Využití bioluminiscenční bakterie Escherichia coli pro stanovení aktivity komplementového systému kaprovitých ryb
| Title in English | The application of bioluminescent Escherichia coli for complement system activity assay in Cyprinidae fish |
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| Authors | |
| Year of publication | 2008 |
| Type | Conference abstract |
| MU Faculty or unit | |
| Citation | |
| Description | The complement system plays an essential role in humoral innate immune response in fish. Together with phagocytosis and systemic inflammation composes main part of the non-specific immunity. Complement system is composed of more than 30 proteins and three pathways of its activation have been described in fish: classical, alternative and possible lectine. The total bacteriolytic activity (TA) including all pathways and activity of alternative pathway (AP) only were assessed in serum and plasma of common carp, Cyprinus carpio, European chub, Leuciscus cephalus and barbel, Barbus barbus, all field collected species belong to Cyprinidae family. The principle of luminometric method described previously on salmonids is based on detection of light production by bioluminescent recombinant strain of Escherichia coli. This Gramm-negative bacterium is sensitive to lysis caused by complement system, so there is a correlation between viability of bacteria and light production. Continuous measurement of bacterial bioluminescence occured 3 hours at laboratory temperature. Time needed for 50% killing of bacteria by sample was used as the measured parameter. The bacteriolytic activity was detectable in both TA and AP. More reliable results were obtained by TA because 50% decrease caused by AP was not always detectable. To optimise the method for exact fish species, different concentration of sample were tested (TA: 0, 350 microlitre/ml; AP: 0 , 400 microlitre/ml). Selected concentrations were used for following analysis. The plasma sample of concentration 100 microlitre/ml needed 1,48 hours for 50% killing of bacteria in Barbus barbus, and 1,97 hours in Leuciscus cephalus respectively. Plasma sample of Cyprinus carpio needed higher concentration of plasma 300 microlitre/ml to reach similar values. |
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