Diagnostic potential of nanoparticle aided assays for MUC16 and MUC1 glycovariants in ovarian cancer.

Our study reports the discovery and evaluation of nanoparticle aided sensitive assays for glycovariants of MUC16 and MUC1 in a unique collection of paired ovarian cyst fluids and serum samples obtained at or prior to surgery for ovarian carcinoma suspicion. Selected glycovariants and the immunoassays for CA125, CA15-3 and HE4 were compared and validated in 347 cyst fluid and serum samples. Whereas CA125 and CA15-3 performed poorly in cyst fluid to separate carcinoma and controls, four glycovariants including MUC16MGL , MUC16STn , MUC1STn and MUC1Tn provided highly improved separations. In serum, the two STn glycovariants outperformed conventional CA125, CA15-3 and HE4 assays in all subcategories analyzed with main benefits obtained at high specificities and at postmenopausal and early-stage disease. Serum MUC16STn performed best at high specificity (90%-99%), but sensitivity was also improved by the other glycovariants and CA15-3. The highly improved specificity, excellent analytical sensitivity and robustness of the nanoparticle assisted glycovariant assays carry great promise for improved identification and early detection of ovarian carcinoma in routine differential diagnostics.

Europium Fluorescent Nanoparticles-Based Multiplex Lateral Flow Immunoassay for Simultaneous Detection of Three Antibiotic Families Residue.

A fluorescent immunoassay based on europium nanoparticles (EuNPs-FIA) was developed for the simultaneous detection of antibiotic residues, solving the problems of single target detection and low sensitivity of traditional immunoassay methods. In the EuNPs-FIA, EuNPs were used as indictive probes by binding to anti-tetracyclines monoclonal antibodies (anti-TCs mAb), anti-sulphonamides monoclonal antibodies (anti-SAs mAb) and anti-fluoroquinolones monoclonal antibodies (anti-FQs mAb), respectively. Different artificial antigens were assigned to different regions of the nitrocellulose membrane as capture reagents. The EuNPs-FIA allowed for the simultaneous detection of three classes of antibiotics (tetracyclines, fluoroquinolones and sulphonamides) within 15 min. It enabled both the qualitative determination with the naked eye under UV light and the quantitative detection of target antibiotics by scanning the fluorescence intensity of the detection probes on the corresponding detection lines. For qualitative analysis, the cut-off values for tetracyclines (TCs), fluoroquinolones (FQs) and sulphonamides (SAs) were 3.2 ng/ml, 2.4 ng/ml and 4.0 ng/ml, respectively, which were much lower than the maximum residue limit in food. For quantitative analysis, these ranged from 0.06 to 6.85 ng/ml for TCs, 0.03-5.14 ng/ml for FQs, and 0.04-4.40 ng/ml for SAs. The linear correlation coefficients were higher than 0.97. The mean spiked recoveries ranged from 92.1 to 106.2% with relative standard deviations less than 8.75%. Among them, the three monoclonal antibodies could recognize four types of TCs, seven types of FQs and 13 types of SAs, respectively, and the detection range could cover 24 antibiotic residues with different structural formulations. The results of the detection of antibiotic residues in real samples using this method were highly correlated with those of high performance liquid chromatography (R 2 > 0.98). The accuracy and precision of the EuNPs-FIA also met the requirements for quantitative analysis. These results suggested that this multiplex immunoassay method was a promising method for rapid screening of three families of antibiotic residues.

EuNPs-mAb fluorescent probe based immunochromatographic strip for rapid and sensitive detection of porcine epidemic diarrhea virus.

Porcine epidemic diarrhea (PED), induced by porcine epidemic diarrhea virus (PEDV) causes acute diarrhea, vomiting, dehydration and high mortality in neonatal piglets, resulting in significant economic losses in the pig industries. In this study, an immunochromatographic assay (ICA) based on a EuNPs-mAb fluorescent probe was developed and optimized for rapid detection of PEDV. The limit of detection (LOD) of the ICA was 0.218 µg/mL (2.725 × 103 TCID50/mL) and its linear detection range was 0.03125-8 µg/mL (3.91 × 102-105 TCID50/mL). The ICA was also validated for the detection of PEDV in swine stool samples. 60 swine stool samples from southern China were analyzed by the ICA and RT-PCR, and the results showed that the coincidence rate of the ICA to RT-PCR was 86.67%, which was significantly higher than that of AuNPs based ICA. The ICA is sensitive and specific and can achieve on-site rapid detection of swine stool samples. Therefore, the ICA has a great potential for PED diagnosis and prevention.

Cross-Subtype Detection of HIV-1 Capsid p24 Antigen Using a Sensitive Europium Nanoparticle Assay.

Accurate and early detection of diverse HIV-1 subtypes using currently available p24 antigen assays have been a major challenge. We report the development of a sensitive time resolved fluorescence (TRF) europium nanoparticle immuno assay for cross subtype detection of p24 antigen using broadly cross-reactive antibodies. Several antibodies were tested for optimal reactivity with antigens of diverse HIV-1 subtypes and circulating recombinant forms. We tested HIV strains using this assay for sensitivity and quantification ability at the pico-gram per millilter level. We identified two broadly cross-reactive HIV-1 p24 antibodies C65690M and ANT-152, which detected all strains of HIV tested. These two antibodies also yielded a better signal to cutoff ratio for the same amount of antigen tested in comparison to a commercial assay. Using an appropriate combination of C65690M and ANT-152 p24 antibodies capable of detecting all HIV types and highly sensitive TRF-based europium nano particle assay platform, we developed a sensitive p24 antigen assay that can detect HIV infection of all HIV subtypes and may be useful in early detection.

Sensitive detection of influenza viruses with Europium nanoparticles on an epoxy silica sol-gel functionalized polycarbonate-polydimethylsiloxane hybrid microchip.

In an effort to develop new tools for diagnosing influenza in resource-limited settings, we fabricated a polycarbonate (PC)-polydimethylsiloxane (PDMS) hybrid microchip using a simple epoxy silica sol-gel coating/bonding method and employed it in sensitive detection of influenza virus with Europium nanoparticles (EuNPs). The incorporation of sol-gel material in device fabrication provided functionalized channel surfaces ready for covalent immobilization of primary antibodies and a strong bonding between PDMS substrates and PC supports without increasing background fluorescence. In microchip EuNP immunoassay (µENIA) of inactivated influenza viruses, replacing native PDMS microchips with hybrid microchips allowed the achievement of a 6-fold increase in signal-to-background ratio, a 12-fold and a 6-fold decreases in limit-of-detection (LOD) in influenza A and B tests respectively. Using influenza A samples with known titers, the LOD of influenza µENIA on hybrid microchips was determined to be ~10(4) TCID50 titer/mL and 10(3)-10(4) EID50 titer/mL. A comparison test indicated that the sensitivity of influenza µENIA enhanced using the hybrid microchips even surpassed that of a commercial laboratory influenza ELISA test. In addition to the sensitivity improvement, assay variation was clearly reduced when hybrid microchips instead of native PDMS microchips were used in the µENIA tests. Finally, infectious reference viruses and nasopharyngeal swab patient specimens were successfully tested using µENIA on hybrid microchip platforms, demonstrating the potential of this unique microchip nanoparticle assay in clinical diagnosis of influenza. Meanwhile, the tests showed the necessity of using nucleic acid confirmatory tests to clarify ambiguous test results obtained from prototype or developed point-of-care testing devices for influenza diagnosis.