Stanovení vybraných parametrů imunitního systému ryb
| Title in English | Assesment of selected fish immune parameters |
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| Authors | |
| Year of publication | 2009 |
| Type | Conference abstract |
| MU Faculty or unit | |
| Citation | |
| Description | Fish immune system cen be divided into cellular and humoral part. Nonspecific cellular part is made by phagocytes and cytotoxic cells, specific by T and B lymphocytes. Nonspecific humoral immunity composes mainly of lectins, complement system and lytic enzymes, e.g. lysozyme, specific immunity is characterized by presence of antibody class IgM. At our department we measure oxidative burst of phagocytes in whole blood, activity of complement in plasma and lysozyme concentration in skin mucus. Oxidative burst of phagocytes is measured by luminescence method. Whole heparinized blood is dilluted and mixed with luminol and activator, which binds to phagocytes receptors and causes its activation. The most important indicators of phagocyte oxidative burst are peaks of luminescence curve, the time of its achievement and total amount of produced reactive oxygen metabolites. Complement activity is assessed by luminescent method in plasma. Uses recombinant bioluminescent bacterial strain of E. coli. The light emission requires presence of ATP produced only by living cells. Intensity of produced light correlates with bacteria viability. From kinetic curve of luminescence the time needed for 50% killing was evaluated. Lysozyme determination is possible in vitro by radial diffusion in agarose. Samples of skin mucus or calibration solution of lysozyme are applied on agarose plates containing Micrococcus luteus. Diameters of bright diffusion zones are converted according calibration curve to mg/ml of sample. Amount of antibodies IgM in fish plasma can be assessed by precipitation with zinc sulphate, which specifically dehydrates proteins, which came out from solution and can be removed by centrifugation. The amount of IgM in the sample (in g/l) is calculated as the difference between total proteins and protein content in the supernatant after precipitation and centrifugation. |
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