The Effects of Sample Matrices on Immunoassays to Detect Microcystin-LR in Water

Abstract

Immunoassays are widely used biochemical techniques to detect microcystins in environmental samples. The use of immunoassays for the detection of microcystins is vulnerable to matrix components and other interferents. This study is an evaluation of the effects of interfering substances commonly found in drinking and ambient water samples using commercially-available immunoassay kits for microcystin toxins. The microplate and strip test immunoassay formats were tested in the study. For the microplate ELISA, the following were found to inhibit microcystin-LR (MC-LR) detection: 250 μg/mL Ca 2+ or Mg 2+ , 0.01% ascorbic acid, 0.1% EDTA chelating agent, 0.05 M glycine-HCl, pH 3. The following exhibited no effect: sodium chloride (NaCl, 1% to 4%) and sodium thiosulfate (0.001% and 0.01%), 0.01 to 0.1 M phosphate buffers (PB), pH 7 and 0.067 M PB at pH 5, 6, 7 and 8. Overall, up to 50 μg/mL of standard and reference natural organic matter (NOM) from various sources did not interfere in the assay system (without MC-LR) but diminished the detection of MC-LR at varying degrees. This is the first study evaluating standard and reference humic and fulvic acids from various sources in immunoassays for microcystins. The strip test also showed variable effects on MC-LR detection in the presence of NOM. This assay format was also sensitive to varying pHs and ionic strengths. MC-LR binding was inhibited at low pH (0.05 M glycine-HCl, pH 3), whereas, 0.067 M PB with pH 6, 7 and 8 can yield false positive results. Lower ionic strength of 0.01 M PB, pH 7 showed no interference in MC-LR binding whereas higher ionic strengths can interfere with MC-LR detection. NaCl at 3% and 4% can interfere with the analysis giving false positive results. Mg 2+ at 50 and 250 μg/mL showed no effect on the analysis while the same concentration of Ca 2+ can yield false positive results. The performance in marine, brackish and hard waters should be tested given the potential sensitivity to salinity. Results of this study may assist in the further refinement of existing assays and the development of practical antibody-based methods to clean-up samples and detect cyanotoxins in water.

Direct Electrochemical Nano‐immunosensor for Microcystin‐LR in Seawater

“…Common potential interferents in water sample analysis , such as PBS (0.01 mol L −1 ), sodium chloride (0.4 mol L −1 ), seawater, sodium sulfate (0.03 mol L −1 ), calcium chloride (1.3×10 −3 mol L −1 ), potassium chloride (9.4×10 −3 mol L −1 ), sodium bicarbonate (2.4×10 −3 mol L −1 ) and potassium bromide (8.2×10 −4 mol L −1 ), were selected for the study on potential interference in the determination of MC‐LR. The results (Figure ) showed a low degree of interference for all potential interferents tested in relation to the control test (less than 10 %), indicating that it is possible to perform the determination of MC‐LR in the presence of these substances.…”

On-chip sample preparation and analyte quantification using a microfluidic aqueous two-phase extraction coupled with an immunoassay

“…Immunoassays are powerful bioanalytical techniques that have been used extensively in clinical diagnosis for decades, either performed in a laboratory [1][2][3] or, more recently, integrated into portable devices. 4,5 Immunoassay applications have been successfully broadened to environmental analysis, 6,7 with a significant and increasing share targeted to mycotoxin quantification in feed and foodstuff. [8][9][10] However, immunoassays suffer from significant matrix interference in samples such as blood, 11 urine, 12 water, 6 and food extracts.…”

“…4,5 Immunoassay applications have been successfully broadened to environmental analysis, 6,7 with a significant and increasing share targeted to mycotoxin quantification in feed and foodstuff. [8][9][10] However, immunoassays suffer from significant matrix interference in samples such as blood, 11 urine, 12 water, 6 and food extracts. [13][14][15][16][17] This interference is responsible for a pronounced decrease in sensitivity and inter-assay reproducibility, usually requiring sample treatment steps prior to analysis, which are often complex and time-consuming.…”

Lake browning counteracts cyanobacteria responses to nutrients: Evidence from phytoplankton dynamics in large enclosure experiments and comprehensive observational data

“…This would suggest that light limitation caused by browning altered the resource allocation of cyanobacteria compared to the situation in clear lakes (Kardinaal et al., 2007). It is also possible that microcystin produced in those conditions was adsorbed to humic substances, leading to an underestimation of toxin production by our ELISA analyses (De la Cruz et al., 2012). However, such matrix effects were unlikely to be important, because the concentrations of humic substances in our experiments were well below the threshold of 10 mg L −1 at which the ELISA test kit manufacturer identified potential for assay interference.…”