Huisheng Zhuang

Biotin-streptavidin enzyme-linked immunosorbent assay for detecting Tetrabromobisphenol A in electronic waste.

Tetrabromobisphenol A (TBBPA) is a widely used brominated flame retardant. A sensitive and selective indirect competitive biotin-streptavidin-amplified enzyme-linked immunosorbent assay (BA-ELISA) was developed for detecting TBBPA. The optimal hapten of TBBPA was 2-(2,6-dibromo-4-(2-(3,5-dibromo-4-hydroxyphenly)propan-2-yl)) acetic acid. Several physiochemical factors that influence assay performance, such as optimal coupling concentration of immunogen and antibody, organic solvent, ionic strength, and pH, were studied and optimized. The limit of detection (IC10) was 0.027 ng/mL and the median inhibitory concentration (IC50) was 0.58 ng/mL. The BA-ELISA was highly selective, with low cross-reactivity with TBBPA analogs. Finally, the assay was used to detect TBBPA in electronic waste samples. The results are consistent with those using liquid chromatography, which proves that the proposed immunoassay is accurate and receptive. This BA-ELISA method is suitable for the rapid and sensitive screening of TBBPA in environmental monitoring.

An ultrasensitive gold nanoparticles improved real-time immuno-PCR assay for detecting diethyl phthalate in foodstuff samples.

A specific polyclonal antibody targeting diethyl phthalate (DEP) with the higher antibody titer at 1:120,000 has been obtained, and an ultrasensitive and high-throughput direct competitive gold nanoparticles improved real-time immuno-PCR (GNP-rt-IPCR) technique has been developed for detecting DEP in foodstuff samples. Under optimal conditions, a rather low linearity is achieved within a range of 4 pg L(-1) to 40 ng L(-1), and the limit of detection (LOD) is 1.06 pg L(-1). Otherwise, the GNP-rt-IPCR technique is highly selective, with low cross-reactivity values for DEP analogs (<5%). Finally, the concentrations of DEP in foodstuff samples by the GNP-rt-IPCR method range from 0.48 to 41.88 μg kg(-1). Satisfactory recoveries (88.39-112.79%) and coefficient of variation values (8.38-12.77%) are obtained. The consistency between the results obtained from GNP-rt-IPCR and gas chromatography-mass spectrometry (GC-MS) is 98.3%, which further proves that GNP-rt-IPCR is an accurate, reliable, rapid, ultrasensitive, and high-throughput method for batch determination of trace amounts of DEP in foodstuff samples.

A heterogeneous biotin–streptavidin-amplified enzyme-linked immunosorbent assay for detecting tris(2,3-dibromopropyl) isocyanurate in natural samples

Tris(2,3-dibromopropyl) isocyanurate (TBC) is a novel brominated flame retardant (BFR) that is widely used to substitute the prohibited BFRs throughout the world. With the development of research, the potential environmental and ecological harms of TBC have been revealed. For sensitive and selective detecting TBC, an indirect competitive biotin-streptavidin-amplified enzyme-linked immunosorbent assay (BA-ELISA) has been established in this study. The small molecular TBC-hapten was synthesized first; it mimicked the chemical structure of TBC and possessed a secondary amine group. The as-obtained hapten was then conjugated with carrier proteins to prepare artificial antigen. After immunization, the anti-TBC polyclonal antibody was obtained from separating rabbit serum. The procedures of this BA-ELISA were optimized. Under the optimal conditions, the limit of detection (IC10) was 0.0067 ng/ml and the median inhibitory concentration (IC50) was 0.66 ng/ml. Cross-reactivity values of the BA-ELISA with the tested TBC analogues were ⩽5%. This immunoassay was successfully applied to determine the TBC residue in river water samples that were collected near a BFR manufacturing plant. Satisfactory recoveries (92.1-109.2%) were obtained. The results indicated that this proposed BA-ELISA is suitable for the rapid and sensitive determining of TBC in environmental monitoring.

A real-time immuno-PCR assay for the detection of tetrabromobisphenol A

In this study, a reliable and ultra-sensitive indirect competitive real-time immuno-PCR (rt-iPCR) was established for the determination of tetrabromobisphenol A (TBBPA). The optimal hapten, artificial immunogen, coating antigen, and polyclonal anti-TBBPA antibodies were successfully prepared. For optimizing rt-iPCR, several physiochemical factors, such as optimal coupling concentration of immunogen and antibody, and the amounts of streptavidin and biotinylated DNA, are discussed. Using this proposed rt-iPCR assay, TBBPA could be determined in the range from 10 pg L−1 to 10 ng L−1, with a detection limit of 2 pg L−1. This rt-iPCR was selective, with low cross-reactivity with TBBPA analogs. Finally, seabed sediment samples were analyzed by rt-iPCR and liquid chromatography. The tested results were demonstrated to be receivable and accurate. The average recovery was between 89.8% and 113.6%. We think this developed rt-iPCR will be very useful in the field of environmental science.

