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1、Piezoelectric Immunosensor for SARS-Associated Coronavirus in Sputum Boli Zuo, Shanmao Li, Zhao Guo,* Jinfang Zhang, and Chuanzhi Chen The Analytical Center, Institute of Chemical Defense of PLA, 1048#, Beijing, 102205, P. R. China A piezoelectric immunosensor was developed for the detection of SARS
2、-associated coronavirus (SARS-CoV) in sputum in the gas phase. Horse polyclonal antibody against SARS-CoV was bound onto the PZ crystal surface in an ordered orientation through protein A. The antigen sample was atomized into aerosol by an ultrasonator, by which the antibody on the crystal could spe
3、cifically adsorb SARS antigen and the changed mass of crystal would lead a frequency shift. A frequency counter was employed to record the admittance frequency, and the plot of changed frequency was displayed on the computer. Under the optimized conditions, the frequency shifts were linearly depende
4、nt on antigen concentration in the range of 0.6-4 g/mL. The device has good reproducibility (could be reused 100 times without detectable loss of activity), stability (the immunosensor was stable for more than two months when stored over silica gel blue at 4-6 C), short analyzing time (less than 2 m
5、in), and specificity. Severe acute respiratory syndrome1(SARS) is a viral respira- tory illness caused by a coronavirus, called SARS-associated coronavirus (SARS-CoV). SARS was first reported in November 2002. Over the next few months, the illness spread to more than 30 countries. According to the W
6、orld Health Organization (WHO), to present, more than 8000 people worldwide became sick with SARS; The SARS global outbreak of 2003 was contained; however, it is possible that the disease could reemerge when the temper- ature is low enough. The main way that SARS seems to spread is by close person-
7、to-person contact. The virus that causes SARS is thought to be transmitted most readily by respiratory droplets (droplet spread) produced when an infected person coughs or sneezes. On the other hand, oral-fecal transmission might also be an important route of transmission of the disease.2 To cope wi
8、th the challenge of SARS, many organizations have begun to study how to detect and diagnose it and have had some success. At present, the methods of SARS detection are mainly based on two different theories. One theory is by examining virus RNA by molecular biology, such as reverse transcription-pol
9、y- merase chain reaction (RT-PCR)3-5and fluorescent polymerase chain reaction.6,7They can identify the SARS-CoV in the first part. They require a high specificity between primers and templates but do not have a very high sensitivity so that false negative results are likely. On the other hand, these
10、 methods need specific equipment, costly reagents, and complicated technique. The other one is based on SARS antibody analysis in patient sputum, such as the enzyme-linked immunosorbent assay method8,9and immu- nofluorescence assay.10But methods require more time (10-20 days). In this period, the vi
11、rus would have been transmitted to others The piezoelectric (PZ) immunosensor is an important biosen- sor. In 1972, Shons11made the first PZ immunosensor to detect cow serum IgG antibody. Since then, this method has been important in biodetection for sensitivity, specificity, simplicity, and swiftne
12、ss. The PZ immunosensor consists of a PZ crystal with an antigen or antibody immobilized on its surface. The biospecific reaction between the two interactive molecules, one immobilized on the surface and the other free in solution or gas phase, can be followed in real time. The PZ crystal is a mass
13、device, which means that any surface mass change reflects on its resonant frequency. The increase of mass at the sensor results in a decrease of the frequency as shown by Sauerbrey, who first introduced the quartz crystal microbalance principle.12This relationship of frequency to mass has been outli
14、ned by the following equation:13 where f is the change in resonance frequency of the coated * To whom correspondence should be addressed. E-mail: zhetiandadi . Tel: 0086-10-66756549. Fax: 0086-10-66756518. (1) He, F. C. Severe Acute Respiratory Syndrome, 1th ed.; Science Press: Beijing, 2003; Chapte
15、rs 2, 6, and 9. (2) Peiris, J. S. M.; Chu, C. M.; Cheng, V. C. C.; Chan, K. S.; Hung, I. F. N.; Pool, L. L. M.; et al. Lancet 2003, 361, 1767. (3) Zhou, B. P.; Chen, X. C.; Wang, H. S.; Li, M. Z.; Hu, Y. W.; Du, F.; Xu, L. M.; Yang, G. L. Chin. J. Exp. Clin. Virol. 2003, 17 (2), 137. (4) Ren, Y.; Di
16、ng, H. G.; Wu, Q. F.; Chen, W. J.; Chen, D.; Bao, Z. Y.; Yang, L.; Zhao, C. H.; Wang, J. Acta Acad. Med. Sin. 2003, 25 (3), 368. (5) Yang, J.; Wang, Z. H.; Chen, J. J.; Hou, J. L. J. First Military Med. Univ. 2003, 23 (5), 424. (6) Wu, B. Q.; Zhong, H. H.; Gao, J. P.; Liu, S. P.; Heng, W. J. E. W.; Gu, J. Chin. J. Pathol. 2003, 32 (3), 212. (7) Wu, X.; Cheng, G.; Di, B.; Yin, A.; He, Y.; Wang, M.; Zhou, X.; He, L.; Luo, K.; Du, L. Chin. Med. J. (Engl.) 2003, 116 (7), 988. (8) Guan, M.; Chen, H.
