Supporting Information

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1 Supporting Informtion Positive Potentil Opertion of Cthodic Electrogenerted Chemiluminescence Immunosensor bsed on Luminol nd Grphene for Cncer iomrker Detection Shoujing Xu, Yng Liu*, Tihong Wng*, Jinghong Li* Deprtment of Chemistry, Key Lb of ioorgnic Phosphorus Chemistry & Chemicl iology, Tsinghu University, eijing 184, Chin b College of Mterils Science nd Engineering, nd Key Lbortory for Micro-Nno Optoelectronic Devices of Ministry of Eduction, Hunn University, Chngsh 4182, Chin Synthesis of Grphene. The grphene ws synthesized ccording to our previous works 4, 41. Grphene oxide ws first synthesized from commercil grphite powder by the Hummers method 52. riefly, grphite powder ws oxidized by the mixture of concentrted H 2 SO 4, K 2 S 2 O 8 nd P 2 O 5 to produce pre-oxide grphite. Then, the pre-oxidized grphite ws re-oxidized by concentrted H 2 SO 4 nd KMnO 4 to get grphene oxide products. Grphene ws prepred by dding hydrtion hydrzine into the fresh-synthesized grphene oxide solution. Finlly, blck hydrophobic powder grphene ws obtined by filtrting the production nd drying in vcuum. The finl products were chrcterized by TEM, XRD nd Rmn spectr (Figure S1 in Supporting Informtion). Preprtion of Gold Nnorods (GNRs). The preprtion of gold nnorods ws reported elsewhere In brief,.6 ml of 1 mm freshly prepred ice-cold NH 4 solution ws dded to the mixed solution of.25 ml 1

2 HAuCl 4 3H 2 O (1 mm) nd 7.5 ml cetyltrimethylmmonium bromide (CTA) (1 mm) to get ple brown gold seed solution. To prepre gold nnorods growth solution,.25 ml of 1 mm AgNO 3 nd.27 ml of 1 mm scorbic cid solution were dded to mixed solution of 4 ml CTA (1 mm) nd 1.7 ml HAuCl 4 3H 2 O (1 mm) successively. Finlly,.42 ml of gold seed solution ws dded to bove-mentioned growth solution, nd llowed to rect for 15 h t 28 o C before centrifugtion. The resulting GNRs were chrcterized by trnsmission electron microscopy (TEM) nd UV-Vis spectr (Figure S2 in Supporting Informtion). Sttisticl nlysis of them shows tht the nnorods hd n verge dimeter of bout 13 nm nd n verge length of bout 4 nm with n spect rtio of ~3.. Preprtion of Ab 2 nd GOx conjugted gold nnorods (GOx-GNR-Ab 2 ). The conjugtion of gold nnorods with GOx nd Ab 2 is shown in Scheme 1A. efore using, the fresh gold nnorods solution ws centrifuged t 14 rpm for 1 min, nd the products t the bottom were then redispersed in PS solution. The bove procedure ws conducted twice to remove extr free CTA. After tht,.1 ml of 1 mg/ml PSS ws dded into 5 ml of the redispersed GNRs solution nd left for 2 min to get PSS functionlized gold nnorods. Then, the mixture ws centrifuged t 8 rpm to get rid of excessive PSS nd the pellet ws redispersed in PS solution (ph 7.4, 1 mm). To obtin GOx-GNR-Ab 2, the colloidl solution of PSS cped GNRs ws dded into the solution of GOx nd Ab 2. The mixed solution ws left to rect for 4 min under shking, nd ws centrifuged to remove extr GOx nd Ab 2 nd redispersed in PS solution. After tht, 3% (w/v) SA solution s blocking solution ws dded into bove GOx-GNR-Ab 2 solution to block extr ctive sites. Finlly, the solution ws centrifuged t 7 rpm to get rid of the superntnt followed by redispersed in PS solution to GOx-GNRs-Ab 2 solution. 2

3 15 Intensity(.u.) Intensity (.u.) C Rmn(cm -1 ) / degree Figure S1 TEM (A), Rmn spectr () nd XRD pttern (C) of the s-synthesized grphene. (A) () Aborbnce Wvelength / nm Figure S2 TEM (A) nd UV-vis spectr () of the s-prepred gold nnorods. 3

4 c b Wvelength / nm Figure S3 Chemiluminescence of luminol on bre GC electrode (curve,.5 V vs. Ag/AgCl, ph 1.5) nd GR-CHIT/GC electrode t.5 V (curve b) nd.1 V (curve c) b Potentil / V (vs. Ag/AgCl) Figure S4 ECL behvior of luminol on grphene oxide-chitosn (curve ) nd grphene-chitosn (curve b) modified GC electrode. 15 A Current / Potentil / V (vs. Ag/AgCl) Potentil / V (vs. Ag/AgCl) Figure S5 ECL behvior (A) nd CV curve () of luminol on GR-CHIT/GC electrode. 4

5 Time / s Figure S6 ECL-time curve of GR-CHIT/GC electrode under continuous cyclic voltmmetry scn in.1 M PS solution (ph 7.6) contining 1 μm luminol t scn rte of 1 mv s -1. Z im ( ) e b c d Z re ( ) Figure S7 Cyclic voltmmogrms (A) nd electrochemicl impednce spectr () of bre GC electrode (), GR-CHIT/GCE (b), Ab 1 /GR-CHIT/GCE (c), PSA/Ab 1 /GR-CHIT/GCE (d) nd GOx-GNR-Ab 2 /PSA/Ab 1 /GR-CHIT/GCE (e) in 5mM [Fe(CN) 6 ] 3-/4- contning.1 M KCl solution (ph 7.). Scn rte ws 1 mvs -1. The impednce spectr were recorded from.1 Hz to 1 khz with signl mptitude of 5 mv t the forml potentil of the redox couple. 5

6 Figure S8 Scnning electronic microgrphs of PSA/Ab 1 /GR-CHIT/GC before () nd fter (b) the incubtion in the colloidl solution of gold nnorods conjugted with GOx nd Ab 2. The inset is the high-resolution imge A Incubtion time / min [GOx] /[Ab 2 ] Figure S9 (A) The ECL intensity s function of immunorection time in the presence of 1 ng/ml PSA. () The influence of mss rtio of GOx to Ab 2 in the preprtion of GOx-GNR-Ab 2 on the ECL intensity of the immunoelectrode in the presence of 5 ng/ml PSA. All experiments were crried out in.1 M PS solution (ph 7.6) contining 1 μm luminol nd 1 mm glucose. The ECL intensity ws obtined t.5 V vs Ag/AgCl. The error brs were clculted from three times prllel experiments. 6

7 ECL Intensity (. u.) b Potentil V vs. Ag/AgCL Figure S1 The ECL behvior of GR-CHIT modified electrode in the bsence () nd presence (b) of luminol in PS solution (ph 7.6) 7