TAP CIJ HAT IIIEK KHiCH, TAP IS, SO Tl - 201S
Nghien CLPU qua trinh hap phu va giai hap phu cue probe bi§n doi thiol tren dien cyc vang bang phu'ang phap quet the vong tuin hoan .^ ,.-
• CaoHCruTi^n ' ^ '"^
• NguySn Quang Thi$n
• Ha Van Linh
• U Van Hi6u
Tnffrng Dgi hpc Khoa hgc Ty nhifin, OHQG-HCM C Bai nhdn ngay 23 ihdng 09 ndm 2014. nhgn dSng ngdy 03 ihdng 08 ndm 2015}
TOM
T A TTrong bdi bdo ndy chiitng tdi nghiSn ci>u cdc tinh chit di$n hda cOa I6p (Son ti/ t$p hop (SAM) diigc t^o bCfi sv hip phi^ vd gidi hip phtj cOa cdc alkanethiol trdn bi mdt di$n ct/c sp'/ nano vdng (AuNW) bing phuvng phdp CV. Kit qud phdn tfch cho thiy llnh hi$u qud cOa quy trinh gidi hip tfaf han 90 %. Vi$c dp m^t hidu <Ti$n thi dm gdy ra
sy gidi hdp cOa SAM, kdt qud Id cdc SAM bj
tdch rai khoi dien ct/c. Muc dich cOa chOng tdi Id nhim tdi sCr dung cdc didn ci/c AuNW sau khi chiing da du^fc chux; ndng hda bdi SAM. Bdn canh dd, kit qua cOa nghidn cCeu ndy cQng giup trdnh cho SAM khdng bi gidi hdp bdi SI/ dien hda vd nhd vdy gidp bdo v$
mSu hidu qua han trong qud trinh khdo sdt.
TO khda: SAM, cdm biin sinh hqc, qudt thi vdng tudn hoan, hip phu, gidi hip phij.
MdDAU
Cdc don ldp thiol tr£n cdc chat nen (substrates) khdc nhau, d^c bi^t tr€n vdng, to ra uu vi^t vdi nhQng d$c tfnh tv t$p hpp cua chiing.
Phan tir thiol dupc gSn len b^ mat ciia chat ngn thdng qua lien ket hoa hpc gli^ nguyen tu chat nen vd nguyen tu thiol trong alkanethiol. Tuong tdc ky nudc ciia cdc chuSi alkyl vdi nhau chii yiu Id do tuong tdc van der Waals [1]. Tfnh chdt nay cho p h ^ sir hdp phu h6a hpc ciia cdc alkylthiol len be m^t ciia chat nen (kim loai, ban dan), tao thdnh don ldp. Ldp don tao thdnh ndy dupc gpi Id ldp don t^ tdp hpp (SAM).
Nhitng SAM cua thiol dd dupc nghien cihi tii nhibig ndm 1980 cua th€ ki 20 [2, 3] vi pham vi img dyng r^ing rai cho nhieu be m$t khde nhau.
Hon n i ^ SAM cd cdc tinh chdt v$t li va hda hpc
rat tot, chiing khdng duoc qua trinh dn mdn, cd tdc dung nhu tac nhdn can quang trong quang khde va co the bien ddi vdi nhung nhom chuc ndng khac nhau (thi du nhu -COOH, -OH, -CH,, -CF3). Tuy thupc vao nhom chiic ndng d ddu ciia SAM md cd the ung dung phu hpp cho cdc cdm bien hda hpc va quang hgc. Hdu h6t cdc nghien ciru cd li^n quan den ldp co thanh phan la thiol b>eo (aliphatic thiol). Cac tinh chdt di£n hda b|
thay ddi dang ke khi cdc phan tu thiol dugc hap phu hoa hoc l€n tie'mat dien circ. Tinh chdt dien hda cua di€n cue bj bi6n ddi hk mat ciing phu thudc vdo tfnh chat cua ban phan tii huu co [4, 5].
