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Fabrication of a Centrifugal Microfluidic System for Cell-free DNA Isolation from whole Blood as a Biomarker

Hatami, Ali | 2021

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  1. Type of Document: M.Sc. Thesis
  2. Language: Farsi
  3. Document No: 54141 (06)
  4. University: Sharif University of Technology
  5. Department: Chemical and Petroleum Engineering
  6. Advisor(s): Saadatmand, Maryam; Garshasbi, Masoud
  7. Abstract:
  8. Today, medical science has come to the conclusion that the best time to start treating a disease is in its early stages. Therefore, the topic of early detection has been widely welcomed in scientific circles. Today, one of the safest methods for early detection in medicine is the study of biomarkers. These biomarkers include CTC, cfRNA, cfDNA, Exosome, etc. For instance, in the discussion of early cancer detection, the US Food and Drug Administration has so far identified two biomarkers, cfDNA and CTC, as the safest markers. The cfDNA marker is also widely used not only in the diagnosis of cancer, but also in the field of sex diagnosis and fetal Down syndrome in the first weeks of pregnancy. One of the biggest challenges in extracting biomarkers is their very low concentration in body fluids. One of the best ways to isolate fluids on a small scale is to utilize microfluidic knowledge. Therefore, the purpose of this project is to fabricate a centrifugal microfluidic system to isolate cfDNA from whole blood. In the fabrication of this microfluidic device, various internal and external forces have been used to increase efficiency. These forces include internal centrifugal forces, inertia, and external magnetic forces. Also, various types of microchannels, siphon valves and septum valves have been used in the design and fabrication of this microfluidic disc. In this Lab-on-a-Disc, seperation of cfDNA from whole blood is based on using of magnetic silica beads, which are controlled from the outside of the disk by a series of powerful magnets. It should be noted that in the design of this disk, in order to achieve the maximum possible response to a sample to answer system, whole blood is used as a sample. Also, to evaluate the function of the disc, fetal cfDNAs were extracted from the mother's blood. Therefore, in the first step, the disk performance was evaluated in terms of hydrodynamics and then in terms of efficiency by real samples. As mentioned, this disk is able to use blood samples and isolate its plasma, therefore to study the purity of the extracted plasma, in the absence of contaminants such as hemoglobin, red blood cells, white blood cells and platelets, various tests were used. These tests include the use of hemoglobin test kits, spectrophotometry, Hemocytometer, etc. The designed disk is capable of extracting 1.3 ml of plasma from 3 ml of blood, and the results of the quality of the extracted plasma show its very high purity. Finally, cfDNAs extracted by disk were examined using a male fetal gender identification kit and real-time PCR. The results of the real-time PCR process indicate the correct function of the disc in extracting cffDNAs in 10 Landa samples from cycle 34 onwards. Since the disk requires only 1 ml of plasma to extract, it leads to a shorter extraction time, as well as fully automatic extraction, which is completely superior to the manual method
  9. Keywords:
  10. Biofluid ; Microfluidic System ; Lab-on-a-CD (LOCD) ; Early Detection ; Cell Free Deoxyribonucleic Acid (cfDNA)Biomarker ; Biomarker ; Magnetic Silica Beads

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