Nanoparticle-assisted chemiluminescence and its applications in analytical chemistry

Trends in Analytical Chemistry, Vol. 29, No. 10, 2010

Trends

Nanoparticle-assisted chemiluminescence and its applications in analytical chemistry Dimosthenis L. Giokas, Athanasios G. Vlessidis, George Z. Tsogas, Nicholaos P. Evmiridis This comprehensive, critical review summarizes the use of nanoparticles in enhanced and amplified chemiluminescence detection, illustrated by different reaction strategies, electrogenerated chemiluminescence sensors, immunoassay or hybridization labels and electrogenerated chemiluminescence immunoassay or hybridization sensors. We discuss the analytical applications on the basis of validity, range and sensitivity, and draw some useful conclusions about the most sensitive approach in each type of application. ª 2010 Elsevier Ltd. All rights reserved. Keywords: Catalysis; Chemiluminescence; Electrochemiluminescence; Hybridization assay; Immunoassay; Nanoparticle; Quantum dot; Reaction strategy; Sensitivity; Sensor

1. Introduction Dimosthenis L. Giokas, Athanasios G. Vlessidis*, George Z. Tsogas, Nicholaos P. Evmiridis Laboratory of Analytical Chemistry, Department of Chemistry, University of Ioannina, 45110, Ioannina, Greece

*

Corresponding author. Tel.: +30 2651008401; Fax: +30 2651008781; E-mail: [email protected]

The unique properties of nanomaterials, which are between those of bulk materials and molecular species, open new opportunities for applications [1–4]. Apart from the general properties (e.g., optical, mechanical, thermal, size and shape), transition-metal nanoparticles (NPs) show catalytic properties for redox reactions, intense size- and shape-dependent catalytic action, controllable charge-transfer events and compatibility with biomaterials. Such properties can be employed to increase the kinetics and the selectivity of chemical reactions as well as to enhance response signals of instrumental methods of analysis. For example, size-dependent catalytic action results in selective reaction, strong catalytic activity in redox reactions offers rapid detection, and biocompatibility provides convenient conjugation to biomaterials. Chemiluminescence (CL) and electrogenerated chemiluminescence (ECL) in aqueous phase systems are generated mainly by redox reactions with proper CL reagents and/or electrochemical-reaction conditions. In batch or flow reactors, the reaction process involves formation of intermediate radicals that result in unstable products that

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decompose to form electronically-excited molecules that deactivate with CL emission. This process can take place either directly (i.e. direct oxidation of the target analytes to produce emitting species) or indirectly {i.e. exploiting the enhancing or inhibitory effects on the CL emission of certain luminescence compounds [e.g., luminol, lucigenin, RuðbpyÞ3 2þ , or pyrogallol} [5–8]. In contrast to bioluminescence systems, the CL emission generated during oxidation of organic molecules is of relatively low intensity due to low quantum yield. Enhancement of CL emission is therefore necessary for application in trace analysis in analytical chemistry. Compounds or reaction strategies that can act as signal enhancers have been proposed for that purpose (e.g., surfactants, hydroxylamine, transition elements, and solvent pre-oxidation) [9–13]. In that sense, metal NPs with their unique redox catalytic properties have gained increasing attention in recent years as a novel alternative to catalyze redox CL reactions, under proper conditions, providing amplified CL emission. Analytes can either enhance or inhibit the NP-induced enhanced CL signal, thus providing the basis for development of a number of sensitive methods of analysis.

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On the other hand, NPs used as labels conjugated to biomolecules or linked to organic molecules to form complexes with the analyte can offer amplified methods by dissolution, followed by CL detection. Moreover, in cases that NPs are used as scaffolds that carry a number of labels (e.g., enzymes or even other NP conjugates or NPs coated by a shell of metal that is more soluble in a less acidic environment), CL detection becomes extremely amplified. Finally, highly-selective determinations can be achieved through coupling of CL detection with enzymatic reactions, bio-affinity or DNA-hybridization reactions. Several reviews gathering the latest advances in nanomaterial science have appeared, but there have been no reviews related to the analytical areas of application, especially with regards to CL analysis. Two reviews have appeared most recently:  Krull et al. [14] deals with the application of quantum dots (QDs), gold NPs (AuNPs), and molecular switches to optical nucleic-acid diagnostics, with emphasis in the areas of: (a) QDs and fluorescence-in situ-hybridization microarrays, fluorescence resonance energy transfer (FRET) and signal enhancement; (b) AuNPs or silver NPs and colorimetric detection, fluorescence quenching, chemiluminescence and electrochemiluminescence; and, (c) advances in the design of molecular beacons, hairpin probes and other diagnostic nucleic-acid constructs.  Chang et al. [15] deals with DNA-functionalized AuNPs for bio-analysis in colorimetry, fluorescence and lightscattering detection of bio-polymers and small solutes, and highlights the effects of size and concentration of AuNPs, length and sequence of DNA, the nature of capping agents, ionic strength, and pH on the sensitivity and specificity of the nano-sensors for the analytes. In this review, we describe the recent trends in the analytical application of NP-assisted CL-reaction strategies and ECL sensors, developed for enhanced and amplified CL detection. We review highly-sensitive methods of analysis proposed to be applied in trace analysis, food chemistry, pharmaceuticals, and biochemical analysis. In addition, we present CL immunoassays and hybridization assays of specific interest in clinical analysis and medical diagnosis (e.g., protein or antibody determinations, DNA analysis, DNA-sequence recognition and single-nucleotide-polymorphism analysis). 2. NP-assisted CL-detection strategies 2.1. CL-reaction studies Although procedures for enhanced CL emission are obtained through different reaction strategies (e.g., catalytic processes, energy transfer, or redox reactions), only a limited number of these studies has been employed for analytical purposes. 1114

