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1 Detection of Epstein Barr virus infection in cancerby using highly specific nanoprobe based on dBSAcapped CdTe quantum dots Yilin Li,abLihong Jing,bKe Ding,bJing Gao,aZhi Peng,aYanyan Li,aLin Shen*aand Mingyuan Gao*bBy using denatured bovine serum albumin (dBSA) molecules as multidentate ligands, surface-functionalizedCdTe quantum dots (QDs) were synthesized and further evaluated for preparingfluorescent probes capableof detecting Epstein Barr virus infection. The optical studies revealed that the dBSA coating effectivelyimproved the optical stability of the resulting QDs The interaction between CdTe@dBSAand the membrane of carcinoma cells further suggested that the nonspecific adsorption of CdTe QDsstabilized by thioglycolic acid (TGA) was significantly eliminated upon the dBSA coating.

2 Streptavidin wascovalently conjugated to CdTe@dBSA to endow the QDs with specific binding affinity in detecting theserum level of anti-Epstein Barr virus (EBV) capsid antigen IgA (VCA-IgA) in nasopharyngeal carcinomapatients for early screening and diagnosing EBV-associated IntroductionQuantum dots (QDs) have been attracting considerable atten-tion owing to their unique electronic and optical 3As the spectral features of QDs in comparison with that ofconventional dyes is characterized by narrow, symmetric, andparticle size-dependent uorescence, as well as a broad excita-tion range and excellent robustness against photobleaching,QDs are ideal on many counts for bioassay and 7 However, due to the strong surface defect-sensitive uores-cence.

3 The emission intensity of QDs only coated with a smallthiol ligand, such as thioglycolic acid (TGA), mercaptopropionicacid (MPA), and so forth, is readily reduced in 10 Most importantly, the surface of the QDs isalways prone to adsorbing biomolecules nonspeci cally,11 13which sharply decreases the speci city of the QDs-basedmolecular probes, and thereby limits the bioapplication of strategies for surface modi cation on QDs have beendeveloped to overcome these 20 However, the majorstrategies, such as the encapsulation of QDs with silica,21 23organic polymers,24 27or amphiphilic polymers15,17,18.

4 20haveconcentrated on passivating the surface defects of QDs therebyimproving their uorescence ,28 Limited approacheshave been developed to suppress the nonspeci c interactionbetween QDs and biomolecules. Polyethylene glycol (PEG)remains the most commonly used material to modify the surfaceof nanoparticles for suppressing their non-speci c adsorptionwith ,14,29 Owing to the antifouling property ofPEG, the binding speci city of PEG-coated particle probes can beeffectively improved for immuno uorescence ,30 Even forin vivoapplications, PEG-coated QDs also exhibit reducednonspeci c accumulation in the reticuloendothelial tissues,which is bene cial for increasing the accumulation of QDs atregion of addition.

5 Multidentate PEG ligands havealso been developed for effectively passivating the 19,31 However, the preparation of multidentate PEGligands requires sophisticated synthetic procedures. Moreover,the functional groups, typical at the end of the PEG molecules,limit the number of bioligands to conjugate to PEGylated QDs,and thereby their potential in bioassays and bovine serum albumin (dBSA) has recently beenused to manipulate the surface chemistry of ,33 Preparedby chemically reducing disul de bonds in BSA, dBSA contains37 thiol groups per monomer, thereby affording them greatpotential as excellent multidentate ligands for from that, BSA is also a well-known blocking reagent toreduce nonspeci c adsorption in immunoassays similar 36It is therefore reasonable to expect that dBSA as mul-tidentate ligands can not only passivate surface defects andimprove the optical properties of QDs.

6 But also suppress theaKey Laboratory of Carcinogenesis and Translational Research (Ministry of Education),Department of GI Oncology, Peking University Cancer Hospital and Institute, FuchengRoad 52, Haidian District, Beijing 100142, China. E-mail: +86-10-88196561bInstitute of Chemistry, the Chinese Academy of Sciences, Bei Yi Jie 2, Zhong Guan Cun,100190 Beijing, China. E-mail: Tel: +86-10-82613214 Electronic supplementary information (ESI) available: The uorescenceintensity and peak position of CdTe@dBSA with different dBSA : QD ratios, andthe optical stability of CdTe@dBSA by optimal dBSA : QD ratio in 1 PBS arerespectively shown in Fig.

7 S1 and S2; the uorescent stability of CdTe@dBSA streptavidin bioconjugate are shown in Fig. S3. See this:RSCAdv.,2014,4, 22545 Received 15th March 2014 Accepted 1st May 2014 DOI: journal is The Royal Society of Chemistry 2014 RSCAdv.,2014,4, 22545 22550 |22545 RSC AdvancesPAPER nonspeci c interaction of QDs with biomolecules. Moreover,compared with PEG polymer, dBSA contains different func - tional groups such as amino and carboxyl groups along thepolymer backbone, which provides more reactive sites forconjugation with bioligands to achieve QD-based probes forboth bioassay and this rationale, dBSA was chosen to encapsulateCdTe QDs stabilized by TGA in this study.

8 The optical propertiesof the resulting CdTe@dBSA were systematically studied. Theinteraction between CdTe@dBSA and carcinoma cells was thenstudied to investigate the resistance of dBSA coating to non-speci c binding. Furthermore, the CdTe@dBSA streptavidinconjugate was synthesized and used as a molecular probe todetect Epstein Barr virus (EBV) infection in nasopharyngealcarcinoma (NPC) patients. EBV, as a member of the herpes virusfamily, has been implicated in the occurrence of variouscancers, such as NPC, Burkitt's lymphoma, gastric carcinoma,and so er infected by EBV, patients always exhibitelevated level of anti-EBV antibodies in sera, particularly theanti-EBV capsid antigen IgA (VCA-IgA).

9 37 39 Therefore, detectionof the serum level of VCA-IgA has served as an effective strategyfor early screening and diagnosis of EBV-associated ,39In this paper, the application of the CdTe@dBSA streptavidinconjugate in detecting the serum level of VCA-IgA in NPCpatients was also demonstrated through indirect immuno uo-rescence Materials and ChemicalsAluminum telluride (Al2Te3, ), cadmium perchloratehydrate (Cd(ClO4)2$6H2O, ), thioglycolic acid (TGA, 97+%),1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC, +%),N-hydroxysulfo-succinimide sodium salt (Sulfo-NHS, ),were all purchased from Sigma-Aldrich.

10 Agarose (BiowestAgarose) was purchased from Gene Tech. Co., Ltd., Shanghai,China. Streptavidin was purchased from Promega Biotech Co.,Ltd. Bovine serum albumin (BSA, Amresco, 99%), acrylamide(AM, Amresco, 99+%),N,N0-methylenebis(acrylamide) (MBA,Amresco, 99%), ammonium persulfate (APS, Amresco, 95%),N,N,N0,N0-tetramethylethylenediamin e (TEMED, Sigma, 99%),tris(hydroxymethyl)aminomethane (Tris, Amresco, ),glycine (Gly, Amresco, ) were distributed by BiodeeBiotechnology Co., Ltd., Beijing, China. Goat anti-humanimmunoglobulin A antibody (antiIgA Ab) was gi ed from theNational Institute for Viral Disease Control and Prevention,China CDC.