan Mas (Cyprinus carpio L.). Fakultas Perikanan dan Ilmu Kelautan, Universitas Padjadjaran. Jurnal Akuatika. 2(1).

McCarthy JS, Marjason J, Elliott S, Fahey P, Bang G, Malkin E. 2011. A Phase 1 Trial of MSP2-C1, a Blood-Stage Malaria Vaccine Containing 2 Isoform of MSP2 Formulated with Montanide® ISA 720. PLoS One ; 6(9): 1-13.

Neil Spooner . 2013. A dried blood spot update:still an important bioanalytical technique. Bioanalysis, 5(8):879-883.

Oliveira R.V., Jack H. & Enaksha W. 2014. Fully-Automated Approach for Online Dried Blood Spot Extractionand Bioanalysis by Two¬D imensional-Liquid Chromatography Coupled with High- Resolution Quadrupole Time-of- Flight Mass Spectrometry. Anal. Chem. 86:1246-1253.

Rizwana Q., Raka J. & Atul A. 2013. The use of dried blood spot samples in screening drugs of abuse. Pharmacology & Pharmacy. 4:152-159.

Sambrook, J., Fritsch, E.F., and Maniatis, T. (1989): Molecular Cloning: a Laboratory Manual, New York, Cold Spring Harbor Laboratory Press

Sharma A., Swati J., Mahendra S. & Jawahar L. 2014 Dried blood spots: Concepts, present status, and future perspectives in bioanalysis. Drug Test. Analysis. DOI 10.1002/dta.1646.

Soto M., Roger P., Valerie A., Mylowagner, Ronya P., Manuel P., James M., Christopher A.J., Kevin L.S. & Marc W.R. 2014. Evaluation of matrix microsampling methods for therapeutic drug candidate quantification in discovery-stage rodent pharmacokinetic studies. Bioanalysis, 6(16):2135-2146.

Wenkui Li., John D., Paul M., Jimmy & Francis L.S.T. 2015. LC- MS/MS bioanalysis of loratadine (Claritin in dried blood spot (DBS) samples collected by subjects in a clinical research study. Journal of Chromatography B, 983:. 117-124.

Wilhelm A.J., Jeroen C.G., Burger D. & Eleonora L.S. 2014. Therapeutic Drug Monitoring by Dried Blood Spot: Progress to Date and Future Directions. Clin. Pharmacokinet, 53:961-973.

Downloads

Published

2020-03-05

Issue

Section

Research Articles