Covalent virus layer for mass-based biosensing.
M13 virus particles were covalently attached to a planar gold-coated quartz crystal microbalance (QCM) through reaction with a self-assembled monolayer of N-hydroxysuccinimide thioctic ester, followed by incorporation of the blocking agent bovine serum albumin. This immobilization chemistry produced a phage multilayer having a coverage equivalent to approximately 6.5 close-packed monolayers of the virus. The properties of this "covalent virus surface" or CVS for the mass-based detection of a 148.2 kDa antibody were then evaluated in a phosphate buffer using a flow injection analysis system. The mass of the CVS increased with exposure to an antibody (p-Ab) known to bind the phage particles with high affinity. Bound p-Ab was removed by washing with 0.5 M HCl thereby regenerating the sensor surface. A calibration plot for p-Ab binding was constructed by repetitively exposing the surface to p-Ab at concentrations between 6.6 and 200 nM and HCl rinsing after each exposure. The mass-concentration relationship was linear with a sensitivity of 0.018 microg/(cm2 nM) and a limit of detection of 7 nM or 1.3 pmol. The CVS could be saturated with high doses of p-Ab enabling the determination that an average of approximately 140 binding sites are available per M13 phage particle. Exposure of the CVS to a second, nonbinding antibody (n-Ab) did not cause a measurable mass change. These results demonstrate that the covalent virus layer is a rugged, selective, and sensitive means for carrying out mass-based biodetection.
Yang
Li-Mei C
LM
Department of Chemistry, University of California, Irvine, CA 92697-2025, USA.
Diaz
Juan E
JE
McIntire
Theresa M
TM
Weiss
Gregory A
GA
Penner
Reginald M
RM
D000937:Antigen-Antibody Reactions; D017104:Bacteriophage M13 / Q000737:chemistry* / Q000648:ultrastructure; D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods*; D002021:Buffers; D002138:Calibration; D003460:Crystallization; D004952:Esters / Q000737:chemistry; D017022:Flow Injection Analysis; D006046:Gold / Q000737:chemistry*; D008970:Molecular Weight; D011791:Quartz / Q000737:chemistry*; D012710:Serum Albumin, Bovine / Q000737:chemistry; D013388:Succinimides / Q000737:chemistry; D013499:Surface Properties; D013997:Time Factors
D002021:Buffers; D004952:Esters; D013388:Succinimides; D011791:Quartz; D012710:Serum Albumin, Bovine; D006046:Gold; C001426:N-hydroxysuccinimide