Monoclonal antibodies specific to haemocytes of black tiger prawn Penaeus monodon (original) (raw)
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Characterisation of shrimp haemocytes and plasma components by monoclonal antibodies
Journal of Cell Science, 1995
Various haemolymph components of the shrimp Penaeus japonicus were identified and characterised by monoclonal antibodies. Three groups of monoclonal antibodies were raised. Their reactivity to haemocyte types and/or secreted molecules was determined by immunofluorescence and the molecular masses of the antigens were analysed by western-blotting. A 170 kDa protein, in reducing conditions, was recognized by four panhaemocytic monoclonal antibodies from group 1. This protein was present both in the plasma and in the haemocytes from which it appears to be secreted. The shrimp haemocytes were separated by isopycnic centrifugation on a Percoll gradient and the different subpopulations were antigenically analysed using the two monoclonal antibodies, 40E2-2A and 40E10-2B, from group 2. The granular cells were labelled by 40E2-2A which was specific for a protein of 142 kDa also present in plasma. By comparison, the 40E10-2B monoclonal antibody was assumed to be the marker for small hyaline a...
Binding of immunoglobulins and immune complexes to erythrocytes of vertebrates
Immunochemistry, 1978
Previous mvestigations which have referred to red cell Fc receptor m rabbit. guinea pig. sheep and man. have been extended to other vertebrates. such as reptiles. amphihia. birds and mammalians. In these investigations. red cells from different animal spectes have been analyzed. As ligand DNP-BSA anti-DNP non-precipitating antibody, aggregated IgG by the bis-diazotized henzidine method. IgM 7s and non-modified IgG. IgA and IgM were used. The ligand binding to red cells was detected by Coomhs test with specific anti-immunoglohulin serum. The analyzed red cells showed exposed Fc receptor. In the case of human red cells. to render such receptor evident it is necessary to submit the cells to trypsinization. All the analyzed erythrocytes hind IgM 7s. Ab-Ag complex and aggregated IgG. and some of these cells hind non-modified immunoglobulins. * Members of the Scientific Researcher Career. National Research Council of Argentina. MATERIALS AND METHODS t+~~rhroc~~~lc~.s o/' tliffiwnt nnimul ~spccics These were obtained from heparinized blood. The cells were washed 5 times with 0. I5 M NaCI. 0.01 M phosphate, pH 7.6. phosphate buffered saline (PBS) before using. These were obtained by the Morton and Pickles (1947) method. The trypsin treatment time was carried out for 10 min. Antigen-antihocij~ wntple.ws ! Ag-Ah J These were prepared by mixing 2 mg ml of antibody w'ith 250 ng,ml of antigen.
Journal of Invertebrate Pathology, 2012
Monoclonal antibodies (MAbs) to hemocytes of mud crab, Scylla serrata, were produced by immunizing mice with formalin-fixed hemocytes. Of the six MAbs produced, two (MAb 1D11 and MAb 1A2) reacted specifically with hemocyte proteins in western blot. MAb 1A2 showed strong immunofluorescent reaction with granular hemocytes and a weak reaction with semigranular cells. However, hyaline cells did not show any reaction. The MAbs also showed strong cross-reactivity with the hemocytes of different crab species but not with other crustaceans. The MAbs produced can be used as a marker for granular hemocytes of crabs.
Aquaculture, 1999
The current status on the antibodies that have been produced against teleost fish immuno-Ž . globulins, immunoglobulin-bearing cells B-cells , thymocytes and T-cells is reviewed. An updated list of monoclonal antibodies is reported together with their antigen specificity, their use in the detection of cross-reacting cells, and their eventual use as markers of fish leucocyte subpopulations. The evidence obtained with monoclonal antibodies on the ontogeny of lymphoid cells and functions of lymphocyte subpopulations is presented. The possibility to create a common nomenclature for fish monoclonal antibodies is discussed. q 0044-8486r99r$ -see front matter q 1999 Elsevier Science B.V. All rights reserved.
