Abstract
High plasma homocysteine levels can cause an increased risk of cardiovascular, cerebrovascular, and peripheral arterial diseases. Besides this, Alzheimer’s disease and other dementias, osteoporosis, diabetes and renal disease due to folate and B-vitamin deficiency, various drugs or pre-existing atherosclerotic diseases may be the result of high homocysteine levels. The presented research work aimedto perform the detection of Homocysteine (Hcy) by using Quartz Crystal Microbalance (QCM) biosensor. The temperature controlled QCM system was a home-made designed and constructed equipment which can use silver electrod quartz crystal. The modification of silver electrod quartz crystals surfaces was achieved by the surface cleaning process with sodium hydroxide, acetone and methanol in a consecutive manner. Then self-assembled monolayer of cysteamine and chemical coupling of glutaraldehyde (GA) to free end of monolayer was achieved to create the new functional surface in order to complete the formation of spacer arm/ligand. Homocysteine specific recognizing ligand, anti-Homocysteine antibody was immobilized to glutaraldehyde coupled surfaces. The change in resonance frequency values were measured for each modification step. The optimization of dilution ratio of the antibody solution was performed to modified surfaces. The least dilution ratio of antibody, 1/10000 v/v, was determined as optimum antibody ratio. The detection of homocystein was analysed at a detection limit of 0.1 µM and the linear ranges of calibration curves were estimated as 0.1-2.0 µM and 10-50 µM. Homocysteine values indicated good linearities (R2=0.9813 and 0.9875, respectively). The relative standart deviation (RSD %) for precision was calculated as less than 10%. In conclusion, it was found that the detection of homocysteine can be done both in nano- and micro-molar concentration levels. Additionally, designed biosensor showed desired stability and reproducibility. Finally, a new method different from the present methods for the use in the analysis of Hcy was proposed and developed which detects homocysteine by designed QCM technique with a rapid, cheeper and less pretreatment processes. Additionally, homocysteine detection was performed in nano- and micro- molar concentration values.
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