Anna Oubiña

Development of a highly sensitive enzyme-linked immunosorbent assay for atrazine Performance evaluation by flow injection immunoassay

Specific polyclonal antibodies to the herbicide atrazine (6-chloro-N-ethyl-N′-isopropyl-1,3,5-triazine-2,4-diamine) have been raised by immunizing three New Zealand rabbits. With the antisera (As) a highly sensitive enzyme-linked immunosorbent assays (ELISA) has been developed to determine atrazine in water samples. Several usable competitive immunoassays have been obtained by screening a battery of nine enzyme tracers (ETs) and three antisera. The optimized ELISA presents an IC50 of 0.28 nM (60 ng l−1) and a detection limit of 0.043 nM (9 ng l−1). Cross-reactivity studies have proved that the immunoassay is specific for atrazine while other triazine compounds are only detected on a minor extent. The flow injection immunoanalysis (FIIA) method has an IC50 of 2 nM (0.47 μg l−1) reaching a detection limit of 0.35 nM (75 ng l−1). The performance of both methods has been evaluated by analyzing water samples containing mixtures of atrazine and other pesticides at the ppb level. For this purpose two candidate reference materials have been used (A and B) and a spiked sample stored on Empore disks (sample C). A close correspondence was found between the results obtained with both immunochemical techniques.

Determination of pentachlorophenol in certified waste waters, soil samples and industrial effluents using ELISA and liquid solid extraction followed by liquid chromatography

The Pentachlorophenol RaPID-ELISA was compared with on-line liquid solid extraction (LSE) followed by liquid chromatography (LC) for the monitoring of pentachlorophenol (PCP) in certified waste waters, a waste water sample from a pulp industry, a soil sample from Brazil and a certified reference material. The water pH (1 to 9), and two cross-reactants (2,3,5,6-tetrachlorophenol and 2,3,4,6-tetrachlorophenol) were also evaluated to know if these parameters could affect the enzyme linked immunosorbent assay (ELISA) determinations. Water samples were spiked with certified material provided by Aquacheck, containing pentachlorophenol and 2,4,6-trichlorophenol at concentration levels ranging from 1 to 10 μg 1−1. Good correlations were found between the data obtained by ELISA and from on-line LSE-LC using PLRP-S sorbent and UV detection. A dried contaminated industrial soil used as certified reference material from the Community Bureau of Reference (BCR) of the European Commission (EC), a soil sample containing pentachlorophenol at low level (μg g−1) and a waste water sample from a pulp industry were also studied using both techniques. Moreover, the confirmation of all analytes present in the waste water was carried out by liquid chromatography-mass spectrometry (LC-MS) using atmospheric pressure interface (APCI). The presence of unknown cross-reactants in the waste water affected the ELISA determinations.

Detection of endocrine-disrupting pesticides by enzyme-linked immunosorbent assay (ELISA): application to atrazine

Abstract An overview of biological and toxicological effects of relevant endocrine-disrupting compounds is given. Special attention is paid to the determination of atrazine, a relevant pesticide that is considered an endocrine disrupter, by ELISA.

Sensitivity of selected bacterial species to UV radiation

The effect of exposure of bacterial suspensions to UV radiation by means of the dose-response curves was assessed. The D37 and D10 values were used for subsequent statistical analysis of the results. The aim of this article is to evaluate the sensitivity to UV radiation of several microorganisms of different habitats (Rhizobium meliloti, Rhodobacter sphaeroides, Escherichia coli, and Deinococcus radiodurans), two mutants with nonfunctional SOS DNA repair system (R.meliloti recA- and E. coli recA-), and a mutant in the synthesis of carotenoids (R. sphaeroides crtD). The results reveal that D. radiodurans was an extremely resistant bacterium, R. meliloti was more resistant than R. sphaeroides, and E. coli was the most sensitive bacterium tested. The high sensitivity of recA- mutants was also verify. Moreover, it seems that the possession of pigments had no important effect in the sensitivity of R. sphaeroides to UV radiation.

