Sublytic C5b-9 triggers glomerular mesangial cell proliferation via enhancing FGF1 and PDGFα gene transcription mediated by GCN5-dependent SOX9 acetylation in rat Thy-1 nephritis.

Rat Thy-1 nephritis (Thy-1N) is an animal model of human mesangioproliferative glomerulonephritis (MsPGN), accompanied by glomerular mesangial cell (GMC) proliferation and extracellular matrix (ECM) deposition. Although sublytic C5b-9 formed on GMC membrane could induce cell proliferation, the mechanism is still unclear. In this study, we first demonstrated that the level of SRY related HMG-BOX gene 9 (SOX9), general control nonderepressible 5 (GCN5), fibroblast growth factor 1 (FGF1) and platelet-derived growth factor α (PDGFα) was all elevated both in the renal tissues of Thy-1N rats (in vivo) and in the GMCs (in vitro) with sublytic C5b-9 stimulation. Then, we not only discovered that sublytic C5b-9 caused GMC proliferation through increasing SOX9, GCN5, FGF1 and PDGFα expression, but also proved that SOX9 and GCN5 formed a complex and combined with FGF1 and PDGFα promoters, leading to FGF1 and PDGFα gene transcription. More importantly, GCN5 could mediate SOX9 acetylation at lysine 62 (K62) to enhance SOX9 binding to FGF1 or PDGFα promoter and promote FGF1 or PDGFα synthesis and GMC proliferation. Besides, the experiments in vivo also showed that FGF1 and PDGFα expression, GMC proliferation and urinary protein secretion in Thy-1N rats were greatly reduced by silencing renal SOX9, GCN5, FGF1 or PDGFα gene. Furthermore, the renal tissues of MsPGN patients also exhibited positive expression of these genes mentioned above. Collectively, our findings indicate that GCN5, SOX9 and FGF1/PDGFα can form an axis and play an essential role in sublytic C5b-9-triggered GMC proliferation, which might provide a novel insight into the pathogenesis of Thy-1N and MsPGN.

Targeting NF-κB c-Rel in regulatory T cells to treat corneal transplantation rejection.

The relevance of Tregs in the induction of tolerance against corneal allografts has been well established. Although it is well known that the conversion of Tregs into effector-like cells contributes to the loss of corneal immune privilege, the underlying mechanism is still not fully understood. Using heterologous penetrating keratoplasty model, we found that Tregs from corneal allograft rejected mice (inflam-Tregs) exhibit impaired function and characteristics of effector T cells. Further study showed that the expression of NF-κB c-Rel, a key mediator of effector T cell function, was significantly increased in inflam-Tregs. Mechanistic study revealed that elevated NF-κB c-Rel level in inflam-Tregs impaired Treg function through the promotion of inflammatory cytokine production and glycolysis. More importantly, we demonstrated that targeting NF-κB c-Rel was able to improve the immune suppressive function of inflam-Tregs in vitro and enhance the potential of them to suppress corneal transplantation rejection. Therefore, our current study identified NF-κB c-Rel as a key mediator of the conversion of Tregs into effector-like cells when under inflammatory environment.

Effects of bulk and surface defects on the photocatalytic performance of size-controlled TiO2 nanoparticles.

Defects are one of the crucial factors influencing the photocatalytic activities of TiO2 nanoparticles (NPs), as they directly affect the charge separation efficiencies. However, we found that the effect of bulk or surface defects on the photocatalytic performance of TiO2 is totally different. Here, we controllably synthesized four kinds of uniform TiO2 NPs, which were used to investigate how the bulk and surface defects influenced the photoactivities of samples with different particle sizes. Through XPS and EPR analysis, bulk and surface defects were observed in all the TiO2 samples and their amounts had been reduced after the calcination process. The reduction of bulk and surface defects enhanced the photoactivities of TiO2 NPs with diameters >10 nm, but the situation was inverse for those samples with diameters <10 nm. Therefore, we demonstrated that bulk defects play a dominate role in the change in photoactivity in particles larger than 10 nm, while surface defects are more important than bulk defects for particles less than 10 nm.

Epigenetic regulation of CDH1 exon 8 alternative splicing in gastric cancer.