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Catalytic reactions that produced a CL reagent followed by further CL reaction were initially employed by Shi et al. [16] for the detection of NH3, based on the oxidation of NH3 to NOx by the catalytic action of perovskite NPs and further oxidation of NO with O3. Another example, reported by Lan et al. [17] for the determination of glucose, employed the conversion of glucose to D-gluconic acid with the production of H2O2 through the enzymatic action of glucose oxidase (GOD). H2O2 then reacted with added luminol in the presence of horseradish peroxidase (HRP) enzyme to generate the enhanced CL signal. Both enzymes being adsorbed on AuNPs were attached on the inner wall-surface of a glass tube that was placed in-line with a flow-injection (FI)/ CL-detection system of analysis. Reactions taking place on the surface of catalysts with production of electronically excited product molecules (cataluminescence reaction) provide another strategy of generating CL emission by direct deactivation of the molecule from the excited to the ground state or by energy transfer to a fluorescent molecule of high quantum yield. The analytical utility of the latter strategy has been explored for the detection of ethanol after its conversion to electronically-excited acetaldehyde via the catalytic action of Eu3+-doped ZrO2 NPs, followed by energy transfer of the deactivated molecule to Eu3+ fluorescent cation [18]. The use of fluorophore molecules is another popular strategy in CL-based assays amplified by the presence of NPs. Non-fluorescent electronically-excited molecules deactivate to the ground state without light emission unless FRET takes place to a nearby fluorescent element. Such energy transfer takes place when the excited energy state of the donor molecule is close to the energy state of an unoccupied orbital energy state of acceptor molecule. Based on this principle, a fluorimetric sensor prepared by Karim and Lee [19] based on Tb3+-acetylacetone complex NPs was applied to the determination of enoxacin via UV excitation of enoxacin and further deactivation through FRET to Tb3+-acac complex. One of the most important features of metal NPs is their ability to catalyze various redox reactions taking advantage of their unique chemical and electrochemical properties (Table 1a). The catalytic activity of Au-colloid and other metal-colloid NPs of the precious elements group (e.g., silver and platinum) on the gas-phase and liquid-phase redox CL reactions has been established, and has become an expanding field of research. The mechanism of the CL reaction of luminol-H2O2 CL reagent was found to be the following [20]: AuNPs

H 2 O2 ! 2HO ðCatalytic reactionÞ HOO þ HO ! O 2 þ H 2 O ðRadical reactionÞ LH  þ HO ! L þ H 2 O ðRadical propagation reactionÞ 2 L þ O ! AP2 þ hm ðCL  reactionÞ 2 ! LðOOÞ ! AP

Method

Analyte

Range (M)

LOD (3r)

RSD (%)

Application

Ref.

Metal-nanoparticle catalysis 5.0 · 109–2.0 · 106 5.0 · 109–5.0 · 106 5.0 · 109–5.0 · 106 2.0 · 108–5.0 · 106 1.0 · 109–5.0 · 106 2.0 · 109–2.0 · 106 2.0 · 109–2.0 · 106

1.7 · 109 2.0 · 109 1.9 · 109 8.4 · 1010 1.9 · 1010 9.3 · 1010 6.4 · 1010

FI/CL-detector Enhanced Luminol-H2O2-AuNPs

Norfloxacin Ciprofloxacin Lomefloxacin Fleroxacin Ofloxacin Levofloxacin

2.5 · 107–4.0 · 106 3.9 · 108–4.0 · 106 4.0 · 108–4.0 · 106 7.6 · 108–4.0 · 106 5.5 · 108–2.8 · 106 2.8 · 108–4.0 · 106

1.0 · 108 2.9 · 108 2.0 · 108 2.4 · 108 2.2 · 108 2.2 · 108

Luminol-H2O2 citrate-coated Pt-colloid NPs inhibited CL

Ascorbic acid Catechol Pyrogallol Phloroglucinol L-tyrosine Tannic acid Gallic acid Uric acid

2.5 · 107–1.0 · 104 1.0 · 108–5.0 · 106 1.0 · 108–2.5 · 106 1.0 · 107–1.0 · 105 5.0 · 108–1.0 · 105 5.0 · 108–1.0 · 105 2.5 · 107––1.0 · 105 5.0 · 108–1.0 · 105

3.6 · 108 2.0 · 109 1.9 · 109 4.4 · 108 1.0 · 108 8.5 · 109 5.0 · 108 1.0 · 108

CL detector Luminol-H2O2-AuNPs (QCL-emission)

Estrone Estradiol Estriol

0.07 · 106–7.0 · 106 0.04 · 106–10.0 · 106 0.10 · 106–10.0 · 106

3.2 · 109 7.7 · 109 49 · 109

2.9% (n = 11 at 1.0 · 106) 2.6% (n = 11 at 1.0 · 106) 1.8% (n = 11 at 1.0 · 106)

CL-detector Luminol-H2O2-Fe2O3 colloid NPs

H2O2

2.0 · 103–0.1 (LogCL vs CH2O2) 2.5 · 105–2.5 · 103 2.5 · 105–2.5 · 104 2.5 · 104–1.9 · 103 2.5 · 105–6.25 · 104 1.25 · 104–6.25 · 104 8.25 · 104–2.25 · 103 2.5 · 104–2.25 · 103

1.25 · 103

6.2% (n = 5 at 2.0 · 103)

1.0 · 105–1.0 · 103 0.1 · 106–1.0 · 106 1.0 · 106–10.0 · 106 10.0 · 106–30.0 · 106

5 · 106 30 · 109

<2% (n = 9 at 1.0 · 105) 1.45 (n = 6 at 0.5 · 106) 0.96 (n = 6 at 5.0 · 106) 1.0 (n = 6 at 30.0 · 106)

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Vitamin E Cysteine Tyrosol Catechin Chlorogenic acid Pyrogallic acid Caffeic acid Ascorbic Acid Gallic Acid CL-flow biosensor GOD + HRP + AuNPs Luminol-hydrazine-AuNPs

Glucose Hydrazine

[20]

9 4:3 > > > 1:5 > > > = 1:9 ðn ¼ 11Þ 1:3 > > > > 1:6 > > ; 2:1

Human urine sample fluoroquinolones Recovery: 92% (n = 5)

[23]

[21]

Estrogen urine samples RSD: 1.6–3.5% Pharmaceutical formulations

[24]

[22]

Glucose in serum %RSD: 1.3% Boiler feed water NH2NH2 RSD: 9.0%, Recovery: 99%

[17] [27]

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Catechol Ascorbic acid Adrenalin Noradrenalin Dopamine L-hystidine L-cysteine

CL-detector AuNPs catalyzed Luminol

Trends in Analytical Chemistry, Vol. 29, No. 10, 2010

Table 1a. Analytical parameters of metal-nanoparticle-catalyzed CL-detection methods

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[19] Pharmaceutical formulations Urine samples 1.35% (n = 6 at 1 · 104)

