Sensitive Direct Enzyme-linked Immunosorbent Assay Research Articles

Development of a monoclonal antibody-based, congener-specific and solvent-tolerable direct enzyme-linked immunosorbent assay for the detection of 2,2′,4,4′-tetrabromodiphenyl ether in environmental samples

A sensitive direct enzyme-linked immunosorbent assay (ELISA) for the specific detection of 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) in environmental samples was developed. A hapten mimicking BDE-47 was synthesized by introducing a butyric acid spacer into 5-hydroxy-BDE-47 and coupled to keyhole limpet hemocyanin to form an immunogen for the production of monoclonal antibodies (Mabs) against BDE-47. The most sensitive direct ELISA was formatted with a Mab, designated as 4F2, in combination with 5-(2,4-dibromophenoxy)pentanoic acid peroxidase as a tracer. The inhibition half-maximum concentrations and limit of detection of BDE-47 in phosphate buffered saline with 25% DMSO were 1.4 ± 0.05 and 0.1 ng mL(-1), respectively. Cross-reactivity values of the ELISA with the tested BDE congeners and metabolites were ≤5.8%. This assay was used to determine BDE-47 in soil, sediment and house dust samples after ultrasonic extraction, simple cleanup and concentration steps. The average recoveries, repeatabilities (intraday extractions and analyses), and intra-laboratory reproducibilities (interday extractions and analyses) were in a range of 92-126%, 8-19% and 9-25%, respectively. Applied to 44 real samples, the results of this assay displayed a statistically significant correlation with those of a gas chromatography-mass spectrometry method (R(2)=0.79-0.85), indicating this ELISA is a suitable tool for environmental analyses of BDE-47.

ELISA for Aging Biomarkers Induced by Telomere Dysfunction in Human Plasma

Background. We identified cathelicidin related antimicrobial protein (CRAMP) secreted from telomere dysfunctional bone marrow cells of late generation telomerase knockout mice (G4mTerc−/−), increased in blood and various tissues. It can represented human aging and disease. The main aim of this study is to investigate the sensitive direct enzyme-linked immunosorbent assay (ELISA) method to analyze the human aging and disease in plasma and the detailed methods to quantify the direct ELISA of these aging biomarkers. Methods. Telomere lengths of 50 healthy persons are measured with real-time PCR in blood cells. Plasma samples from all subjects are analyzed using direct ELISA. Results. From 25 years old person to 78 years, the telomere length becomes shorter during aging. In blood plasma, the expression levels of CRAMP increases during human aging. There is the reverse correspondence between the telomere length and the plasma CRAMP level. We also find that the fresh plasma, the frozen plasma which thawed less than 3 times, and the plasma kept in the room temperature less than 3 hours are better for the ELISA analyze of CRAMP in the plasma. Conclusion. This CRAMP ELISA could become a powerful tool for investigating the relationship between human aging and telomere length shortening.