BACKGROUND: The tumor suppressor gene CDH1 is critical for intercellular adhesion. In our previous work, we reported a nonfunctional CDH1 transcript that lacks the final 83 base pairs of exon 8 (1054del83). In this work, we probed the role of histone epigenetic modifications as well as DNA methylation in selection of this isoform. METHODS: RT-qPCR was used to detect CDH1 RNA expression. Methylation of CDH1 was analyzed by bisulphite sequencing PCR. ChIP assay was performed to show histones level. Cell lines were treated with DNA methyltransferase inhibitor AZA, HDAC inhibitor TSA, or siRNA oligonucleotides to test regulation of CDH1 splicing. RESULTS: Greater CDH1 1054del83 transcripts were observed in gastric cancer (GC) cell lines than human gastric mucosal epithelial cell line GES-1. All the cell lines showed significant methylation pattern at the CpG sites of CDH1 exon 8. AZA treatment did not influence selection of 1054del83 transcripts. A significant decrease in acetylation for histones H3 and H4K16Ac in an internal region of the CDH1 gene surrounding the alternative exon 8 were detected in GC cell lines. Treatment with TSA preferentially expressed the correctly spliced transcript and not the exon 8 skipped aberrant transcripts, showing that histone acetylation was involved in the splicing regulation. SiRNA-mediated knockdown of SETD2 (The specific methyltransferase of H3K36) decreased exclusion of exon 8, suggesting that the presence of this mark correlates with increased skipping of the final 83 base pairs of CDH1 exon 8. However, CDH1 splicing was not affected by SRSF2 knockdown. CONCLUSIONS: H3K36me3 correlates with increased skipping of the final 83 base pairs of CDH1 exon 8. Histone acetylation was involved in the splicing regulation as well.

Serum LINC00339 is a promising biomarker for prognosis prediction of nasopharyngeal carcinoma.

BACKGROUND: Long non-coding RNAs (lncRNAs) in serum were useful and promising biomarkers for diagnostic and prognostic application. Herein, we investigated the serum lncRNA LINC00339 expression and its role in nasopharyngeal carcinoma. METHODS: In this study, we recruited a cohort of 129 nasopharyngeal carcinoma patients, 68 patients with nasopharyngitis, and 80 healthy controls. Serum LINC00339 levels were measured by reverse transcription-quantitative polymerase chain reaction. Receiver operating characteristic (ROC) and Kaplan-Meier curve analyses were conducted to evaluate th e clinical role of LINC00339. The effects of linc00339 on cellular activities were measured using CCK-8 and Transwell assays. RESULTS: We observed that serum LINC00339 expression was upregulated in nasopharyngeal carcinoma patients and closely associated with tumor node metastasis stage, lymph node metastasis, and overall survival rate. Meanwhile, ROC analysis showed serum LINC00339 had diagnostic value to distinguish nasopharyngeal carcinoma patients from healthy individuals and nasopharyngitis patients. Silencing of LINC00339 could repress cellular behaviors by targeting miR-152. CONCLUSION: This study clarified that LINC00339 was upregulated in nasopharyngeal carcinoma, and that serum LINC00339 may act as a diagnostic or prognostic marker, and a hopeful therapeutic target for patients with nasopharyngeal carcinoma.

ERK1/2-dependent activity of SOX9 is required for sublytic C5b-9-induced expression of FGF1, PDGFα, and TGF-ß1 in rat Thy-1 nephritis.

Mesangial proliferative glomerulonephritis (MsPGN) and its related rat model Thy-1 nephritis (Thy-1N) are associated with C5b-9 deposition and are characterized by proliferation of glomerular mesangial cell (GMC) and expansion of extracellular matrix (ECM) expansion, alongside overexpression of multiple growth factors. Although fibroblast growth factor 1 (FGF1), platelet-derived growth factor alpha (PDGFα), and transforming growth factor beta 1 (TGF-ß1) are well known for their proproliferative and profibrotic roles, the molecular mechanisms responsible for regulating the expression of these growth factors have not been thoroughly elucidated. In this study, we found that sublytic C5b-9 induction of sex-determining region Y-box 9 (SOX9) transactivated FGF1, PDGFα, and TGF-ß1 genes in GMCs, resulting in a significant increase in their mRNA and protein levels. Besides, sublytic C5b-9 induction of activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) phosphorylated SOX9 at serine 181 and serine 64, which enhanced SOX9's ability to transactivate FGF1, PDGFα, and TGF-ß1 genes in GMCs. Furthermore, we demonstrated that inhibiting ERK1/2 activation or silencing either ERK1/2 or SOX9 gene led to reduced SOX9 phosphorylation, decreased generation of FGF1, PDGFα, and TGF-ß1, and ameliorated glomerular injury in rat Thy-1N. Overall, these findings suggest that expression of FGF1, PDGFα, and TGF-ß1 is promoted by ERK1/2-mediated phosphorylation of SOX9, which may provide a valuable insight into the pathogenesis of MsPGN and offer a potential target for the development of novel treatment strategies for MsPGN.