An indirect competitive enzyme-linked immunosorbent assay for the determination of 3,4-dichlorobiphenyl in sediment using a specific polyclonal antibody

A specific polyclonal antibody targeting non-dioxin-like PCB 3,4-dichlorobiphenyl (PCB12) was obtained, and a sensitive indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) was developed for the determination of PCB12 in sediment samples. Under optimal conditions, good linearity was achieved within a range of 0.06 to 6 μg L−1. The observed half-maximal inhibition concentration (IC50) was 2.37 μg L−1, and the limit of detection (LOD) was 0.021 μg L−1. This method was used for the detection of PCB12 in the sediment samples collected from the East China Sea adjacent to Shanghai, China. The concentrations of PCB12 in the samples ranged from 0.21 μg kg−1 to 8.59 μg kg−1. The recovery was from 81% to 105% and the CV values were from 2.8% to 8.4%. The consistency between the results obtained from ic-ELISA and GC-ECD was 98%. It further confirmed the reliability and accuracy of the ic-ELISA for rapid detection of PCB12 in the environment.

A carbon nanotube-enhanced real-time immuno-PCR for ultrasensitive detection of AHTN in water

Polycyclic musks (PCMs) in the aquatic environment have become an emerging environmental issue because of their potential risk. The most commonly used method for analysis of PCMs is gas chromatography-mass spectrometer (GC-MS) with different sample extractions, which are somewhat expensive to operate, laborious and complex. In this paper, a carbon nanotube-enhanced real time immuno-PCR was developed for ultrasensitive detection of AHTN in water for the first time. The SWCNTs were used to immobilize numerous amino-DNA and polyclonal antibody to form polyclonal antibody-CNTs-DNA conjugates, which were used as a signal-amplifier in the proposed immunoassay system. This proposed carbon nanotube enhanced real time immuno-PCR assay was used to determine AHTN in water samples ranging from 5 pg/L-0.1 ng/L; using sample size as low as 10 μL. This proposed carbon nanotube enhanced real time immuno-PCR is the most ultrasensitive one for determination of AHTN in water without pre-concentration or extractions; and it provide a potential way for ultra-trace AHTN detection in the aquatic environment.

Determination of multi-residue PCBs in air by real-time fluorescent quantitative immuno-PCR assay

Based on the prepared group-specific antibodies against polychlorinated biphenyls (PCBs), a direct competitive real-time fluorescent quantitative immuno-polymerase chain reaction (rt-IPCR) assay was developed. The purpose of the assay was the determination of multi-residue PCBs in indoor air samples. In the assay, male New Zealand white rabbits were immunized with an immune antigen mixture composed of PCB12-O-BSA, PCB37-O-BSA, and PCB77-O-BSA. The specific polyclonal antibodies (pAbs) to multi-residue PCBs were obtained and used to develop a direct competitive rt-IPCR assay. The specificities of the pAbs were examined by the indirect competitive enzyme-linked immunosorbent assay (id-ELISA). The assays were found to be highly specific for PCB congeners as well as Aroclors 1248 and 1242. The effect of optimal reagent concentrations on reducing background fluorescence was also investigated. Using the optimized assay, a standard curve for Aroclor 1248 was prepared. The linear range for the determination of PCBs was 10 to 106 fg mL−1 with a correlation coefficient of 0.98 and a detection limit of 10.25 fg mL−1. The entire procedure was then evaluated using spiked air samples. The rt-IPCR results for the air samples were confirmed by gas chromatography/mass spectrometry and ELISA. Recovery was lower or higher with agitation but would still be acceptable for use in an on-site field test to provide rapid, semiquantitative, and reliable test results for detection of PCBs in air samples.

Competitive immunoassay for monitoring polybrominated diphenyl ethers in PM2.5 in Shanghai

ABSTRACT In this paper, a sensitive biotin–streptavidin (BS)-ELISA was developed for determining the polybrominated diphenyl ethers in PM2.5. For establishing this proposed BS-ELISA, we prepared the biotinylated antibody primarily. And for reducing the background interference, some influencing factors and procedures for this immunoassay were also discussed and optimised. Under the optimal conditions, the IC50 = 0.53 ng/mL; IC10 was 0.002 ng/mL; and the results were almost consistent with those using the gas chromatography and electron capture detection (GC-ECD). Less procedures and simpler sample preparation were required for this method compared with the GC-ECD. The results showed that the highest value of BDE-47 concentration occurred in December, which might reflect the combination of heating and industrial pollution. In our analysis, we studied the Pearson correlations between BDE-47/PM2.5 and gaseous pollutants (such as NO2, SO2, CO, O3, PM10 and PM2.5). BDE-47 showed a higher correlation with NO2 than that with PM10, PM2.5 and SO2, which implied that the BDE-47 emission process might be accompanied by the emission of NO2. Moreover, during the Spring Festival, the concentration of BDE-47 in PM2.5 decreased significantly, whereas the PM2.5 changed little. This suggested factories and vehicles might be the major contributors to BDE-47 emissions (but not to PM2.5).