[16] NH3 in environment

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Surface NPs of low coordination were suggested to be responsible for the catalytic activity of AuNPs. Some examples with potential analytical utility following this field of research are studies of the effect of Au-colloid NPs on the luminol-H2O2 (model CL reaction) and the inhibition effect of several organic compounds containing -OH, -NH2 and -SH groups [20] and the catalytic action of Pt-colloid NPs on luminol-H2O2 (model CL reaction) when prepared by citrate reduction [20,21]. In all cases, it was suggested that the CL enhancement was due to the ability of metal NPs to form surface radicals of OH from the disruption of the O–O bond (H2O2) and to facilitate the electron-transfer process between the radical intermediates formed. In a similar approach, Fe2O3-colloid NPs were shown to exhibit catalytic activity on the luminol-H2O2 model CL reaction [22], which was modified in the presence of various oxidant and antioxidant compounds of analytical interest. The step-wise reaction pathway follows the Weis reaction in the first step and FentonÕs reaction in the last step, as follows:  FeðIIIÞFe2 O3 þ HO 2 ! FeðIIÞ þ HO2 ðWeis reactionÞ

HO2 þ OH  ! O 2 þ H 2 O ðRadical propagationÞ

3 · 108 2 · 107–1 · 104 Enoxacin PL-detector Tb-acac NPs

15 mg L1 1.0 · 103–12.4 · 103 12.4 · 103–24 · 103 Ethanol ETCTL- detector Eu3+-doped ZrO2 NPs

8.2 · 107

  O 2 þ L ! LðOOÞ ! AP2 þ N 2 þ hm ðCL-reactionÞ

2.3 · 106–0.6 · 103 NH3 Other catalytic reactions Nano-converter LaCoO3 NPs

1.1 · 10

[18]

[26] Tap water (catechol) Synthetic samples 2.9% (n = 11, 1 · 109) 2.3% (n = 11, 5 · 109) 1.7% (n = 11, 5 · 109) 0.9 · 1012 1.9 · 1011 7.5 · 1011 3.6 · 1012–5.4 · 109 7.3 · 1011–9.1 · 109 1.8 · 1010–3.6 · 108 FI/CL-detection Luminol-NaIO4-AuNPS

Catechol Hydroquinone Resorcinol

2.9 · 1010 1.1 · 109 1.0 · 109–7.0 · 106 4.0 · 108–7.0 · 106 L-cysteine 6-Mercaptopurine

2.1% (n = 11 at 1.0 · 106) 1.8% (n = 11 at 7.0 · 107)

[25] G-MP in medicine tablets GSH in human serum Recovery:90% 7.0 · 10 –1.0 · 10 Glutatione

8 7

CL-detector Luminol-H2O2-AuNPs

Method

Table 1a (continued)

Analyte

Range (M)

5

LOD (3r)

RSD (%)

6

Application

1.4% (n = 11 at 1.0 · 10 )

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Ref.

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FeðIIÞ þ H 2 O2 ! HO þ HO þ FeðIIIÞ ðFenton reactionÞ The analytical utility of the catalytic effect of NPs on CL-reaction systems stems from the fact that the CL intensity of the above reactions can be enhanced or inhibited by several analytes and this effect can be employed for developing CL methods of analysis. Based on this principle, several methods have been proposed, such as the determination of:  fluoroquinolone derivative-antibiotics in pharmaceutical preparations and human urine through the strong enhancement effect of the CL signal [23];  estrogens in pharmaceutical preparations and human urine [24];  glutathione, L-cysteine, and 6-mercaptopurine [25];  catechol, hydroquinone, and resorcinol by luminolIO4  reaction CL system [26]; and,  hydrazine in boiler feedwater based on AuNPmediated catalytic oxidation of hydrazine by dissolved O2 in aqueous solutions [27]. Nevertheless, several exceptions to the catalytic action of metal NPs have been observed {e.g., the lack of effect on pyrogallol oxidation by periodate or the quenching effect of NP materials coated with cationic surfactants (i.e. CTAB) as stabilizers [28]}. In all the above studies, sensitivity is the major benefit from the involvement of metal NPs in CL reactions. However, routine samples include interfering ingredients that, in some cases, can seriously modify the response signal. To secure selectivity for a CL-analysis method,

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especially during in vivo analysis, it is often necessary to restrict the access of interfering species into the reactor environment. Except conventional separation procedures (e.g., delay coils, chromatographic columns or capillary electrophoresis), which continue to be major techniques of choice to resolve selectivity barriers of CL detection, NPs could offer promising options. On this basis, a nano-reactor made of porous nano-shells of Caphosphate was devised by Wingert et al. [29]. These nano-shells were hollow and filled with luminol-H2O2 solution that contained hematin-fluorescein as a fluorescent system. The nano-reactor was prepared by coating a phospholipid liposome with a 1-nm thick layer of Ca-phosphate grown on the outer surface. 2.2. Immunoassay and hybridization labels Metal NPs by themselves do not have target-recognition abilities for selectively binding analytes of interest. Antibodies, proteins, short-chain organic acids, aminothiols and DNA are used to functionalize the surface of metal NPs to obtain target-recognition sites. These NPsurface-functionalization agents are conjugated through electrostatic attraction or covalent bonding. The electrostatic attraction technique is easy and quite simple, but the attachment is not strong enough, unlike that of the covalent bonding technique, which is strong enough. Aptamers act as antibodies in target-recognition processes depending on their sequence and conformation. They can fold into complex three-dimensional configurations forming binding pockets and clefts for specific recognition of ions, small and large molecules, proteins and complexes, cells, viruses and parasites. In addition, aptamers possess high structural stability and high affinity, and are amenable to structural modification. 2.2.1. CL immunoassays (CLIAs). Assays based on CLIAs commonly rely on indirect approaches based on labeling strategies involving the use of chemiluminescent reagents [e.g., isoluminol derivatives, acridan esters, and tris(2,2_-bipyridyl)ruthenium(II)] and enzymes (e.g., peroxidase, alkaline phosphatase, glucose oxidase, and oxidoreductases) [30,31]. Although very successful, the labeling procedures often involve multi-step reactions and further purification of the reaction products, which are complicated, tedious, time consuming and costly. Furthermore, enzyme labels are unstable under CL-detection-analysis conditions and prone to interference from inhibitors present in the sample. The method of labeling with CL reagents is also intricate and can cause protein denaturation and/or induce partial loss of enzymatic activity. In several cases, the use of signal enhancers is necessary to increase the reaction efficiency and to amplify the CL signal, but the analytical features vary with different enhancers. Another issue worth mentioning is that CLIAs based on CL-reagent systems and enzyme labeling are not rapid