Poly(pyrocatechol-3,5-disodiumsulfonate)/multi-walled carbon nanotubes composite for simultaneous determination of dopamine, ascorbic acid and uric acid.

A novel poly(pyrocatechol-3,5-disodiumsulfonate)/multi-walled carbon nanotubes (PPD/MWCNT) composite film modified glassy carbon electrode (GCE) was fabricated and successfully used to simultaneously determine ascorbic acid (AA), dopamine (DA) and uric acid (UA). Owing to the synergistic effects of PPD and MWCNTs, the electrochemical responses of AA, DA and UA at PPD/MWCNTs/GCE were clearly superior to those at bare GCE. Additionally, the PPD/MWCNTs composite film could reduce the potential overlap of AA, DA and UA so as to favor simultaneous determination of the three substances. Under the optimum conditions, the linear calibration curves of AA, DA and UA were obtained by differential pulse voltammetry in the range of 10-1000, 1-100 and 4-1200 microM, and the detection limits were 5, 0.5 and 0.5 microM (S/N = 3) respectively. The results showed PPD/MWCNTs/GCE possessed good reproducibility and stability, and it could be applied to determine UA in human urine with satisfying recovery by standard addition method.

Colorimetric determination of melamine by pyridine-3-boronic acid modified gold nanoparticles.

A rapid, simple and sensitive colorimetric detection method for melamine was proposed based on pyridine-3-boronic acid (PBA) modified gold nanoparticles (AuNPs). The formation of supramolecular hydrogen-bonded structure between PBA and melamine resulted in the aggregation of PBA modified AuNPs and the color change from red to blue. Melamine could be detected by colorimetric response of AuNPs that could be detected by naked eyes or a UV-vis spectrophotometer. The detection concentration of melamine ranged from 6.0 x 10(-8) to 1.6 x 10(-6) M, and the detection limit (3sigma) was 3.0 x 10(-8) M (i.e., 3.2 ppb). This provided an effective and facile colorimetric sensor for real-time and on-site determination of melamine. Particularly, the proposed method could be used to detect melamine in pretreated liquid milk products with high sensitivity and low interference, and the recoveries were from 95% to 102%.

Colorimetric detection of Cd2+ using gold nanoparticles cofunctionalized with 6-mercaptonicotinic acid and L-cysteine.

We have developed a colorimetric assay for the highly sensitive and selective detection of Cd(2+) using gold nanoparticles (AuNPs) cofunctionalized with 6-mercaptonicotinic acid (MNA) and L-Cysteine (L-Cys) through the formation of an Au-S bond. In the presence of Cd(2+), the aggregation of functionalized AuNPs occurred by means of a metal-ligand interaction that led to visible color changes. Most importantly, cofunctionalized AuNPs had better responses for Cd(2+) than that functionalized by either MNA or L-Cys. Cd(2+) could be detected by the colorimetric response of AuNPs that could be detected by the naked eye or a UV-vis spectrophotometer. The absorbance ratio (A(620)/A(523)) was linear with the Cd(2+) concentration in the range of 2.0 × 10(-7) to 1.7 × 10(-6) M. Under optimum conditions (2.0 × 10(-5) M MNA, 2.0 × 10(-6) M L-Cys and 0.020 M NaCl at pH 10.0), the detection limit (3σ) of Cd(2+) could be as low as 1.0 × 10(-7) M. Interference experiments showed that Pb(2+) and Cu(2+) caused a slight interference for Cd(2+) determination while other metal ions caused no interference. The proposed method was successfully applied to determine the concentration of Cd(2+) in environmental samples (lake water).

Simultaneous determination of 3,4-dihydroxyphenylacetic acid, uric acid and ascorbic acid by poly(L-arginine)/multi-walled carbon nanotubes composite film.

The poly(L-Arginine)(PArg)-multiwalled carbon nanotubes (MWCNTs) composite film was used to modify glassy carbon electrode (GCE) to fabricate PArg/MWCNTs/GCE through electropolymerization of L-Arginine on MWCNTs/GCE. The PArg/MWCNTs/GCE exhibited high electro-catalytic activities towards the oxidation of 3,4-dihydroxyphenylacetic acid (DOPAC), uric acid (UA), and ascorbic acid (AA), and could be sensitively used for simultaneous determination of DOPAC, AA, and UA in pH 7.4 phosphate-buffered solution (PBS).The linear ranges were 7 microM to 2.7 mM for DOPAC, 3 microM to 1.2 mM for UA, and 70 microM to 1.4 mM for AA. The detection limits were 1.3 microM for DOPAC, 0.7 microM for UA and 20 microM for AA.

