A biotin-hydrogel-coated quartz crystal microbalance biosensor and applications in immunoassay and peptide-displaying cell detection

作者：Chen Hsiu Mei^*; Huang Tsun Han; Tsai Ruey Ming

来源：Analytical Biochemistry, 2009, 392(1): 1-7.

DOI：10.1016/j.ab.2009.05.029

• Biotin
• Hydrogel
• Quartz crystal microbalance
• Streptavidin

A biotin-coated quartz crystal microbalance (QCM) chip was prepared by dip-coating a long-chain alkanethiol-modified crystal with precoupled dextran-biotin hydrogels. The resulting biotin chip was used to affinity-immobilize streptavidin (SAv) and was then further employed for various biosensor assays. First, the SAv chip allowed efficient on-line binding of biotinylated bovine serum albumin (bBSA), followed by a sensitive and specific response toward anti-bovine serum albumin (BSA) antibodies. Three consecutive immunoassays were reproducibly demonstrated with a single chip. Tire apparent binding kinetics with k(on) = 5.9 mu M(-1) h(-1), k(off) = 10.1 h(-1), and K(D) = 1.71 mu M was readily resolved by fitting the real-time sensorgrams. Second, the capability of the SAv chip to selectively recognize recombinant Escherichia coli with flagella displaying an artificial SAv binding peptide, Strep-tag II, was demonstrated by QCM analysis and verified by scanning transmission electron microscope (STEM) image analysis with biotin-coated gold nanoparticles as the label. Finally, the affinity of the cell-displayed Strep-tag II peptide to surface-coated SAv, K(D) = 6.8 x 10(8) CFU/ml, was resolved on-line using equilibrium binding kinetics by QCM. This Study presents an easy, economical, and reliable method of preparing high-performance SAv-coated biotin chips with potential for application in real-time repetitive immunoassays, on-line binding kinetics Studies, and high-affinity peptide screening.

• 出版日期2009-9-1