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enough to be suitable for the CL detection of ultra-trace amounts of antigens or antibodies and in situ analysis. Accordingly, some new labeling strategies that exhibit enormous signal amplification have been explored, but they also involve complicated analytical processes, so they still remain a challenge in practical applications. To overcome these issues, metal NPs have been deployed, since they offer certain advantages:  a variety of nanostructures with unique properties at nanoscale dimensions;  good biocompatibility and similar size range to many macro biomolecules;  ability to tailor the NP surface, thus forming conjugates with small molecules and/or biomolecules;  capacity to be used as scaffolds for carrying multifunctional molecules;  ability to act as labels that provide catalytic action to CL (and ECL)-reaction systems; and,  stable conjugates with enzymes with no loss of activity. Metal NPs in chemiluminescence immunoassays (CLIAs) are used mainly as functionalized labels conjugated to antibodies or aptamers. These labels enhance sensitivity or amplify detection (Table 1b). They are stable and do not alter the biological properties of its conjugates. Solid-phase CLIA procedures involve non-competitive or competitive sandwich-type reactions for the formation of specific immunocomplexes based on antigen-antibody or aptamer-bioaffinity systems [32–38]. The NP label is mainly conjugated to the antibody or aptamer moiety of the immunocomplex, whereas the biomaterial analyte that is immobilized on the solid support is an antigen, an antibody or an aptamer (Fig. 1). However, it is important for the resultant conjugate to be adequately characterized for each method and specific biomolecule, because: (1) colloidal particles can also adsorb other components of the chemical system, causing undesired or nonspecific labeling [39]; (2) antibodies can adopt many different orientations due to their numerous functional groups; (3) NP conjugation can affect the protein structure, whilst (4) excessive biomolecule coverage can reduce functionality due to steric crowding [40]. In general, the reports have described AuNPs based on their catalytic action on the oxidation of luminol for enhanced CL-detection signal, obtained by NP oxidative dissolution followed by oxidation of luminol-AuCl4  or another CL system for a super-amplified CL-detection signal [32–37] (Fig. 1) and as scaffolds loaded with other labels (e.g., enzymes or fluorophores) [38]. Amplified CL based on NP-dissolution methods explore the fact that each AuNP contains thousands of Au atoms (e.g., 1.1 · 105 Au atoms are estimated theoretically to be contained in a 15-nm, spherical AuNP), so higher CLemission signals [i.e. lower limits of detection (LODs)] http://www.elsevier.com/locate/trac

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Table 1b. Analytical applications of chemiluminescence immunoassays and hybridization assays Method

Analyte

Range

LOD (3r)

RSD (%)

Application

Ref.

Chemiluminescence immunoassays Amplified CLIA-detection MBs/AuNPs label

hIgG

GFQIA-sensor FITC fluorescent agent

a-fetoprotein

CL-imaging IA Au NP-protein (or chit.) composite(Lum.+H2O2+PIP)

H2O2 rHu-IL-6

1.0 · 106–1.0 · 104 M 2.0–312 pg/mL

2.0 · 107 0.5 pg/mL

3.8% (n = 6 at 3.0 · 105 M) 4.4% (n = 6 at 39 pg/mL)

Serum samples

IA amplified CL-detection Ag on AuNPs (Ag+-K2S2O8-Mn2+)/(H3PO4-luminol)

hIgG

0.02–50 ng/mL

0.005 ng/mL

3.5% (n = 7 at 0.5 ng/mL)

Human serum samples

[33]

IA amplified CL-detection Au-NPs label(AuCl4–Lum.)

goat-anti-human IgG

5.0 ng/mL to 10 lg/mL

1.5 ng/mL

4.2–4.6% (n = 3 at 5–10 ng/mL)

Immuno-serum

[34]

IA amplified CL-detection DC-AuNPs labels

AFP

0.008–0.3 ng/mL

5 pg/mL

Chemical serum samples Recovery: 96.2–103.0%

[35]

IA amplified CL-detection Multi.Ab-Au NP conjugates (Luminol-AgNO3-Au NPs)

hIgG

25–5000 ng/mL

12.8 ng/mL

8.6% (at 100 ng/mL) 7.1% (at 500 ng/mL) 3.0% (at 1000 ng/mL)

[36]

2.0 · 1014–2.0 · 1012 M

4.8 · 1015 M

3.4% (n = 11 at 4.0 · 1014 M)

[45]

3.1 · 1012 0.5 ng/mL

[32]

0.17 nM

HA amplified CL-detection NanoCuS tags (Cu2+ ASV preconcentrated + Luminol)

Target DNA

2.0 · 1012–1.0 · 1010 M

5.5 · 1013 M

HA amplified CL-detection NH2OH extra gold loaded Au NPs Capture target DNA

CL det fi target DNA Nucleotide

0.01–1.0 fM >100 fM

10 a M

Patients serum samples

[38] [37]

[43]

4–11% ((n = 11 at 0.075–1.0 fM))

105 ± 2.1% (at 0.075 fmol) 101.0 ± 5.0% (at 0.25 fmol) 111.0 ± 12.0% (at 1.0 fmol)

[44]

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Chemiluminescence hybridization assays HA amplified FI/CL-detection CuS.NPTarget DNA labeled-DNA.probe-functionalized AuNPs

0.1–100 ng L1

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Figure 1. Non-competitive sandwich-type reactions with amplified or sensitized detection employed in chemiluminescence immunoassays.

can be attained by determining the Au ion. The latter also offers improved selectivity, since it is less prone to interference than enzyme-assisted CL assays. 2.2.2. CL-hybridization assays (CLHAs). After MirkinÕs work on the detection of DNA through hybridization, DNA-functionalized AuNPs acquired widespread acceptance, specifically for single-nucleotide polymorphism (SNP) methods of analysis. Aptamers and DNA enzymes that both contain single-stranded (ss) DNA or RNA are used to prepare bio-conjugates of AuNPs with specific recognition ability for proteins and small analyte molecules (Table 1b). CLHA protocols mainly comprise a sandwich-type non-competitive hybridization reaction that forms biocomplexes between an immobilized molecule on a solid support capture DNA probe, or a target (ss) DNA analyte and a labeled reporter DNA probe, or by a hybridization reaction between target (ss) DNA analyte and DNAprobe-functionalized NP label (Fig. 2) [41–45]. Labels mostly used are metal-colloid NPs (e.g., Au, Ag, and Pt) and inorganic metallic compounds (e.g., CuS).

The metal NPs used are biocompatible and are able to form bio-conjugates with DNA probes without loss of hybridization ability with target (ss) DNA strands. AuNP bio-conjugation to DNA can be easily performed through covalent bonding or electrostatic attraction. Amplified CL detection is obtained by combining the dissolution of the metal NPs (which produces many metal ionic species per label) with the metal-enhanced CL emission during oxidation of CL-reagent systems [41– 45]. However, the dissolution process takes place under unfavorably acid conditions (i.e. highly acidic media), and is time consuming, while the acid conditions vary with the nature of the NP metal; the order of solubility between the different metal colloids used as labels for CLIAs or CLHAs is Cu>Ag>Pt>Au. 2.3. ECL sensors Electrogenerated CL emission is the CL-emission phenomenon generated at the working electrode in an electrochemical-redox process that is taking place between a reagent immobilized on the working electrode and species within electrolyte solution. The modified

Figure 2. Non-competitive sandwich-type reactions with amplified or sensitized chemiluminescence detection employed in chemiluminescence-hybridization assays.

