Characterisation of fish leucocytes : an immunocytochemical and functional study in carp (Cyprinus carpio L.)
A panel of monoclonal antibodies (MAbs) against carp serum immunoglobulin (Ig), WCIs or carp thymocytes (T), WCTs were used for the characterisation of carp leucocytes. Unfortunately, all WCTs and some WCIs react with common carbohydrate determinants present on all leucocytes and Ig. Most WCIs react specific with protein determinants at the heavy chain of Ig. Consequently, B lymphocyte (sub) populations, plasma cells and Ig-binding cells could be studied. Ig molecules are found in clusters at the cell membrane of B cells and plasma cells, and in contrast to mammalian plasma cells, most carp plasma cells still have Ig at their surface membrane. Mainly the dull surface Ig-positive (sIg +) cells were stimulated by the mammalian B cell mitogen LPS and not by PHA (T cell mitogen) in vitro , whereas the sIg-negative (sIg -) cells were stimulated by PHA and not by LPS. The percentages of B cells and plasma cells showed an increase during ontogeny and reached a plateau at about 3 months and 8 months of age respectively. It is suggested that full development of the carp (humoral) immune system needs at least 8 months (at 21-22 °C). Three different subpopulations of B cells and plasma cells and at least two Ig isotypes can be distinguished based upon their reactivity with WCI 4 arid WCI 12. The distribution of the three B cell subpopulations appeared to be organ and age dependent which indicates functional differences between the Ig isotypes. Fc-like receptors were mainly demonstrated on gut macrophages while pronephros macrophages and neutrophilic granulocytes did not show Ig binding. Consequently, other forms of antigen opsonisation (e.g. complement) may play a role in phagocytosis by these non Ig-binding cells. Several procedures were tested for obtaining MAbs specific for Ig -lymphoid cells. It is concluded that the presence of immunodominant carbohydrate determinants is the major problem for obtaining specific MAbs. Tolerisation of mice against these determinants or the use of isolated membrane lysates from (sIg -) PBL appeared promising but till now only specific thrombocyte markers have been obtained. The use of more purified antigen is recommended in further attempts. The data presented in this thesis can be used for fundamental studies on cell interactions in the immune response, but also for more applied investigations on fish health control.
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| Format: | Doctoral thesis biblioteca |
| Language: | English |
| Published: |
Landbouwuniversiteit Wageningen
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| Subjects: | antibodies, blood, blood plasma, blood serum, carp, cyprinidae, erythrocytes, fibrin, immunocytochemistry, immunoglobulins, leukocytes, platelets, reticuloendothelial system, antilichamen, bloed, bloedplaatjes, bloedplasma, bloedserum, erytrocyten, fibrine, immunocytochemie, immunoglobulinen, karper, leukocyten, reticulo-endotheliaal systeem, |
| Online Access: | https://research.wur.nl/en/publications/characterisation-of-fish-leucocytes-an-immunocytochemical-and-fun |
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| Summary: | A panel of monoclonal antibodies (MAbs) against carp serum immunoglobulin (Ig), WCIs or carp thymocytes (T), WCTs were used for the characterisation of carp leucocytes. Unfortunately, all WCTs and some WCIs react with common carbohydrate determinants present on all leucocytes and Ig. Most WCIs react specific with protein determinants at the heavy chain of Ig. Consequently, B lymphocyte (sub) populations, plasma cells and Ig-binding cells could be studied. Ig molecules are found in clusters at the cell membrane of B cells and plasma cells, and in contrast to mammalian plasma cells, most carp plasma cells still have Ig at their surface membrane. Mainly the dull surface Ig-positive (sIg +) cells were stimulated by the mammalian B cell mitogen LPS and not by PHA (T cell mitogen) in vitro , whereas the sIg-negative (sIg -) cells were stimulated by PHA and not by LPS. The percentages of B cells and plasma cells showed an increase during ontogeny and reached a plateau at about 3 months and 8 months of age respectively. It is suggested that full development of the carp (humoral) immune system needs at least 8 months (at 21-22 °C). Three different subpopulations of B cells and plasma cells and at least two Ig isotypes can be distinguished based upon their reactivity with WCI 4 arid WCI 12. The distribution of the three B cell subpopulations appeared to be organ and age dependent which indicates functional differences between the Ig isotypes. Fc-like receptors were mainly demonstrated on gut macrophages while pronephros macrophages and neutrophilic granulocytes did not show Ig binding. Consequently, other forms of antigen opsonisation (e.g. complement) may play a role in phagocytosis by these non Ig-binding cells. Several procedures were tested for obtaining MAbs specific for Ig -lymphoid cells. It is concluded that the presence of immunodominant carbohydrate determinants is the major problem for obtaining specific MAbs. Tolerisation of mice against these determinants or the use of isolated membrane lysates from (sIg -) PBL appeared promising but till now only specific thrombocyte markers have been obtained. The use of more purified antigen is recommended in further attempts. The data presented in this thesis can be used for fundamental studies on cell interactions in the immune response, but also for more applied investigations on fish health control. |
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