TY - THES T1 - The application of surface plasmon resonance (SPR) immuno-biosensors for medical diagnosis A1 - Chung,Jiwon Y1 - 2007/05/02 N2 - A biosensor is an analytical device to detect target analyte in a complex mixture by using high selectivity of biomolecules as the molecular recognition tool. Especially, the immunoaffinity (IA) biosensor exploits the highly specific and selective interaction between antigen and antibody for the detection of a target analyte. The surface plasmon resonance (SPR) sensor was used for label-free detection and real-time monitoring, and SpreetaTM was used for this work. The major objective of this work is the development of SPR biosensor for medical diagnosis with the features of a cost-effective test by 'additive assay';, short analysis time through 'simultaneous detection'; and high sensitivity by 'signal amplification';. [1] First topic is a reuse method of IA biosensor called 'additive assay';. In the 'addtive assay'; method, the sample is repeatedly injected to the IA layer without removing the already bound analytes by chemical treatment and then the concentration of sample is calculated from the actually measured signal by using previously prepared correlation curve between the accumulated concentration of additively injected sample and accumulated signal which represents the number of occupied binding sites. The application of additive assay for real medical diagnosis was demonstrated by using tumor marker (CA 19-9) as a target analyte. When the concentrations of four samples were analyzed by using the correlation curve of CA 19-9, the average deviation of the calculated concentrations from the real concentrations was estimated to be 5.3 %. [2] The second topic is the 'simultaneous detection'; which enables the detection of multiple analytes on the single sensor element with single sample treatment. In the 'simultaneous detection';, a sample with several analytes is treated to the single sensing region which has multiple receptors for each target analyte and then the concentrations of each analyte is analyzed sequentially by using the respective correlation between the concentration and signal. In this work, two simultaneous detection models (Model 1 and Model 2) were devised for the samples with the following composition: (1) one target analyte resulting in a sensor response without any label and the other analyte with only additional label (Model 1), (2) both target analytes requiring additional labels for detetion (Model 2). The real medical diagnosis based simultaneous detection (Model 2) was demonstrated by analysis of human chorionic gonadotropin (hCG) and human albumin (hA) in human urine for the diagnosis of preterm delivery of patients with diabetes. The hCG has been used for the qualitative pregnancy test. The hA can be used for diabetes, who have the steeply increased prevalence of preterm delivery. The average errors of analysis based on Model 2 were 6.5 and 5.9 % for hCG and hA, respectively. [3] The third topic is the improvement of sensitivity and detection limit through two 'signal amplification'; methods by using mass label (A) and by the orientation control of IA layer (B): (A) The mass label attached to the already bound target analyte increases the total mass attached to the sensor surface and then it induces the increase of SPR signal. Among several labels, the amplification using the PAP complex was selected as the most efficient method. The feasibility of this signal amplification method was demonstrated by analysis of human hepatitis B virus (hHBV) antibody. The result from PAP method shows that the detection limit of SPR biosensor (0.64 nM) approached closely to cut-off value for medical diagnosis (0.24 nM) by using the commercial ELISA kit. (B) The sensitivities of immunosensors are known to be improved by the control of IA layer. As more target analytes can be attached to this controlled IA layer, the signal at the same concentration of target analyte can be increased. In this work, among several controlled IA layers, the NeutrAvidin-protein A complex on gold surface of SPR biosensor showed the highest surface density of receptor and ligand antibody. And the binding ratio of ligand per unit receptor antibody was also one of the highest values. When the NeutrAvidin-protein A complex was prepared on biotin-labelled SAM, the binding ratio of ligand per unit receptor was found to be significantly improved. For the feasibility test of orientation control, the NeutrAvidin-protein A complex was applied for the detection of a tumor marker, carcinoembryonic antigen (CEA). The sensitivity was improved to be 1.5-fold higher than bare gold surface and the detection limit of 30 ng/ml was achieved. These results demonstrated the 'additive assay'; for cost-effective test, 'simultaneous detection'; for short analysis time and 'signal amplification'; for high sensitivity. The presented three methods in this study will be applied for the development of a practical SPR biosensor for the various medical diagnosis. KW - Biosensor KW - Oberflächenplasmonresonanz KW - Untersuchung KW - Antigen-Antikörper-Reaktion CY - Saarbrücken PB - Universitäts- und Landesbibliothek AD - Postfach 151141, 66041 Saarbrücken UR - http://scidok.sulb.uni-saarland.de/volltexte/2007/1115 ER -