Differentiation and Proliferation of B lymphocytes in a Culture of Carp Hematopoietic Cells
Fish Pathology, 1998
Cultivation of cells from carp (Cyprinus carpio) kidney tissues including hematopoietic tissues resulted in the formation of a feeder-like cell layer. The carp hematopoietic cells were found to grow in contact with the cell layer. Numerous non-attaching cells were observed in the supernatant of the culture medium. These non attaching cells were constantly produced in the culture medium, half of which was renewed at 1-4 days intervals for about 1 month. To characterize the proliferating cells, we observed the non-attaching cells with a scanning electron micro scope. At a higher magnification, the cells had many short microvilli on their cell surface which is a common feature of carp lymphocytes. Immunofluorescent staining by monoclonal antibodies against carp IgM demon strated that 1.6-5.6% of the cells were surface immunoglobulin (sIg) positive and 0.1-0.5% of the cells were cytoplasmic immunoglobulin (cIg) positive. Moreover, the percentage of sIg or cIg-positive cells was main tained during the culture, even after the total number of non-attaching cells rapidly increased (more than 20 population doublings) between 10-30 days. These results suggest that immature B-lymphocytes (sIg-negative cells) differentiated into a more matured type of B lymphocytes (sIg or cIg positive cells) during the culture.
Molecular Immunology, 1981
Spleen cells of mice immunized with rabbit spleen and mesenteric lymph node (MLN) 5 cells were fused with mutant mouse myeloma cells. Twenty-six clones which react with rabbit lymphoid cells were obtained. By membrane immunofluorescence, as analysed visually and by the fluorescence-activated cell sorter, one of these clones, 9AE10, produced an antibody that reacted with nearly all thymocytes (> 90%) and with from 46 to 78% of spleen, MLN and peripheral blood lymphocytes. Double-membrane immunofluorescence with the 9AElO monoclonal antibody (MAb) and anti-Ig showed that 9AE10f and Ig' cells of spleen, MLN and peripheral blood were distinct and non-overlapping populations. Thus, the 9AElO MAb is a T-cell-specific antibody.
Haemocytes of the clam Tapes philippinarum (Adams & Reeve, 1850): morphofunctional characterisation
Fish & Shellfish Immunology, 2000
Tapes philippinarum is a bivalve mollusc of the Pacific Ocean, successfully imported for human consumption into the northern Adriatic Sea (Europe). For better knowledge of its considerable adaptive ability in comparison with similar autochthonous species, a morpho-functional characterisation of its haemocytes was carried out with the establishment of short-term cell cultures (60 min at 25 C). Various methods of cytochemical staining identified four cell types in the haemolymph: granulocytes (48•05% 1•43), hyalinocytes (32•18% 0•99), haemoblasts (18•97% 0•63) and serous cells (0•8% 0•19). The granulocytes, possessing cytoplasmic granules with di#ering dye a$nity, included basophils, neutrophils and acidophils. Such granules stained vitally with Neutral Red, and correspond to lysosomes. Hydrolytic and oxidative enzymes were mainly detectable after stimulation in the presence of yeast cells. Both granulocytes and hyalinocytes were positive for alkaline phosphatase, non-specific esterase, peroxidase, and cytochrome C oxidase, whereas only granulocytes were positive for-glucuronidase, acid esterase, and arylsulphatase. Both cell types were competent phagocytes towards yeast and plasma had an opsonising e#ect. Moreover, the respiratory burst accompanied phagocytosis with superoxide anion production, recognisable through cytoplasmic deposits of formazan after treatment with nitro blue tetrazolium. Haemoblasts were small undi#erentiated cells which, due to their morphology and positivity to the anti-CD34 antibody, show the typical features of stem cells. Serous cells, probably arising from Keber's gland and belonging to another di#erentiation pathway, contained non-sulphate acid mucopolysaccharides and play an important role in early defence mechanisms, taking part in the formation of clots.