Chieu dai cua chuoi hydrocarbon co the dieu chinh tinh chdt vat li vd hoa hpc ciia cdc thiol
SAM tao ra tren be mdt kim lo?i. /
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Scleice & Techiiliiy h n l M i n t , Vol 18, Ko.TI- 2015
Qud trinh hdp phy/giai hdp phu dd duoc nghiSn cim bdng nhieu phuong phdp bao gdm cac ky thu^t dien hda [6-15], bi^n ddi Fourier quang phd hdng ngoai (Fourier transform infrared spectroscopy (FT IR) [13, 15, 16], electrochemical quartz crystal microbalance (EQCM) [17 - 19] vd kfnh hign vi q u a xuySn hdm t^i chd (in situ scanning tunneling microscope (STM)) [20 - 23]. Mpt trong nhung ky thuat quan trpng nhdt cho qua trinh nghign Clhl hdp phy vd giai hap phy ciia SAM la phirong phdp q u a tha vdng tudn hodn (CV). Phuong phdp ndy cung cdp thdng tin vt chdt Iupng cua mang don ldp. Phuong phdp CV dd dupc sit dung d^khao sdt tfnh chdt di^n mdi cua SAM tren be mdt vdng. Cd thl thdy rdng cdc alkylthiol vdi chudi alkyl ddi hon ldm chuyen ddi peak khu sang thfi am hon. Di^u ndy phan dnh thdng qua tuong tdc van der Waals manh m€ giEta cac chudi alkyl ciia phdn tu thiol [6], Qud frinh gidi hdp SAM dupc bdt dka Xii nhCtng sai hdng trong don Idp thiol, chdng hjm nhu khuylt cdc hdng vd ria cua cdc lo tr6ng ndm tdch bi^t. Cdc phdn tu alkylthiol da gidi h i p ket t§p lai trong dung djch H2SO4 [25].
Widrig vd cs [6] dd mo td su gidi hap cua cdc phan tli alkanethiol khdi b€ m^t vdng vdo trong dung djch cd tfnh ki€m nhu qud trinh khu mdt di^n tli theo phdn iing sau:
Au-SR + e" ^ Au + RS" (*) Di^n tfch ciia peak, hinh dang peak cue am cho viec khii gidi hap vd peak thfi cung cdp thong tin hitu fch ciia SAM, chdng ban nhu Id so luong hap phy, d$ dn djnh, ndng iupng hdp phu, su djnh hudng, hlnh thdi bh m?t [6-23]. Mpt nghign ciru chi tiet ciia sy khu giai hdp vd oxi hda tdi ldng ddng ciia cdc thiol SAM nhdm khdng djnh rdng peak thl cua sy khir giai hap Id tiiy thupc vdo chieu ddi ciia chuoi hydrocarbon, nhdm chiic ndng d d^u vd cdc tinh thl b l mdt cua vang [15].
Ngirdi ta tim thdy rdng Idp hap phy ciia cdc phan tir hiiu co dupc giai hap khdi b l mdt dien cue thdng qua mdt qud trinh tao 16 (poration process), hodc tao ra sai hdng trong Idp hdp phy khi dien thl trd nen dm hon. Myc dfch ciia bdi bao nay Id d l nghi€n ciru qud trinh hdp phy/giai hdp phu cua SAM trSn b l m$t vdng bdng phuong phdp CV vdi be mdt di^n c\rc vdng dupc bien doi bdi DNA probe vd mercaptohexanol (MCH).
VAT L I $ U VA PHlTCfNC P H A P Cdc chdt hda hpc dupc sit dyng Id KCl (Merck), K3Fe(CN)6, Ethanol, H2SO4, Phosphate Buffer Saline (PBS) (Sigma-Aldrich). Nudc khii ion vd tiet triing (dien trd sudt 18 M^.cm) dd dupc sii dyng trong sudt qyd trinh thi nghiem.
Cam biln dd dupc chl tao bdng cdch sii dung dien cue spi nano vdng (AuNW) (dai: 1000 pm;
rpng- 2 |im; cao: 60 nm) [8].
Trinh ty ciia DNA probe; 5'-SH-C6- GAGGGTTGTGATGAATTCTCAGCCCTCTT CAAAAACT TCTCCACAACCCTC-MB-3'
Myc lieu Id sdn phdm ciia phdn img RT-PCR (vdi c^p m6i IP: CCAAGGAAAACTG GGTGCAG vd IR: CTTGOATACCACAGA GAATGAATTTTT) [24] tir mRNA cua InterIeukin-8 md khdng can trai qua qud trinh tinb sach gi them (dupc cung cdp bdi Trung tam Y sinh hoc phdn tir, Dai hpc Y Dugc TP HCM).
Trinh ty: cua DNA muc tieu:
-. ...GAGGGTTGTGGAGAAGTTTTTGAA GAGGGCTGA GAATTCAT...3'
Qudt thl vdng tudn hodn Id mot kJ thudt trong dien hda, nd sg do ddng dien dugc tao ra trong binh do dien hda vdi mdt di^n thl dugc dp vdo vd tudn theo djnh ludt Nemst. CV dugc thyc hi§n bdng thl vdng ciia di|n cyc Idm viec vd do ddng dien dugc tao ra.