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Table 2a. Analytical parameters of ECL sensors Sensor/assay Sensing element

Analyte

Quantum dots on modified working electrode ECL-sensor (Cd/Se NCs on PIGE) H2O2 Ru(bpy)32+ CL-reagent on modified working electrode ECL-sensors LBL films of clay or SiO2 TPrA Ru(bpy)32+. oxalate ECL-senso Chit-Ru-DSNPs composite

Range

LOD (3r)

RSD (%)

Applied

2.5 · 107–6 · 105 M

1.0 · 107 M

60 nM–0.66 mM

20 nM 100 nM

[46] [49]

8.5 · 109–8.1 · 105 M

2.8 nM

[50]

1.18% (10 lM)

[47]

ECL-sensor Ru-DSNP/AuNPs LBL assembly

TPA

6.1 · 108–1.0 · 104 M

5.2 nM

5.5% (n = 8 at 8.6 lM) 7.0% (n = 5 at 8.6 lM)

ECL-sensor Ru-DSNPs/Chitosan biopolymer composite film

Itopride

1 · 108–2 · 105 g/mL

3 · 109 g/mL

2.3% (n = 11 at 8 · 108 g/mL)

Pharmaceuticals, human serum samples

ECL-sensor Ru-DSNPs / Nafion nano composite film

Metoclopromide

2 · 108–1 · 105 M

7.0 · 109 M

3.2% (n = 11 at 1.2 · 106 M)

Pharmaceutical preparations Human urine Recovery: 96.5–104.5

Isoniazid

Metallic compounds NPs on modified working electrode ECL sensor Nafion stabilized Fe3O4 TPrA multilayer film ECL-sensor polymer-coated Mag NPs immobilization matrix film

TPrA

[53]

[54]

[55] [52]

<5% (n = 11 at 1 · 108 M)

Real samples Recovery: 95–105%

[60]

Isoniazid in pharmaceutical formulations Recovery: 100%

[61]

1.0 · 1010–1 · 106 g/mL

2 · 1011 g/mL

0.1 lM–1.0 mM

50 nM

3.9% (n = 10, 0.5 mM)

[56]

49 nM

5% sensor to sensor reproducibility

[57]

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ECLIA-biosensor Ru-AuNPs-ADHEtOH 1 · 105–1 · 102 M 3.3 · 106 M Mod. ITO Luminol-, or bioenzyme-functionalized AuNP-CL-reagent system on modified working electrode ECL-sensor Core/shell NPs chit. Pyrogallol 3 · 109–2 · 105 M 1.0 · 109 M coated Luminol-doped SNPs ECL-sensor Luminol-based SiO2 NPs two step emulsion method

Ref.

Sensor/assay Sensing element ECL immunoassays ECL-IA aptasenso Ru-DSNPsaptamer conjugate ECL-sensor DBMA labeled BSA biotin

Analyte

Range

Thrombin

2 pM–2 fM

BSA IgG

1.0–80.0 lg/mL 5.0–100.0 lg/mL

hIgG

3.0 · 1011–1.0 · 109 g/mL

ABEI ECL-aptasensor Ru-DSNPs via target protein displacement

Thrombin

ECL-aptamer biosensing system Ru(bpy)32+ DMAET-Cd/Te NPs

Lysozyme

ECL-IA homogenous ABEI-antihIgG conjugate

ECL-hybridization assays ECL-DNA sensor Ru-doped S.NPs ssDNA probe

LOD (3r) 1.0 fM

RSD (%) 4.46%

Application Thrombin in plasma

Ref. [63]

8.4% (n = 5 at 10 lg/mL) 10.2% (n = 6 at 20 lg/mL)

[64]

1 · 1011 g/mL

3.1% (n = 11 at 1 · 1010 g/mL)

[66]

3.0 · 1014–1.0 · 1010 M

2 · 1014 M

1.9% at 1 · 1010 M

10 pM–10 fM

1.0 fM

4.37% (n = 4, plasma dilution >1000)

[62]

0.5 nM

7.04% (n = 6 at 50 nM)

[65]

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DNA-hybrid

2 · 1013–2 · 109 M

1 · 1013 M

ECL-DNA sensor Ru(bpy)2(dcbpy) NHS-DNA probe-AuNPs scaffold

Target ss DNA

1.0 · 1011–1.0 · 108 M

5 · 1012 M

ECL-DNA sensor Ru(bpy)2(dcbpy) NHS-DNA probe-AuNPs modified electrode

Target DNA

1.7 · 1011–1.7 · 109 M

6.7 · 1012 M

ECL-DNA sensor Mn-doped CdS NPs-MPA assembly film/(ss) DNA hairpin probe – AuNP

Target DNA

50 aM–5.0 fM

2100 copies/70 lL sample

DNA segments related to cystic fibrosis

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Table 2b. Analytical parameters of ECL-immunoassay and hybridization assay detection methods

[67]

[68]

4.3% (n = 7)

[69]

[70]