Haemocytes of the clam Tapes philippinarum (Adams & Reeve, 1850): morphofunctional characterisation
Fish & Shellfish Immunology, 2000
Tapes philippinarum is a bivalve mollusc of the Pacific Ocean, successfully imported for human consumption into the northern Adriatic Sea (Europe). For better knowledge of its considerable adaptive ability in comparison with similar autochthonous species, a morpho-functional characterisation of its haemocytes was carried out with the establishment of short-term cell cultures (60 min at 25 C). Various methods of cytochemical staining identified four cell types in the haemolymph: granulocytes (48·05% 1·43), hyalinocytes (32·18% 0·99), haemoblasts (18·97% 0·63) and serous cells (0·8% 0·19). The granulocytes, possessing cytoplasmic granules with di#ering dye a$nity, included basophils, neutrophils and acidophils. Such granules stained vitally with Neutral Red, and correspond to lysosomes. Hydrolytic and oxidative enzymes were mainly detectable after stimulation in the presence of yeast cells. Both granulocytes and hyalinocytes were positive for alkaline phosphatase, non-specific esterase, peroxidase, and cytochrome C oxidase, whereas only granulocytes were positive for -glucuronidase, acid esterase, and arylsulphatase. Both cell types were competent phagocytes towards yeast and plasma had an opsonising e#ect. Moreover, the respiratory burst accompanied phagocytosis with superoxide anion production, recognisable through cytoplasmic deposits of formazan after treatment with nitro blue tetrazolium. Haemoblasts were small undi#erentiated cells which, due to their morphology and positivity to the anti-CD34 antibody, show the typical features of stem cells. Serous cells, probably arising from Keber's gland and belonging to another di#erentiation pathway, contained non-sulphate acid mucopolysaccharides and play an important role in early defence mechanisms, taking part in the formation of clots.
Classification of Hemocytes from Four Crustaceans and Cross-Reactivity of Their Antisera
Journal of Shellfish Research, 2018
Common antigens and analyzed the hemocytes of the lady crab (Charybdis japonica) were studied. Chinese mitten crab (Eriocheir sinensis), Dungeness crab (Cancer magister), and red king crab (Paralithodes camtschaticus) using a laser scanning confocal microscope (LSCM), flow cytometry (FCM), Western blotting (WB), and transmission electron microscopy. In this paper analyzed hemocyte classification and immunological characteristics of the whole proteins in these species and their antisera. Four antisera positively cross-reacted, to varying degrees, with hemocytes of the four species. Two subpopulations were classified in C. japonica, P. camtschaticus, and E. sinensis and three in C. magister. The LSCM results showed that four antisera were positive with the hemocytes of the four crustacean species, respectively. Dot fluorescent signals on the hemocyte membranes show a cricoid intensive distribution. FCM analysis showed a positive rate of cross-reaction, in which the reaction of the antisera with its own hemocytes was higher than that with other speciesÕ hemocytes. The positive rate of granulocytes was always higher than that of hyalinocytes. Western blotting results showed that the four antisera mainly recognized antigens with molecular weights of about 75 or 80 kDa in the hemocytes of C. japonica, 80 or 90 kDa in the hemocytes of P. camtschaticus, 30 and 82 kDa in the hemocytes of E. sinensis, and 43 or 70 kDa in the hemocytes of C. magister. More lanes reacted between hemocytes of E. sinensis and C. magister and their antisera. The results indicate antigenic similarities among the hemocytes of the four crustacean species.
Journal of Fish Biology, 1990
The ontogenetic development of IgM-containing cells is described as demonstrated by immunoperoxidase staining with a mouse anti-trout IgM monoclonal antibody and the differentiation of enzyme-histochemical markers in the non-lymphoid cells forming the stroma of the thymus, spleen and kidney of the rainbow trout. The first lymphoid cells staining with the monoclonal antibody occurred at day4-5 after hatching in the renal lympho-haemopoietic tissue. By I month after hatching IgM-positive cells also appeared in the spleen and thymus. Enzyme-histochemical demonstration of the alkaline and acid phosphatase and non-specific a-naphthyl acetate esterase enzymatic activities in the non-lymphoid cells indicated that a certain degree of maturation of the cellular stroma of the developing lymphoid organs of trout was reached before or at the time when IgM-expressing cells could be observed. The relationships of the stromal components of the various lymphoid organs to the development of IgM-positive cells, and the possible role of the renal lympho-haemopoietic tissue as a primary lymphoid organ for B-cell differentiation in the trout are discussed.