T r a D g 6
TAP cri HiT Tiifu n t n , T(P IS, sf Tl - 2S1S
Cyclic Voltammogratn Excitation Signal
S
•s , t
ti
0,
%
d
g / \ c / \ b/ U / \
/ \ / \ '
a / i-V \ , cycle 1 cycle 2
Time (s)
Hlnh 1. Bieu dien qud uinh khu xay ra khi qu6t theo chilu dm tir (a) din (d) vd qud trinh oxy hoa xay ra khi qu6t theo chilu duong tfr (d) din (g). Qu^ thl vdng t u ^ hodn thu dupc kit qua ve ddng dien d di^n cyc ldm v i ^ Uong qud trinh quet the.
Khi qudt theo hudng tdi, difn t h l sS qu^t theo chilu dm bdt ddu d difn thl cao (a) vd k i t thiic d dien thl thip (d) vd difn thl tai (d) gpi Id thl chuyen, t^i ddy qud trinh qu^t the sS ddo ngugc 1^, qu^t tir (d) den (g) vd the dugc q u ^ theo chieu duong (Hinh 1).
Qua trinh khu xay ra khi qudt theo chilu dm tir (a) den (d), difn cyc sS nh§n difn tii vd trd thdnh chdt khu, ket qua xudt hifn ddng catod, ddng catod tdng nhanh den khi nong d$ chSt oxi hda trfn bf m^t difn cyc bdng khdng, liic dd ddng catod sS dat cyc dai, tai dd ndng d^ chdt oxy hda trfn bk mdt difn cyc bdng 0. Sau dd sS giam ddn do n6ng df chdt oxy hda ngay sdt b l mdt difn cue bj bift tieu. Sau khi chdt khu dugc t^o thdnti, qud trinh quet thf sS ddo ngugc lai (quft theo chieu duong). Mdc dCi quet theo chieu duong nhimg ddng catod van cdn tir qud trinh kbit, khi difn cyc trd thdnh chdt oxy hod m^uih thi ddng anod xuat hifn vd tdng nhanh cho den khi n6ng dp cua chdt khCr trdn b l m^t difn cyc bdng 0, Mc dd cd dugc ddng anod cyc dai. Sau dd sS giam ddn do chdt khit trong dung dich tai vimg
O R
O
Hlnh 2. D$c trung CV cua he oxy hda khii thudn nghjch O + ne- f2 R.
riep gidp t>l m$t difn cue trift tifu dan bdi phan ling oxy hda. Chu k^* qudt ddu tifn k i t thiic khi difn the quay v l gid t n ban ddu.
Trong hf oxy hda khiir thu§n ngtych, dinh ddng lpc vd Ipa sS ngang bdng nhau. Tuy nhifn, khi tdc dp quft tdng thl dinh ddng lpc vd Ipa sS khdng bdng nhau ntta, vdi mpt tdc dp dii tdn thl he sS trd nSn bdt thuan nghjch. Cdc peak cao trong gian dd tdng tuyin tinh vdi tdc d$ q u ^ vd df ddc ti If vdi hf s6 khuich tdn (Hhih 2). Mpt sy thay dpi ve ndng dp hay chdt difn giai cQng cd thl Idm dnh hudng den k i t qua ciia cdc thf nghifm. Cdc thdng sd can quan tdm khi thyc hifn ph6p do CV: tdc d0 qu^t thl, nhift df thf nghifm, nong dp ctia cdc chdt hay Id sd lugng ion cua chdt difn giai.
Mpt han che ciia CV Id nd sg tao ra difn dung ldn d ddng n l n . Dinh ddng Faradtac phy thufc vdo su trao ddi dien tich vd tdc dd cua cdc chdt oxy hda khti khufch tdn, ndng dd ciia cdc chdt oxy tai t>l m$t sS gidm khi difn t h l cdng dm vd b l mdt difn cyc bj nhiem bdn cQng dnh hudng din hf thdng do.
