Development of optical bio-sensing methods for detection of toxins produced by algae.

The main aim of this work is to develop an optical method for the detection of microcystin-LR, one of the most dangerous toxins released by cyanobacteria. Photosynthetic cyanobacteria (also known as blue-green algae) exist in any type of water (including drinking water) and possess a serious threat to humans and animals, and generally to the environment. Microcystin-LR (MC-LR) is perhaps the most toxic from a large family of cyanotoxins; it is known to cause liver damage and also acts as a carcinogen. The World Health Organization has set the limit of 1pg/l for MC-LR in drinking water. However existing detection methods, such as ELISA, cannot provide such high sensitivity. The relatively low molecular weight of MC-LR (995.2 m/mol) makes it difficult to detect using conventional QCM and SPR based analytical methods. Therefore, the development of highly sensitive, reliable, and (at the same time) inexpensive and easy-to-use methods of detection of microcystin is of very high importance today. The method of Total Internal Reflection Ellipsometry (TIRE) is particularly attractive for the above task considering its high sensitivity and particular suitability for detection of low molecular weight analytes. In this work MC-LR was detected in direct immunoassay with MC10E7 monoclonal antibodies raised against MC-LR in mouse, using TIRE as a detection method. Also the TIRE method was used to study the protein- protein interaction (the protein chaperon with its receptors in the chloroplast) and to detect other types of toxins such as Aflatoxin B1.At first, the TIRE immunoassay was calibrated using aqueous solutions with known concentration of MC-LR. The study of binding kinetics of MC-LR to MC10E7 yielded the association constant close to 108 l/mol which is typical for highly specific immune reactions. The detection limit for MC-LR between 0.1 ng/ml and lng/ml was achieved using the TIRE detection. Then, the concentration of the MC-LR toxin produced naturally by algae microcystis aeruginosa was evaluated using calibration data obtained for solutions spiked with known concentrations of MC-LR. The role of environmental factors (temperature, pH, nutrition contents, and salinity) on the efficiency of production of MC-LR by microcystis aeruginosa was studied. Purification of solutions contaminated with MC-LR (both commercial and naturally produced) was achieved using MnCO[3] microparticles coated with polyelecrolytes and functionalised with MC10E7 antibodies as an absorbent for MC-LR. Also, the concentration of MC-LR in solutions was reduced as a result of direct electrolysis.