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working electrode is the sensor. A variety of ECL sensors have been reported over the past decade. These differ by the nature of the modifier used {e.g., semiconductor cluster (QDs), [Ru(bpy)3]2+ or [Ru (bpy)2(bipyridine derivative)]2+, metal NPs, CL-reagent-doped silica NPs and organic-polymer NPs (Table 2). 2.3.1. ECL sensors based on semiconductor clusters (QDs). QDs are semiconductor nanocrystals that exhibit bright, narrow, size-tunable emission as a consequence of quantum confinement; they have a number of interesting properties [e.g., broad absorption spectra (StokesÕ shift in excess of 100 nm), lifetimes longer than 10 ns, and, in many cases, better resistance to photobleaching than fluorophores]. However, QDs exhibit blinking behavior that can be a problem in some analytical work. Where the surface of QDs is used as a scaffold for the immobilization of biomolecules, control must be exercised over the surface, so that it does not limit diagnostic applications. In this area of research, a strong effort was devoted to passivating the surface of NPs or nanocrystals (NCs) and increasing sensitivity. Passivation of NP surfaces is found when surface-ECL emission gets closer to the fluorescence wavelength of the bulk material [46]. The ECL emission is generated due to an electron-transfer reaction between reduced QDs and oxidants [47]. 2.3.2. ECL sensors based on [Ru(bpy)3]2+. A relatively large number of ECL sensors have been reported by a number of research groups. Most of the reports referred to modified solid electrodes with [Ru (bpy)3]2+ found within films attached to the electrode, while some others had [Ru(bpy)3]2+ in electrolyte solution. In nearly all cases, the co-reactant is tri-n-propylamine (TPrA) unless otherwise mentioned (Table 2). ECL sensors with [Ru(bpy)3]2+ immobilized on the working electrode are regenerable thus providing sensors with relatively long life. Reports with [Ru(bpy)3]2+ modified electrode generally used different methods of immobilization and model co-reactant TPrA or derivatives in order to enhance sensitivity of the sensor [48–55]. 2.3.3. Sensors with catalysts of CL reagents immobilized on modified electrodes. Some of the most popular types of ECL sensors are those with catalysts of CL reagents immobilized on modified electrodes. These sensors, if not contaminated by oxidized products, are stable and have a long life. Several sensors have been proposed based on Naflon-stabilized Fe3O4-NP-modified Pt-electrode [56] or Pt-electrode modified by poly(3-thiopheneacetic acid)-coated magnetic NPs [57], but the most frequent reports concern cysteine film-coated Au electrodes modified by AuNPs [58,59]. The enhanced ECL signal is generated through a series of electrochemical and charge-transfer reactions between Ru[(bpy)3]2+ and a co-reactant (R) [56,57]: 1122

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Ru½ðbpyÞ3 2þ  e ! Ru½ðbpyÞ3 3þ

ðElectrochemical reactionÞ

3þ

Ru½ðbpyÞ3  þ RExcited species ! Ru½ðbpyÞ3 3þ þ ROxidized ðCharge-ztransfer reactionÞ

or on the surface of the sensor during CL-reagent oxidation (usually lucigenin or luminol), as in the following example [57]: Luc2þ þ e ! Lucþ ðElectrochemical reactionÞ O2 þ e ! O 2 ðElectrochemical reactionÞ  Lucþ þ O 2 ! LucðOO ! NMA ðECL-reactionÞ   2O ðCharge-transfer reactionÞ 2 þ H 2 O ! O2 þ HO2 þ OH

2.3.4. ECL sensors with luminol on modified electrodes. The main disadvantage of such electrodes is that the co-reactant is decomposed or converted to inactive oxidized products during the CL reaction, but these sensors have low background-CL emission. Sensors in this category include:  a chitosan-film-coated Au electrode, modified by selfassembled luminol-doped SNP sensor [60]; and,  a glassy-carbon electrode modified by a core/shell luminol-doped SNP sensor [61]. The ECL and CL signals are generated upon oxidation of luminol with O2  produced from other reactions (e.g., pyrogallol-O2). 2.4. Immunoassay or hybridization-labeled complex ECL sensors 2.4.1. ECL immunoassays (ECLIAs). Recently reported NP-assisted ECLIAs are based on the preparation of immunoassay complexes between NP-labeled bioconjugates and a bio-modified electrode. ECL emission is mainly generated by the [Ru(bpy)3]2+-co-reactantreaction system. The NP label includes [Ru(bpy)3]2+ ECL reagent or the co-reactant. Different types of ECLIA procedures have been developed depending on the CLreagent position {i.e. whether [Ru(bpy)3]2+ is involved in the electrode-modifying moiety or dissolved in the electrolyte solution, or both CL reagent and co-reactant are in electrolyte solution}. 2.4.1.1. RuðbpyÞ3 2þ label attached on the biomaterial-NP conjugate. Applications of ECLIAs with an RuðbpyÞ3 2þ label attached on the biomaterial-NP conjugate are ECL aptasensors [62,63] for determination of thrombin based on aptamer-DNA probe hybridization or a sandwich-type immunoassay complex that is aptamer-linked to an Au electrode. 2.4.1.2. Co-reactant or enhancer on the biomaterial-NPconjugate label. This category of reported ECLIAs is divided into:

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(a) techniques with the co-reactant label on the ECLIA complex linked to the modified electrode (heterogeneous ECLIAs); and, (b) homogeneous ECLIA techniques. An example of heterogeneous ECLIA with co-reactant label on the ECLIA complex linked to modified electrode is the ECL sensor described by Yin et al. [64] based on: (a) avidin-biotin reaction for the determination of bovine serum albumin (BSA); and, (b) sandwich-type complex for the determination of immunoglobulin G (IgG). ECL measurements were obtained when the electrode was brought into contact with Ru[(bpy)3]2+ solution and scanned from +0.5 to +1.3 V vs. Ag/AgCl. Applications of homogeneous ECLIAs with the coreactant label on the ECLIA complex linked to a solid substrate and the ECL-detection reaction performed with CL reagent and co-reactant labeled bio-conjugate moiety detached from the complex in electrolyte solution are exemplified in the ECL aptasensor reported by Guo et al. [65]. Another application of homogeneous ECLIAs was reported by Qi et al. [66] for the determination of human immunoglobulin G (hIgG) with N-(aminobutyl)-N-ethylisoluminol (ABEI)-labeled anti-hIgG antibody conjugate in the electrolyte solution. The ECL signal was observed at an AuNP-modified paraffin-impregnated graphite electrode (PIGE) at a constant potential, which was enhanced by binding to hIgG. 2.4.2. ECL-hybridization assays (ECLHAs). ECLHAs are used for enhanced sensitivity of DNA, recognition and sequence specification. The target DNA is attached on an electrode and [Ru(bpy)3]2+ is simply Ru or Ru-doped SNPs, covalently conjugated to appropriately capped DNA probes [67–70]. ECL measurements can be performed by applying a constant potential [68,69] or a variable potential (in pulse or step mode) [67,70].