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Science & Techoloir Vol18,No.T1-ZD15
Phep do CV da dugc tien hanh bdng cdch sii dung mpt thilt bj do difn hda AUTOLAB PGSTAT12/30/302 (Netherlands). Tdt ca cac phep do deu dugc thyc hien trong hf ba difn cue vdi Ag/AgCI/3M KCl lam dien cue so sanh, sgi platinum ldm dien circ ddi va dien cyc sgi nano vang (AuNW) nhu la mdt dien circ lam viec
Tnrdc khi tien hdnh ii vdt probe (qua trinh hdp phu) cdc dien cue phai trai qua qua frinh xCr li ldm sach ban dau bdng each quet thl CV trong H^SO^ 0,005 M (thl dp dung tir 0 V din 1,6 V).
Viing hoat dpng difn hda dupc udc tinh tir peak oxi hoa khOf [25]. Tir dung djch probe (100 nM) da dugc khii ldn ddu tien trong TCEP 10 mM;
dung djch ndy dugc giii d 4 "C khoang 2 gid d l khit cdc lien kit disulfide gicta cdc DNA probe.
Dung djch trdn dupc pha lodng vdi PBS 10 mM (pH 7,4) (PBS dugc su dyng trong tdt ca cdc thi nghifm trir khi cd ghi chit khac). Cdc difn cgc sau khi Idm sach dugc ngdm trong dung djch probe dd qua xit li cd ndng dd thich hgp trong 36 gid. Sau qud trinh it vd hinh thanh ldp don ty tap hgp (SAM), tiln hanh loai bd bdt ky DNA probe ndo khdng dugc hap phu tdt tren b l mdt dien cue bang cdch rira vdi nudc cdt khit ion trong khoang 30 s d nhift do phdng. Tai thdi dilm ndy, DNA probe se gdn vdi be mdt difn cyc thdng qua sy hinh thdnh SAM bdi lifin kit giOa thiol vdi vdng.
Ndng df probe dugc su dyng trong budc ndy Id 500 nM Difn cyc sau khi sita ddi dugc ldm khd
bdng khi nitrogen vd ngay lap nic chuyfn vao dung dich MCH 2 mM ii khoang 2 gid tai nhiet dp phdng, trdnh anh sang [26]. Riia sach MCH hap phy vat ly tren be mat dien cyc bdng nudc khit ion trong 1 phiit d nhiet dp phdng. Nhe nhdng lau khd cdc vet nudc quanh dien cue (khdng sit dung nitrogen). Chuyfn difn cyc tryc tiep vao PBS de bao quan.
De kifim tra hifiu qua hdp phy va giai hap phu chiing tdi tien hanh quet CV trong dung mdi K3Fe(CN)6 0,5 M/ PBS 0,1 M, thl quet tit -0,2 V din 0,6 V. Phuong phdp gidi hdp dupc tiln hanh bdng phep do CV trong PBS 0,1 M, thfi quet tir - 0,2 V din -1,7 V [27]. Tdt ca cac thf nghiem dd dugc tiln hdnh d dieu kifn dp sudt va nhiet dp phdng
KET QUA VA T H A O LUAN
Hieu qua ldm sach dien cyc dupc thdy ro khi cdc chu ki quet thfi sau gdn nhu ldp lai chu ki quet trudc do (Hinh 3), cudng dd ddng peak khir ciia Au tai 0,8 V khi quet thfi trong dung dich H3SO4 cua la -0,67 |iA. Sau khi giai hdp (Hinh 4B), cudng dp ddng peak khii ndy la -2 pA. Dilu ndy giup nhgn thay SAM tren b l mat dien cyc van chua dugc giai hdp hodn toan. Dk cai thifn hieu qua gidi hdp, chiing tdi tiln hdnh Idp lai them vdi chu ky quet thl CV trong PBS. Bfin each do viec thay mdi dung mdi cung Id mdt ylu td kha quan trpng nhdm trdnh hien tugng tdi hdp phu SAM.
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TAP CSi PIAT THIEII KHACH, TAP 18, SO Tl - Z01S
E m VS Ag/AgCI
Hinh 3. Kit qua lam sgch dien cyc trin bang CV [rong 0,005 M H2SO4
Hlnh 4. (A) Dudng qufil the giai hip SAM bdTig phuong phap CV trong PBS vdi ihl ap dgng la tit-0.2 V d i n - V. (B) Ket qua lam sach difn cyc trong 0,005 M H2SO4 sau khi giai hip
Tai the -1,3 V chiing ta nhdn thdy peak khit giam di ro ret sau vdng quet ddu tifin, d gia trj thl ndy cac probe sS dugc gidi hap khdi b l mdt difin cyc. Tai thdi diem ndy phdn tu luu huj-nh^nhdn
Au-S se bj pha vd (*). Qua trinh nay bdt ddu difin ra tai nhung vi tri cd mat dp probe thap, nhihig khuyfit hdng d be mdt dien cyc. Thdi gian dien ra nhanh hay ch§m Id tuy thupc vdo dd ddng nhdt vfi mft dien til- tir dien cyc vdng va do dd lifn kit cau triic, mite dp khuyfit hdng be m$t
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ScieiGi S TechHiBil HvilMiiit, Vil 18, NB.TV Z015
3.0x10*
2.0x10*
1.0x10*
E (V) VS Ag/AgCI
Hlnh 5. Hifu qua giai hdp ldp m^ng SAM. (A). (B) cudng dO ddng khi g3n probe Idn 1 va lan 2; (C), (D) cudng df ddng khi Iai vdi myc tieu lan 1 vd ldn 2.
