Prceedings of 2006 In ternational Conferenceon Microtechnologies in Medicine and Biology

Okinawa, Japan 9-1 2 May 2006

Development of Biosensor Chip for Clinical Diagnosis Using SurfacePlasmon Resonance Imaging with Multi-Microchannels

T. Suzuki, Y. Teramura*, K. Inokuma, I. Kanno, H. Iwata** and H. KoteraDept. of Microengineering, Kyoto University

Yoshida-honmachi, Sakyo-ku, Kyoto, 606-8501, JapanTel +81-75-753-3559, Fax +81-75-771-7286, E-mail takaakigme.kyoto-u.ac.jp

* Dept. of Polymer Chemistry, Kyoto University53 Kawara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507, Japan* * Institute for Frontier Medical Sciences, Kyoto University53 Kawara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507, Japan

Abstract

A compact biosensor chip for clinical diagnosis is presented. The proposed biochip integrated three independent microchannels onone chip partially coated with Au thin film as a Surface Plasmon Resonance (SPR) excitation layer. For clinical diagnosis, theaffinity binding of unlabeled biological molecules onto the Au surface can be quantitatively analyzed by SPR imaging with themulti-microchannels, i.e. one biosample and two reference flows to obtain an analytical curve.

Keyword: Surface Plasmon Resonance Imaging, Clinical Diagnosis, Multi-microchannels

1 INTRODUCTION

Miniaturized SPR sensors with multichannels have beendeveloped as an alternative to laboratory SPR systems torealize mobile, compact, and cost-effective sensing devices[1-3]. However, most of the systems in which microchannelsand substrates are not bonded decrease the detectionsensitivity due to cross contamination and dehydration ofimmobilized materials.In this paper, a compact and high-sensitive biochip forclinical diagnosis is presented. The proposed biochip arecomposed of three independent microchannels on one chip,and quantitatively measures the affinity binding of unlabeledbiological molecules using the Surface Plasmon Resonance(SPR) imaging. The protein adsorption by the change in themicrochannel height and the flow velocity was observed forgeometry optimization of microchannels in the chip in orderto improve the detection sensitivity and the throughput ofscreenings.

2 EXPERIMENTAL

2.1 Fabrication Process

Sensing microchannels based on the SPR imaging is rapidlyand easily made from poly(dimethylsiloxane) (PDMS) usingsoft micromachining techniques as shown in Fig. 1. The thickphotoresist SU-8 was first spin coated with a thickness of

20pm on a glass substrate. A master of microchannels wasfabricated on the glass substrate using photolithographyprocedure. Then, liquid PDMS prepolymer was poured on themaster and degassed using vacuum-forming to mold themicrochannels. The PDMS microchannels were peeled offthe master. On the other hand, thin Cr and Au films as a SPRexcitation layer were partially deposited on glass substrates(refractive index n=1.52) using a vacuum evaporator with ametal mask. Finally, the PDMS microchannels and the glasssubstrates were bonded using 2 plasma bonding technique[5] in order to increase the detection sensitivity of the SPRimaging. Irreversible bonding increases upper limit of fluidvelocity and liquid switching speed without crosscontamination and dehydration of immobilized materials. Thedimensions of the fabricated multi-microchannels were500pm wide, 20pm height, and 2mm intervals. The biosensorchips can be used by putting on the optical apparatus, thusallowing it to be used as a low-cost disposable device.

2.2 Measurement

To demonstrate sensing characteristics of the proposed chip,the adsorption of bovine serum albumin (BSA) onto the goldsurface was evaluated by putting the prepared chip on anoptical apparatus as shown in Fig.2. Three samples withdifferent concentrations of BSAlwater were simultaneouslyinfused to the microchannels by syringe pumps connected toinlets of microchanmles by Teflon microtubes.

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3 RESULTS AND DISCUSSION

3.1 Evaluation of Quantitative Measuremenit

Three samples with 0.1%, 0.5% and l.0% BSA/water weresimultaneously iMnfsed to thi fabricated microchannels. SPRimages as a fuintion of time were clearly bserved by a CCDcamera as shown in Fig.3. From SPR sensorgam denrved bythe images m Fig.4, it is seen that the quantity an the rate ofthe adsorption depend on the concenrtio6n of thi sample imaccordance with ldinetics. Therefore, the proposed system isapphcable ftr chlaracteiing and quaifyin biomblecular

interactions, e.g. detection of the tuimor marker im clinicaldiagnosis using the anten-antibody reaction.

3.2 Microchannel Height and Flow Velocity Dependlences

We evaluated the protein adsorption by the change im themicrochannel heig adi the flow velocity for geometryoptimization of multi-microchaneIs in SPR iaing sensorsin orde to improve the detection sensitivity nd thethrohput of screenings. hi consequence, the adsorption rateis proportional to one-third power of the flow velocity, and isinversely proportional to one-third power ofthe heigt.

(2-a) t7hini Cr 1aerV with Im thicknev, iJ paftially opofttdon a gIas sUbWtrac using vacuum evaportinM

2-b) Thin Au laayr with 49nni thickness is partially depositedon a4 glass substratoe Osing vacuum evaporation

(I c LNiOPid MPMS prepolymnr is poured ontote MOld deaetated and cured,

DMS ntiiFig.l F

j, g* Leis

Waste

(WhVite light soure)Narrow band passtser

(905 n) o e

CCD camera aC

Fig. 2 S1Oib- of! SP iiian _pnelast f. *hdfud s-ns

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ACKNOWLEDGEMENTS

(a) Omin (b) 8minFig. 3 SPR images as a fnction of time.

5 20000 _B:MII

t10000 -

0 10 20 30 40Time [mi]

Fig.4 SPR sen:sorgram of BSA/waterat flow velocity 3.6mmlsee.

50 60

This study is a part of Kyoto City Collaboration of RegionalEntities for the Advancement of Tecniological Excellence ofJST on the basis of research results suported in part byMizuho Foundation for the Promnotion of Sciences. andCenter of Excellence for Research and Education onComplex Functional Mechanical Systems (COE program) ofMEXT, Japan.

REFERENCES

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[2] C. E. H. Berger, T. A. M. Beumer, R. P. H. Kooyman, J.Greve, 1Suface plasmon resonance multisensing," AnaLChem., Vo1.70, pp.703-706, 1998.

[3] H. J. Lee, T. Goodrich. R. M. Com, "SPR imagmgmeasurements of 1-D and 2-D DNA microarrays createdfrom microflouidic channels on gold films," AnaLChae., Vo1.73, pp.5525-5531, 2001.

[4] J. C. McDonald, D. C. Duffy, J. R. Anderson, D. T.Chin, H. K. Wu, 0. J. A. Schueller, G. M. Whitesides,"Fabrication of microtluidic systems inpoly(dimethylsiloxane)," Electrophoresis, Vol.21,pp.27-40, 2000.

[5] S. Bhattacharya, A. Datta, J. M. Berg, S. Gangopadhyay,"StLdies on Sure Wettability of Poly(Dimethyl)Siloxane (PDMS) and Glass Uhder Oxygen-PlasmaTreatment and Correlation With Bond Sttength,'Mcroclectromcchanical Systmsr, Vol.14, pp.590-597,2005.

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