3. Discussion Although there are numerous reports in the literature concerning CL phenomena in the presence of nanomaterials, only a few research works finally resulted in analytical methods for the determination of analytes in real samples, the most popular being those of biochemical analyses with assays of high specificity. The analytical methods developed based on NP-assisted CL and ECL detection are widely distributed between the fields of analysis of organic compounds and biochemical analysis by immunoassays and hybridization assays. However, methods based on the enhancement or inhibitory effect of the analyte on the NP-enhanced CL signal, although demonstrating high sensitivity, do not always address

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the issue of selectivity, so chromatography or capillary electrophoresis is invoked to overcome this drawback and to explore the benefits of enhanced CL detection (Tables 1 and 2). 3.1. Chemistry strategies CL emission is the energy released during a relatively strong exothermic (>41 kcal/mol) reaction (e.g., oxidation). It was already known from previous research that NP-sized redox catalysts exhibited unique behavior in catalysis because their electrochemical properties differed from those of their bulk phase. Obviously, research in CL detection, in past decade, was directed at the study of the activity of nanosized redox catalysts in CL-reaction systems to generate enhanced CL signals in gas-phase and, especially, liquid-phase systems. This was necessary because various important analytes were found in samples of high complexity (e.g., clinical, food, environmental and biological fluids) at trace or ultra-trace quantities. Apart of the conventional catalytic processes that were known to generate CL emission, (i.e. cataluminescence reactions or reactions that yielded H2O2 or NOx or CO2 or peroxidation products), it was found from experimental work with nano-sized catalysts that metalcolloid NPs of Au, Ag, and Pt were excellent catalysts for enhanced CL emission [16–29]. Most research was focused on preparing different-sized colloid NPs, investigating reaction parameters on CL signal, and using spectroscopic methods of analysis to explain the possible mechanisms. On the other hand, [Ru(bpy)3]2+- and QD-electrogenerated CL (well known to be enhanced by various agents) took a new perspective with the fabrication of sensors. For the former reagent, sensors were prepared by modifying the working electrode with: (a) film assemblies of [Ru(bpy)3]2+ adsorbed on clays or SiO2 porous materials; (b) Ru-AuNP stabilized aggregates, Ru-doped SiO2 NPs, Ru-DSNP-chitosan (or Nafion) composites, or polymer-functionalized [Ru(bpy)3]2+ [48–50,52–55]. For QDs, sensors were prepared by modifying working electrodes with film assemblies of various QDs/NPs [47]. However diffusion problems, reaction rate and chargetransfer kinetics limited the ECL response. The background CL emission was, also, a significant sensitivity factor limiting CL and especially ECL detection. Another strategy employed was to immobilize the coreactant, instead of the CL reagent, in an effort to diminish background CL. This approach employed coreactants {e.g., tri-substituted amine groups compounds for [Ru(bpy)3]2+, or co-reactants or enhancers for other CL reagents}. In addition, ECL sensors were fabricated by immobilizing AuNPs or FexOy that catalyze luminol-CL reactions.

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Despite the large amplification of the CL signal, specificity and selectivity remains an issue. To date, specificity and selectivity in NP-assisted assay were achieved through labeling bioaffinity or hybridization complexes using biomolecule-functionalized NPs. Labeling techniques were used for analytical applications in biochemistry by using protein-, antibody-, aptamer- or DNA-probe-functionalized noble-metal NPs or AuNPs as scaffolds for carrying labels (e.g., enzymes, fluorescent compounds, bio-molecule functionalized CuS NPs) [33– 35,37,38,41,43,62–70]. 3.2. Analytical parameters The analytical parameters of analytes determined by different CL-detection procedures and different types of ECL sensor reported in applications in different branches of analysis are given in Tables 1 and 2, respectively. 3.2.1. CL detection. On examination of the analytical parameters listed in Table 1 (in the part referring to those catalyzed by metal NPs), it is apparent that: (1) methods employing the enhanced CL emission of the luminol-H2O2-Au NPs CL-reagent system performed in  an FI/CL reactor system with analytes that inhibit CL emission, in most cases, exhibited a three orders of magnitude valid range with an LOD at the nM level and RSD less than 3%;  an FI/CL reactor system with analytes that enhance CL emission exhibited a mean of about two orders of magnitude with an LOD in the range 10–100 lM;  a batch-reactor system with analytes that inhibit CL emission exhibited a two orders of magnitude valid range with an LOD in the range 2.5–100 lM. (2) FI/CL-detection methods employing the strategy of energy transfer to a fluorophore from an electronically-excited molecule exhibited a variable valid range (mM–lM) and LODs (nM–lM) depending on the energy-transfer conditions on the analyte. (3) FI/CL-detection methods employing a precursor catalytic reaction usually exhibited a valid range of about two orders of magnitude and LODs in the range nM–lM. Based on these data, we infer that CL detection using an FI system is more sensitive and reproducible, and strategies involving the enhanced CL detection provide wider valid ranges but with slightly less precision. In order to alleviate this drawback, strict adherence to NPpreparation protocols is required to avoid wide distribution in the size and the shape of NPs, since different preparation procedures yield NPs of different electrochemical properties [4] (even for small changes in the nature and the concentrations of reductants and stabilizers). As a result, characterization of NPs is necessary 1124

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in many cases, which goes beyond the aims and the scope of analytical-method development and complicates the interpretation and the reproducible reproduction of an analytical method. To this end, commerciallyavailable NP solutions or certified NP materials could offer a convenient solution. 3.2.2. NP-labeled CLIAS and CLHAs. The analytical parameters of CLIAs suggest that amplified CL emission provides the best sensitivity and wider valid range compared to the enhanced CL emission obtained by the catalytic activity of metal NPs on luminol-H2O2. This is illustrated by the analytical parameters for the determination of hIgG by the different amplification methods [51–53] reported in Table 1, with valid range of three orders of magnitude and LODs in the range 0.005– 1.5 ng/mL, compared to enhanced CL emission [56] with a valid range of two orders of magnitude and an LOD of 12.8 ng/mL. The different sensitivities obtained are due to the different amplification methods, using either different-sized AuNPs (which determined the available Au ions) or Ag-coated-AuNPs (to facilitate the charge-transfer process). However, in amplified analysis, the dissolution of metal NPs is usually obtained under severe conditions with toxic and poisonous reagents (highly concentrated HNO3-HCl or HBr-Br2) and exhibits high CL background. Non-stripping CL methods based on enhanced detection were also pursued, but they usually required enhancers for better sensitivity [36]. The use of irregularly-shaped AuNPs seems to be a promising option, provided that synthesis of irregular NPs can be simplified and controlled [71]. As far as CLHAs are concerned, the methods tabulated for amplified CL emission and the analytical parameters in this group are quite similar with a valid range of about two orders of magnitude, LODs in the range of fM and RSD < 5%. 3.2.3. ECL detection. Table 2 shows the analytical parameters of various applications by ECL sensors or by ECL assays. The various types of ECL sensors with RuðbpyÞ3 2þ CL reagent located in the layers of the filmmodified working electrode of the ECL cell showed, in most cases, a valid range of about four orders of magnitude and LODs at nM-concentration levels with satisfactory reproducibility (RSD < 5%). With regards to ECL immunosensors with immunocomplex carrying various functionalized labels, the reported analytical parameters indicated that aptamer- or (ss) DNA-functionalized Ru-doped SiO2 NPs labels, in homogeneous or heterogeneous ECLIAs, exhibited wider valid ranges and much lower LODs than ECL immunocomplexes labeled otherwise, maybe due to the low background CL emission. By contrast, labels of DMBA had the lowest sensitivity.

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Finally, the analytical parameters of ECLHA sensors for DNA detection using DNA-probe-functionalized Rudoped SiO2 NPs exhibited variable valid ranges and LODs at pM levels. According to these studies, fabrication parameters had a significant effect on the ECL signal (e.g., the immobilization technique on the Au-disk working electrode). In addition, it seems that the AuNP scaffold for carrying the DNA-functionalized label somehow decreased the CL signal compared to the single DNA probe. 4. Conclusions and future work From the above discussion, it becomes clear that NPassisted CL detection is quite sensitive depending on the reaction strategy applied to generate CL emission. The strategy based on NPsÕ ability to catalyze CL-reagent systems generates enhanced CL detection as a function of the redox catalytic properties of the NP species, with activity related to their nature, composition, size, and shape. On the other hand, the emission in ECL sensors depends on the amount of CL agent that can be loaded onto the immobilization matrix, the facility of co-reactant to have easy mass transport through the immobilization matrix and charge-transfer kinetics in the heterogeneous reaction system. The analytical methods designed with these enhanced CL-detection detectors or ECL sensors have an LOD that is hardly better than the nM level. A far more sensitive method is amplified CL detection, based on the formation of stable metal NP-labeled biocomplexes, followed by dissolution and CL detection of the metal content with or without pre-concentration. Such methods have been applied mainly to immunoassays and hybridization assays, providing LODs at fmol levels. These are generally better and faster than those observed in enzyme-assisted CL assays, while they alleviate the drawbacks of instability, interferences and partial loss of enzymatic activity when conjugated with a CL reagent or enhancer. Apart from low LODs, NP-assisted CLIAs, CLHAs, ECLIAs, or ECLHAs afford improved selectivity by combining the immuno-complexes and hybridization complexes with enhanced CL detection catalyzed by Au ions. Furthermore, NP-assisted CL emission in combination with the very sensitive photo-multiplier instrumentation available nowadays makes it possible to fabricate molecular devices for: (a) observing molecular collision events; (b) designing molecular switches; (c) getting information about heterogeneous electrontransfer kinetics; (d) enabling analysis within live cells with nanoreactors; and, (e) obtaining sensitivities of less than amol quantities.

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Despite the fact that NP-assisted CL reaction strategies open new opportunities in contemporary analytical science, we must emphasize that the plethora of strategies proposed to date has not been exploited fully for their analytical potential. Refinement of NP-preparation technology (with regards to size, agglomeration and stability) is needed for the following challenging issues: (a) better precision in analytical practice; (b) more research work focused on conjugation or NP labeling with emphasis on labeling degree, bioactivity, stability and storage; and, finally, (c) the development of NP materials for nucleic-acid diagnostics, especially in living cells. The wide range of NP-assisted CL strategies and sensor devices developed provide new opportunities to expand the analytical utility of CL detection, especially with regards to selectivity issues, which become even more demanding as the complexity of samples and the number of target analytes increase progressively. References [1] H. Wang, Y. Wang, J. Jin, R. Yang, Anal. Chem. 80 (2008) 9021. [2] S.T. Dubas, V. Pimpan, Talanta 76 (2008) 29. [3] N.I. Kapakoglou, D.L. Giokas, G.Z. Tsogas, A.K. Lantavos, A.G. Vlessidis, Analyst (Cambridge, UK) 134 (2009) 2475. [4] N.P. Koutsoulis, D.L. Giokas, G.Z. Tsogas, A.G. Vlessidis, Anal. Chim. Acta 669 (2010) 45. [5] L.G. Gracia, A.M.G. Campan˜a, J.F.H. Pe´rez, F.J. Lara, Anal. Chim. Acta 640 (2009) 7. [6] G.Z. Tsogas, D.L. Giokas, P.G. Nikolakopoulos, A.G. Vlessidis, N.P. Evmiridis, Anal. Chim. Acta 573–574 (2006) 354. [7] D.L. Giokas, G.Z. Tsogas, A.G. Vlessidis, Anal. Chim. Acta 651 (2009) 188. [8] G.Z. Tsogas, A.V. Stergiou, A.G. Vlessidis, N.P. Evmiridis, Anal. Chim. Acta 541 (2005) 151. [9] A. Safavi, M.A. Karimi, Anal. Chim. Acta 468 (2002) 53. [10] N.P. Evmiridis, Analyst (Cambridge, UK) 112 (1987) 825. [11] G.Z. Tsogas, D.L. Giokas, A.G. Vlessidis, N.P. Evmiridis, J. Chromatogr., A 1107 (2006) 208. [12] G.Z. Tsogas, D.L. Giokas, A.G. Vlessidis, N.P. Evmiridis, Anal. Chim. Acta 565 (2006) 56. [13] D.L. Giokas, A.G. Vlessidis, N.P. Evmiridis, Anal. Chim. Acta 589 (2007) 59. [14] W.R. Algar, M. Massey, U.J. Krull, Trends Anal. Chem. 28 (2009) 292. [15] Y.W. Lin, C.-W. Liu, H.-T. Chang, Anal. Methods 1 (2009) 14. [16] J. Shi, R. Yan, Y. Zhu, X. Zhang, Talanta 61 (2003) 157. [17] D. Lan, B. Li, Z. Zhang, Biosens. Bioelectron. 24 (2008) 934. [18] Z. Zhang, K. Xu, W.R.G. Baeyens, X. Zhang, Anal. Chim. Acta 535 (2005) 145. [19] M.M. Karim, S.H. Lee, J. Fluoresc. 18 (2008) 827. [20] Z.-F. Zhang, H. Cui, C.-Z. Lai, L.-J. Liu, Anal. Chem. 77 (2005) 3324. [21] S.-L. Xu, H. Cui, Luminescence 22 (2007) 77. [22] T.M. Triantis, K. Papadopoulos, E. Yannakopoulou, D. Dimotikali, J. Hrbac, R. Zboril, Chem. Eng. J. 144 (2008) 483. [23] L. Wang, P. Yang, Y. Li, H. Chen, M. Li, F. Luo, Talanta 72 (2007) 1066. [24] Y. Li, P. Yang, P. Wang, L. Wang, Anal. Bioanal. Chem. 387 (2007) 585. [25] P. Yang, Y. Chen, Q. Zhou, F. Wang, Y. Li, Microchim. Acta 163 (2008) 263.

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