Development of QCM biosensor to detect a marine derived pathogenic bacteria Edwardsiella tarda using a novel immobilisation method.
QCM technology offers a real time output, simplicity of use and cost effectiveness in addition to high sensitivity. Sensitivity of QCM immunosensor can be enhanced by improving the immobilisation procedure on the quartz surface. The immobilisation strategy should be able to control both the amount and the orientation of the antibody (immunoglobulin; IgG) on the transducer for high affinity to antigens. This study introduced a new methodology recruiting oxidised IgG to expose aldehyde group in Fc region to cross-link to hydrazide conformed on self assembled monolayer (SAM) and compared with three conventional methods. Consequently, it was proved that considerable amount of antibody was immobilised and the sensitivity of new methodology was higher than other methods while ability of new methodology to immobilise IgG was lower than the conventional methods. The frequency shifts following bacterial cell injection were positively related to the frequency shifts after the injection of IgG and the amounts of bacterial cells, revealing that the frequency shifts after bacterial cell injection fully represented the weight change by specific attachments of bacterial cells to the IgG cross-linked on the gold surface. Specificity was tested on different bacteria including E. coli, V. vulnificus and A. hydrophila and showed no significant non-specific affinity on the tested bacteria. It was also demonstrated that the prepared sensor chip was stable enough to withstand repeated surface regeneration. Indeed, polyclonal antibody was more effective to detect antigen than monoclonal antibody which binds to only one epitope of antigen. Conclusively, the new methodology is appeared to be more sensitive than conventional methods tested and reusable for 10 times.
Department of Chemistry, Faculty of Marine Bioscience and Technology, Kangnung National University, Kangnung, Republic of Korea.
Sung-Rok
SR
Hong
Suk-Jung
SJ
Choi
Hyun Do
HD
Jeong
Suhee
S
Hong
D001422:Bacterial Adhesion; D015374:Biosensing Techniques / Q000295:instrumentation*; D015169:Colony Count, Microbial / Q000295:instrumentation*; D020609:Edwardsiella tarda / Q000276:immunology / Q000302:isolation & purification*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D007118:Immunoassay / Q000295:instrumentation* / Q000379:methods; D008386:Marine Biology / Q000295:instrumentation* / Q000379:methods; D055617:Micro-Electrical-Mechanical Systems / Q000295:instrumentation*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity