The detection and separation of small biomolecules from complex mixtures and the possibility of their recovering for further analyses have great benefits for the early diagnosis and prognosis of diseases. Developing simple, sensitive, and cost-effective tools that allow the rapid and accurate assembly and isolation of molecular biomarkers has the potential to improve both patient care and hospital logistic efficiency towards personalized and affordable treatments of diseases.In this work, we presenta method consisting ofUV-vis-spectroscopy assisted-magnetophoresis for the monitoring of DNA hybridization. For this purpose, a magnetic device generating 7.5 T/m uniform magnetic field gradient was designed and incorporated to a commercial spectrophotometer. Different batches of colloidal superparamagnetic particles (SMPs), with different elemental compositions, were functionalized with twenty-mer complementary oligonucleotides, TB1 and TB2. When the functionalized SMPs-TB1 and SMPs-TB2 are mixed and incubated, the hybridization process of TB1 and TB2 occurs resulting in the formation of colloidal aggregates. When brought under the magnetic field, depending on the magnetic strength (Γ) of the formed aggregates, they separate either faster or slower than the non-functionalized SMPs. The difference in magnetic separation time (Δt) is optically monitored by measuring the real time transparency of the suspension at specific wavelengths. The detection of aggregates at concentrations of 0.001% w/v was achieved, showing Δt ranging from 113-228 s. Based on the changes of Δt, the study addresses how electrosteric, magnetic, and hydrogen bonding interactions affect the hybridization process and suggests optimum experimental conditions for accurate monitoring of TB1-TB2 hybridization.
Keywords: Colloidal-stability; Magnetic-isolation; Magnetic-separation; Magnetophoresis; Molecular-diagnosis; Oligonucleotide-hybridization; Optical-monitoring; Surface-functionalization.
Copyright © 2020 Elsevier B.V. All rights reserved.