Determination of PAHs in river water samples by ELISA

Abstract A commercially available enzyme-linked immunosorbent assay (ELISA) kit for carcinogenic polycyclic aromatic hydrocarbons (PAHs) has been evaluated for the trace determination of PAHs in river water samples. The data reported for the total concentration values of PAHs were around one order of magnitude higher using ELISA than the results obtained using conventional GC–MS techniques. The ELISA kit produced no false negative determinations for the different water samples analyzed.

Multianalyte effect in the determination of cross-reactivities of pesticide immunoassays in water matrices

Abstract An extension of Abrahams method to calculate cross-reactivity (CR) is presented. Over a wide concentration range the response curves of a cross-reactant were compared with those of a standard applied to four immunoassay commercial kits (benomyl/carbendazim, chlorothalonil, metolachlor and parathion-ethyl). Various doses of cross-reactants (0–1000 μg/l) were tested in the presence of various doses of the standard (0–100 μg/l). The results showed that the percentage of the cross-reactivity (%CR) decreased as a function of the concentration of cross-reactant present in the water and high %CR values were found in comparison with those reported in commercial kits. This indicates a better sensitivity for many analytes as compared to values actually reported in the ELISA (Enzyme-Linked ImmunoSorbent Assay) kits. Applications of the CR studies are also presented that should help in the acceptance or rejection of a concentration range of cross-reactants in a real sample. When determining the CR in estuarine waters, the matrix effect of estuarine water underestimated the true extent of cross-reactivity of the standard, making it necessary to calculate for cross-reactivity to compensate for matrix effect.

Performance of two immunoassays for the determination of atrazine in sea water samples as compared with on-line solid phase extraction-liquid chromatography-diode array detection

Two immunoassay formats, magnetic particles-based assay (Atrazine RaPID assay and Atrazine High-Sensitivity RaPID assay) and microtiter plate based assay (Department of Entomology and Environmental Toxicology, University of California in Davis) were evaluated for the determination of atrazine in sea water samples. The results obtained were compared and validated with those obtained by using on-line solid phase extraction followed by liquid chromatography-diode array detection (on-line SPE-LC-DAD). The correlation between both techniques was good when analyzing levels of atrazine ranging from 0.01 to 5 μg/l in samples showing salt concentration values varying from 0 to 35 g/l and pH values from 2 to 10. One of these immunoassays (Atrazine High-Sensitivity RaPID assay) was employed to directly analyze atrazine in real estuarine and coastal water samples. The same samples were analyzed after filtration and C18 Empore disks extraction.

Monitoring of Insecticides and Fungicides in Water and Sediment Samples in the Brazilian Environment

Abstract Immunoassay techniques, either microtiter plate or magnetic particle-based ELISA and various gas chromatographic techniques (GC-ECD, GC-NPD and GC-MS) were applied to the determination of priority pesticides (chlorothalonil, metalaxyl, carbendazim, parathion-methyl and folpet) in water and sediment samples from Paty do Alferes at Rio de Janeiro State (Brazil). Water samples were directly analyzed in the field by immunoassay techniques and transported to Barcelona after being stored in SPE disks. Sediment samples were frozen and transported to Barcelona for further analysis. All results were confirmed by GC-MS under negative chemical ionization and using SIM and the protocol was extended to other pesticides like naled, fenitrothion and azinphos-ethyl that could not be measured with ELISA. The results showed that the majority of the water samples had pesticide levels below 5 ng/l with the exception of chlorothalonil and folpet that showed values up to 114 and 210 ng/l, respectively. The sediment sa...

A Master Analytical Protocol for Determining a Broad Spectrum of Organic Pollutants in Industrial Effluents

Contaminated industrial effluents often contain a variety of organic pollutants which are difficult to analyze by standard GC/MS methods since often miss the more polar or nonvolatile of these organic compounds. The identification of highly polar analytes by chemical or rapid biological techniques is needed for characterization of the effluents. By correlating chemical characterization with toxicity a better picture of the effluent can be achieved.