Hieu qua gdn prol>e sau khi tdi sir dyng difn cyc (B) dat dugc 95,2 % iing vdi cudng dp ddng dat 2,08 pA so vdi cudng dp ddng gdn probe ldn ddu tifn (a) (2,12 pA). Qua dudng (C) vd (D) ta thdy hifu sudt lai hda ldn 2 giam cdn khoang 82,8 % so vdi ldn 1 (Hinh 5). Vdi nhihig kit qua ban ddu chiing tdi nhan thdy qud trinh hdp phu vd gidi hdp phu tudn theo co che thufn nghjch nhu trong phuong trinh (*).
K E T LUi^N
Qua nghifin cim ndy cho thdy mft co sd cho vifc tai sir dung/ cdu tnic lai bf m^t cdm bien
sinh hpc bdng cdch sit dung qud trinh khii gidi hdp bdng difn hda de loai bd cdc phdn tu SAM gdn kit vdi be mdt difin cue. Bdng vifc sii dyng phuong phdp difn hda quet t h l vdng tudn hodn (CV) chung tdi cd the khdo sat dugc qud trinh hdp phy vd gidi hap phy cua SAM tren difn cyc AuNW. Ben canh dd phuong phdp ndy cQng cd thl dp dyng cho nhihig loai cam bien cd sit dyng lien kit Au-S de cd dinh cac thu thl sinh hpc.
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TAP CII PIAT THEN KKCII, TAP 18, SO n - Z015
Study on the adsorption and desorption process of thiol-modified probe on gold electrodes using cyclic voltammetric method
• Cao Huu Tien
• Nguyen Quang Thien
• Ha Van Linh
• Le Van Hieu
University of Science, VNU-HCM
ABSTRACT
In this paper. we study the electrochemical properties of self- assembling monolayers (SAM) created by the adsorption and desorption of the alkanethiol at the gold nanowires (AuNW) electrode surface by using the CV method.
The analysis results show that the effectiveness of the desorption process reached over 90 %. The adoption of a
Key words: SAM. biosensor, CV, adsorption,TAI LIEU THAM K H A O
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[2]. V. Sato, B.L. Frey, R M. Com, K. Uosaki, Polarization modulation fourier transform infrared studies of the effects of self- assembly time on the order and orientation of 11-Ferrocenyt-l- undecanethioI Monolayers on Gold, Bulletin of the Chemical Society of Japan, 67(1), 21- 25(1994).
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Scieice ft Teclioloiy leveloiHil, Vol 18, No.TI- Z01S
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[9]. C J . Zhong, ). Zak, M.D. Porter.
Voltammetric reductive desorption characteristics of alkanethiolate monolayers at single crystal Au (111) and (1)0) electrode surfaces. Journal of Electroanaiylical Chemistry, 421, 1, 9-13 (1997).
[10]. C J . Zhong. N.T. Woods, O.B. Dawson.
M.D. Porter, Formation of thiol-based monolayers on gold: implications from open circuit potential measurements.
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TAP Cii PIAT TRIEN KISCH, TAP 18, SO Tl - 201S [21]. T. Kakiuchi, H. Usui, D. Hobara, M.
Yamamoto, Voltammetric properties of the reductive desorption of alkanethiol self- assembled monolayers from a metal surface, Lan^uir, 18, 13, 5231-5238 (2002).
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Electrochemical oxidative adsorption and reductive desorption of a self-assembled monolayer'of decanethiol on the Au (111) surface in KOH ethuiol solution. Journal of Electroanalytical Chemistry, 550, 321-325 (2003).
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Zimmermann, D.T. Wong, CM. Ho, Elecfrochemical detection of low-copy number salivary RNA based on specific signal amplification with a hairpin probe.
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