Microcystin-LR Enrichment from Freshwater by a Recombinant Plant-derived Antibody Using Sol-Gel-Glass Immunoextraction

Abstract

Eutrophication of water bodies can promote cyanobacterial (blue-green algae) blooms, which has become a source of increasing concern for both recreational and drinking water use. Many bacterial species can produce toxins that pose threats to wildlife, domestic animals and humans. Microcystin-leucine-arginine (MC-LR) is the most frequent and most toxic microcystin congener. For the first time, lab-scale investigations were performed to test the application of a recombinant plant-derived anti-MC-LR antibody immobilized on an immunoaffinity support material to selectively extract the toxin from spiked freshwater samples. As a comparison, its hybridoma-derived counterpart (murine monoclonal antibody) was evaluated. The antibody-doped material was prepared via an optimized sol-gel process; its stability and binding efficiency of MC-LR in spiked freshwater samples were thoroughly tested using the ELISA and orthogonal LC-MS methods. For removal, two column-based procedures with sequential or continuous cyclic sample addition and a suspension mode (moving adsorbent) were tested. Noteworthy the results obtained with a crude antibody fraction were fully compatible with the highly purified preparation. This study paves the way for further investigation being focused on novel applications of plant-derived anti-MC-LR antibodies in bioremediation to selectively deplete the toxin from freshwater: a green and promising technology without secondary pollution.

Recombinant antibodies and their use for food immunoanalysis

“…The chimeric antibody was applied to the development of a lateral flow immunoassay for the detection of [Arg4]-MCs in the range of 100-300 ng/L in freshwater samples. In a different approach, the plant-derived antibody encapsulated in a sol-gel matrix was applied to the removal of MC-LR from freshwater samples in lab-scale experiments [118]. The authors point out that plant-based and microbial expression systems are attractive alternatives for the production of recombinant antibodies at large quantities and low cost.…”

Immunoaffinity Extraction and Alternative Approaches for the Analysis of Toxins in Environmental, Food or Biological Matrices

“…However, the resulting non-covalent binding is quite strong under physiological conditions but can be easily disrupted by decreasing the pH of the surrounding solution. The sol-gel method can also be used to entrap Abs [73,89]. In this case, Abs are then immobilized in the pores of a hydrophilic glass matrix that reduces the non-specific adsorption of apolar analytes.…”

“…The proportion of active antibodies can be deduced from the experimentally determined capacity value. As an example, values of 39% or 65% of active antibodies were reported for ISs prepared by the grafting of a poly(GMA-co-EGDMA) monolith [93] or sol-gel entrapment [73], respectively. Concerning laboratory-made ISs, there are only a few studies that give data about the repeatability of the preparation of ISs [71].…”

“…ISs constitute a good mean to concentrate the targeted toxin(s) while removing matrix effect thus allowing the analysis of the extract with simple and fast analytical methods adapted to numerous samples such as bioassays achieved in 96-well plates (ELISA or enzymatic inhibition assay) [16,73,74,77,98] as it was performed mainly for microcystin analysis. In addition, it was shown by Chiavaro et al that the high selectivity of the IS allows the direct determination of AFs B 1 and M 1 at 1 µg/kg in pig liver extracts using only fluorescence detection [17].…”

Flutuação temporal de cianotoxinas (Microcistina) no rio Tapajós (Santarém, Amazônia-Brasil)

“…A comprovação da toxicidade de uma população de cianobactérias em um determinado corpo d'água não implica necessariamente perigo ambiental e humano enquanto as células permanecerem finamente dispersas, todavia o desenvolvimento massivo, com formação de escuma na superfície representam riscos para a saúde [22]. As cianotoxinas em águas de uso recreativo podem constituir um perigo grave para a saúde humana devido à exposição por ingestão acidental de água contaminada, inalação e por contato dérmico [23,24,25,26]. O valor tolerado para exposição recreativa é de no máximo 10 µg.L -1 para MC-LR [22,27].…”