Determination of Cu2+ using poly(2-aminothiazole)/ multi-walled carbon nanotubes composite film modified glassy carbon electrodes.

2-Aminothiazole was electropolymerized by cyclic voltammetry (CV) on the multi-walled carbon nanotubes (MWCNTs) modified glassy carbon electrode (GCE) surface. Poly(2-aminothiazole)/MWCNTs/GCE was used for determination of copper ions. The anodic peak currents of copper ions evaluated by differential pulse stripping voltammetry (DPSV) are linear with the concentrations in the range from 1.0 x 10(-7) M to 2.0 x 10(-5) M with a linear coefficiency of 0.9985. The detection limit is 2.0 x 10(-9) M calculated for a signal-to-noise ratio of 3 (S/N = 3). The proposed method was applied successfully to the determination of copper ions in drinking water, and the recovery was 96%.

Rutile TiO2 single crystals delivering enhanced photocatalytic oxygen evolution performance.

Owing to their scientific and technological importance, the development of highly efficient photocatalytic water oxidation systems with rapid photogenerated charge separation and high surface catalytic activity is highly desirable for the storage and conversion of solar energy. A promising candidate is rutile phase titanium dioxide (TiO2), which has been widely studied over half a century. Specifically, oriented single-crystalline TiO2 surfaces with high oxidative reactivity would be most desirable, but achieving these structures has been limited by the availability of synthetic techniques. In this study, a facile and green synthetic approach was developed for the first time to synthesize rutile TiO2 single crystals with regulable reductive and oxidative facets. Glycolic acid (GA) and sodium fluoride (NaF) are used as the crucial and effective phase and facet controlling agents, respectively. The selective adsorption of F- ions on the facets of rutile TiO2 crystals not only plays a key role in driving the nucleation and preferential growth of the crystals with desired facets but also significantly affects their photocatalytic gas evolution reactivity. With heat treatment, the highly stable F--free rutile TiO2 single crystals with a high percentage of oxidative facets exhibit a superior photocatalytic gas evolution rate (≈116 µmol h-1 per 0.005 g catalyst), 8.5 times higher than that of previous F--containing samples.

Sublytic C5b-9 induces glomerular mesangial cell proliferation via ERK1/2-dependent SOX9 phosphorylation and acetylation by enhancing Cyclin D1 in rat Thy-1 nephritis.

Glomerular mesangial cell (GMC) proliferation is a histopathological alteration in human mesangioproliferative glomerulonephritis (MsPGN) or in animal models of MsPGN, e.g., the rat Thy-1 nephritis (Thy-1N) model. Although sublytic C5b-9 assembly on the GMC membrane can trigger cell proliferation, the mechanisms are still undefined. We found that sublytic C5b-9-induced rat GMC proliferation was driven by extracellular signal-regulated kinase 1/2 (ERK1/2), sry-related HMG-box 9 (SOX9), and Cyclin D1. Here, ERK1/2 phosphorylation was a result of the calcium influx-PKC-α-Raf-MEK1/2 axis activated by sublytic C5b-9, and Cyclin D1 gene transcription was enhanced by ERK1/2-dependent SOX9 binding to the Cyclin D1 promoter (-582 to -238 nt). In addition, ERK1/2 not only interacted with SOX9 in the cell nucleus to mediate its phosphorylation at serine residues 64 (a new site identified by mass spectrometry) and 181 (a known site), but also indirectly induced SOX9 acetylation by elevating the expression of general control non-repressed protein 5 (GCN5), which together resulted in Cyclin D1 synthesis and GMC proliferation. Moreover, our in vivo experiments confirmed that silencing these genes ameliorated the lesions of Thy-1N rats and reduced SOX9 phosphorylation, acetylation and Cyclin D1 expression. Furthermore, the renal tissue sections of MsPGN patients also showed higher phosphorylation or expression of ERK1/2, SOX9, and Cyclin D1. In summary, these findings suggest that sublytic C5b-9-induced GMC proliferation in rat Thy-1N requires SOX9 phosphorylation and acetylation via enhanced Cyclin D1 gene transcription, which may provide a new insight into human MsPGN pathogenesis.

