Graduate Thesis Or Dissertation
 

B-cell heterogeneity in salmonids

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  • Salmonid B lymphocytes were stimulated both in vivo and in vitro and shown to produce antibody specific for the immunogen. A passive hemolytic plaque assay was employed to demonstrate B cells producing specific antibody. Characterization of plaque forming cells (PFCs) involved sequential inhibition with a series of graded concentrations of free antigen. Splenic and posterior kidney derived PFCs exhibited greater heterogeneity in the population of responding B cells compared to anterior kidney derived PFCs. Anterior kidney PFCs were restricted in heterogeneity and lacked high affinity antibody, however, the splenic and posterior kidney PFCs possessed high affinity antibody producing B cells. Thus, the anterior kidney possessed immature B cell populations, characteristic of a lymphopoietic organ. Splenic and posterior kidney B cell populations were demonstrated to be functionally mature. The in vitro antibody production system facilitated investigations into B cell activation signals. T cell independent (TI) B cell activation by trinitrophenylated lipopolysaccharide (TNPLPS) was shown to occur via anti-TNP specific B cell receptors. The ability of salmonid B cells to discern different molecular forms of antigen (protein and lipopolysaccharide) was investigated indicating that the same subpopulation of B cells responds to both forms of antigen. Low dose gamma irradiation effects on B lymphocyte responses illustrated differential sensitivity of responses to this cellular insult, indicating the presence of a radiosensitive regulatory cell. Fine specificity analyses assessed the ability of in vitro generated antibody to recognize analogs of TNP. The effect of different molecular substituents on antibody recognition of the hapten analog indicated that a carboxyl group present on the substituent molecule proximal to the hapten analog hindered the antibody's ability to recognize the hapten. The simplest analog which revealed the heterogeneity of the antibody population was the dinitrophenol analog bearing two nitrite groups.
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