Date of Award
Open Access Thesis
Medical Doctor (MD)
Dr. John Dwyer
The clinical potential of specific passive mucosal immunity is discussed and it is proposed that bulk quantities of hybridoma produced specific dimeric IgAs could be clinically useful as made to order super colostrums. SIgAs function on mucosal surfaces to prevent the adherence of pathogens. They are poor opsonins and do not activate complement except by the alternate pathway. In principle mesenteric lymph note (MLN) or Peyer's patch (PP) lymphocytes aseptically harvested from enterically immunized and boostered mice displaying blastogenic anamnestic response might be fused with log phase cells of a non secreting murine myeloma to generate specific dimeric IgA (+J) secreting hybridomas. Lipoteichoic acid (LTA) has been demonstrated to be the adhesin of the group A streptococci and possibly many other gram postive mucosal pathogens. The polyglycerolphosphate backbone common to all LTAs is a 25-30 residue polymer affording many identical binding sites to a single specificity (monoclonal) antibody. Preincubation of streptococci with backbone specific antibody has been shown to block adherence to human buccal epithelial cells in vitro. Murine hybridoma derived dimeric IgA anti LTA backbone will likely have considerable ability to block adherence of streptococcal pathogens in vitro. In vivo it may be useful as a safe clinical agent providing passive mucosal immunity against a wide variety of gram positive pathogens, and might indeed be employed as a super colostrum. A step toward this goal was taken by demonstrating IgA secreting murine hybridomas can be produced using gut associated lymphoid tissue (GALT) from enterically immunized and boostered animals. MLN and PP lymphocytes from two groups of 5 mice intragastrically immunized and boostered with Freund's complete adjuvant (FCA) were separately pooled and fused with SP2 cells. MLN cells were found to be poor fusion partners with a hybridization frequency 1/10 that of PP or spleen with SP2 cells, paralleling the 1:10 concentration ratio of surface immunoglobulin bearing blasts in these tissues. The 20 day post fusion supernatants of all 12 MLN hybrids and 25 of the 36 (71%) of PP origin failed to demonstrate antibody production of ELISA. The difference is significant at P=0.1, x2c(1df)=2.8, and it is conjectured these MLN blasts were at a stage of differentiation less adept than PP blasts at promoting antibody secretion in SP2 hybrids. The observation of 4 IgA, 2 IgG, and 4 IgM containing supernatants is consistent with the 54:16:30 proportion of PP blasts displaying these surface antibodies, x2c=.185. No attempt was made to determine antibody specificities as FCA was assumed to produce generalized GALT stimulation. Work is currently in progress to assay the IgA containing supernatants for polymeric antibody.
Staszewski, Robert, "Hybridomas Derived from Murine Gut Associated Lymphoid Tissue" (1984). Yale Medicine Thesis Digital Library. 525.