Correction: Effect of aspect ratios of rutile TiO2 nanorods on overall photocatalytic water splitting performance.

Correction for 'Effect of aspect ratios of rutile TiO2 nanorods on overall photocatalytic water splitting performance' by Bing Fu et al., Nanoscale, 2020, DOI: 10.1039/c9nr10870j.

Effect of aspect ratios of rutile TiO2 nanorods on overall photocatalytic water splitting performance.

The spatial separation of reduction and oxidation reaction sites on the different facets of a semiconductor is an ideal and promising route for overall photocatalytic water splitting due to efficient charge carrier separation. Rutile TiO2 has separate oxidation and reduction crystal facets and can be used to achieve direct splitting of pure water under ultraviolet (UV) light irradiation. In order to improve the rate of water oxidation reaction, the ratio of different crystal facets of rutile should be regulated controllably. However, the preparation of rutile TiO2 architecture has been limited by the availability of synthetic techniques. In this study, rutile TiO2 nanorods with various aspect ratios were accurately prepared in the presence of Cl- anions and H+ cations, which were found to play a crucial role in forming the morphology of rutile TiO2 nanorods. In addition, the mechanism involving the growth of rutile TiO2 nanorods with different aspect ratios is proposed. Rutile TiO2 nanorods with a high proportion of oxidative (111) facets provided higher overall water splitting reactivity.

KLF6 Acetylation Promotes Sublytic C5b-9-Induced Production of MCP-1 and RANTES in Experimental Mesangial Proliferative Glomerulonephritis.

Rat Thy-1 nephritis (Thy-1N) is an experimental mesangial proliferative glomerulonephritis (MsPGN) for studying human MsPGN. Although sublytic C5b-9 complex formation on glomerular mesangial cells (GMCs) and renal MCP-1 and RANTES production in rats with Thy-1N have been proved, the role and mechanism of MCP-1 or RANTES synthesis in GMCs induced by sublytic C5b-9 are poorly elucidated. In this study, we first found the expression of transcription factor (KLF6), co-activator (KAT7) and chemokines (MCP-1 and RANTES) was all up-regulated both in renal tissue of Thy-1N rats (in vivo) and in sublytic C5b-9-induced GMCs (in vitro). Further in vitro experiments revealed that KLF6 bound to MCP-1 promoter (-297 to -123 nt) and RANTES promoter (-343 to -191 nt), leading to MCP-1 and RANTES gene transcription. Meanwhile, KAT7 also bound to the same region of MCP-1 and RANTES promoter in a KLF6-dependent manner, and KLF6 was acetylated by KAT7 at lysine residue 100, which finally promoted MCP-1 and RANTES expression. Moreover, our in vivo experiments discovered that knockdown of renal KAT7 or KLF6 gene obviously reduced MCP-1 and RANTES production, GMCs proliferation, ECM accumulation, and proteinuria secretion in Thy-1N rats. Collectively, our study indicates that sublytic C5b-9-induced MCP-1 and RANTES synthesis is associated with KAT7-mediated KLF6 acetylation and elevated KLF6 transcriptional activity, which might provide a new insight into the pathogenesis of rat Thy-1N and human MsPGN.

A novel graphene-DNA biosensor for selective detection of mercury ions.

A novel electrochemical biosensor for sensitive and selective detection of mercury (II) ions (Hg²âº) based on a DNA grafted graphene is proposed. Graphene oxide (GO) was reduced by dopamine, and then the single-strand probe DNA modified at the 5'-end with an alkylamino modifier (NH2-ssDNA) was grafted on the reduced graphene oxide (RGO) surface via Michael addition reaction. In the presence of Hg²âº, the target DNA with four thymine-thymine (T-T) mismatches would hybridize with the probe DNA on the glassy carbon electrode (GCE) through T-Hg²âº-T coordination chemistry. The hybridization of the two oligonucleotides leads to the increase in the peak currents of [Ru(NH3)6]³âº, which could be used for electrochemical sensing of Hg²âº. The difference in the value of the peak currents of [Ru(NH3)6]³âº before and after DNA hybridization was linear with the concentration of Hg²âº in the range from 8.0×10⁻9 to 1.0×10⁻7 M with a linear coefficiency of 0.996. The detection limit was 5.0×10⁻9 M (S/N=3). The proposed electrochemical biosensor is rapid, convenient and low-cost for effective sensing of Hg²âº. Particularly, the proposed method was applied successfully to the determination of Hg²âº in real environmental samples.

Colorimetric sensing of clenbuterol using gold nanoparticles in the presence of melamine.

