Optimization of Binding Buffer Composition (Polyethylene Glycol, Sodium Chloride and pH) for Extraction of DNA from Biological Fluids Using Polyethyleneimine Functionalized Iron Oxide Nanoparticle-Based Method

Imran Khan,1 Gaurav Kaushik,1 Chaitenya Verma,2 Richa Vashishtha,3 Vinay Kumar4 1School of Allied Health Sciences, Sharda University, Greater Noida, Uttar Pradesh, 201310, India; 2Department of Biotechnology, Sharda University, Greater Noida, Uttar Pradesh, 201310, India; 3Department of Medicine, All India Institute of Medical Sciences, New Delhi, 110029, India; 4Pennsylvania State University Hershey Medical Center, Hershey, PA, 17033, USACorrespondence: Gaurav Kaushik; Vinay Kumar, Email [email protected]; [email protected]: Efficient extraction of DNA from biological fluids is crucial for applications in molecular biology, forensic science, and clinical diagnostics. However, traditional DNA extraction methods often require costly reagents and lengthy procedures. This study aims to optimize the binding buffer composition for DNA extraction using polyethyleneimine-coated iron oxide nanoparticles (PEI-IONPs), which offer the dual benefits of magnetic separation and high DNA-binding efficiency.Methods: The effects of three key binding buffer components—polyethylene glycol (PEG-6000), sodium chloride (NaCl), and pH—on DNA adsorption efficiency were systematically evaluated. Blood samples were treated with PEI-IONPs under various conditions, and DNA concentration, yield, and purity were quantified. Nanoparticle functionalization was confirmed through characterization, and DNA quality was validated via agarose gel electrophoresis.Results: The optimized binding buffer composition consisted of a PEG-6000 concentration of 30%, NaCl concentration of 0M, and pH of 4, which yielded the highest DNA concentration (34 ± 1.2 ng/μL), yield (6.8 ± 0.2 μg), and purity (A260/A280 ratio of 1.81). These conditions significantly improved DNA recovery compared to suboptimal buffer compositions.Conclusion: The findings highlighted the critical role of binding buffer composition in maximizing DNA recovery. The use of optimized PEI-IONPs provided a rapid and efficient method for DNA extraction, supporting its potential for applications in scientific and clinical research. Future studies should explore the robustness of these optimized conditions across diverse biological fluids and extraction settings.Keywords: polyethyleneimine, nanoparticles, DNA, TEM, FTIR, blood