A highly sensitive method for the detection of trace amount of clenbuterol based on gold nanoparticles (AuNPs) in the presence of melamine was described in this paper. Hydrogen-bonding interaction between clenbuterol and melamine resulted in the aggregation of AuNPs and a consequent color change of AuNPs from wine red to blue. The concentration of clenbuterol could be determined with naked eye or a UV-vis spectrometer. Results showed that the absorption ratio (A(670)/A(520)) was liner with the logarithm of clenbuterol concentration in the range of 2.8×10(-10) to 2.8×10(-7)M and 2.8×10(-7) to 1.4×10(-6)M with linear coefficients of 0.996 and 0.993, respectively. The detection limit was 2.8×10(-11)M (S/N=3), which was much lower than most existing methods. The coexisting substances including dl-epinephrine, phenylalamine, tryptohan, alamine, uric acid, glycine, glycerol, glucose, MgCl(2), CaCl(2) and NaCl did not affect the determination of clenbuterol. The proposed method could be successfully applied to the determination of clenbuterol in human urine.

The comparison of different gold nanoparticles/graphene nanosheets hybrid nanocomposites in electrochemical performance and the construction of a sensitive uric acid electrochemical sensor with novel hybrid nanocomposites.

In this paper, water soluble poly(diallyldimethylammonium chloride)-graphene nanosheets (PDDA-GNs) were synthesized and characterized by UV-visible absorption spectroscopy, X-ray diffraction (XRD) and X-ray photoelectron spectroscopy (XPS). On the basis of PDDA-GNs, three different types of gold nanoparticles/graphene nanosheets (AuNPs/GNs) hybrid nanocomposites were obtained by one-pot synthesis, in situ reduction and adsorption methods, respectively. These nanocomposites were used as electrode materials for electrochemical determination of uric acid (UA). The results indicated adsorption to be the best method to synthesize hybrid nanocomposites from the electrochemical point of view. Given the fact positively charged PDDA-AuNPs could interact with negatively charged UA molecules, we then synthesized PDDA-protected gold nanoparticles/graphene nanosheets (PDDA-AuNPs/GNs) hybrid nanocomposites by adsorption method, for the first time. As were expected, PDDA-AuNPs/GNs gave better performance for UA than AuNPs/GNs obtained by adsorption, and the anodic peak current of UA obtained by cyclic voltammetry (CV) increased 102.1-fold in comparison to bare GCE under optimizing conditions. Differential pulse voltammetry (DPV) was used to quantitatively determine UA. The linear range of UA was from 0.5µM to 20µM and the detection limit was 0.1µM (S/N=3) with a high sensitivity of 103.08µAµM(-1)cm(-2). The assay results of urine sample provided satisfying recoveries by standard addition method. In addition, the anodic peaks of adrenaline (AD) and UA were well resolved at PDDA-AuNPs/GNs modified electrode, while they were too overlapped to separate at bare electrode, as a result of that UA was successfully detected in the presence of AD. In conclusion, rapid synthesis of PDDA-AuNPs/GNs were realized and applied as an advanced hybrid electrode material for UA determination.

Electrochemical immunosensor for casein based on gold nanoparticles and poly(L-Arginine)/multi-walled carbon nanotubes composite film functionalized interface.

In this paper, a novel electrochemical immunosensor for the determination of casein based on gold nanoparticles and poly(L-Arginine)/multi-walled carbon nanotubes (P-L-Arg/MWCNTs) composite film was proposed. The P-L-Arg/MWCNTs composite film was used to modify glassy carbon electrode (GCE) to fabricate P-L-Arg/MWCNTs/GCE through electropolymerization of L-Arginine on MWCNTs/GCE. Gold nanoparticles were adsorbed on the modified electrode to immobilize the casein antibody and to construct the immunosensor. The stepwise assembly process of the immunosensor was characterized by cyclic voltammetry and differential pulse voltammetry. Results demonstrated that the peak currents of [Fe(CN)(6)](3-/4-) redox pair decreased due to the formation of antibody-antigen complex on the modified electrode. The optimization of the adsorption time of gold nanoparticles, the pH of supporting electrolyte and the incubation time were investigated in details. Under optimal conditions, the peak currents obtained by DPV decreased linearly with the increasing casein concentrations in the range from 1 × 10(-7) to 1 × 10(-5) g mL(-1) with a linear coefficiency of 0.993. This electrochemical immunoassay has a low detection limit of 5 × 10(-8) g mL(-1) and was successfully applied to the determination of casein in cheese samples.