CHARACTERIZATION OF THE 2,4-DINITROPHENYL SPECIFIC B-CELL REPERTOIRE USING NEONATAL HYBRIDOMAS

The neonatal immunoglobulin repertoire has been studied to help clarify the processes by which an individual acquires the large array of antibody specificities present in the adult. Previous investigations have noted the restricted antibody populations shared by genetically identical neonates as well as the reproducible order of appearance of responsiveness to defined antigens in these animals. However, technical limitations have prevented further characterization of these early antibody specificities and their representation in the adult repertoire. Therefore, hybridoma technology was adapted to derive cultured cell lines from neonatal BALB/c murine B-lymphocytes secreting anti-2,4-dinitrophenyl (DNP) antibodies. These hybridoma antibodies were characterized by isotype, isoelectric point and intrinsic affinity and used to derive anti-idiotypic reagents to examine the anti-DNP response at both the B-lymphocyte and serum levels in various ages and strains of mice. Three of the idiotypes studied showed strict temporal patterns of expression among the DNP-specific B-lymphocytes in BALB/c neonates, each characteristically expanding to a maximum frequency at a certain day after birth and subsequently declining. Thus the development of the B-lymphocyte repertoire appears to be extremely reproducible not only in the sequence of appearance of antigen reactivities and major idiotypes, but also in the sequence of appearance of idiotypes within a response which comprise only a small proportion of the adult repertoire. This strict temporal patterning was observed in idiotypes expressed early when few antibody specificities were present and also in an idiotype undergoing maturation and expression during the second week of life when the majority of DNP-specific B-lymphocytes were already mature. All idiotypes studied were expressed repeatedly in DNP-specific B-lymphocyte and serum antibody populations of BALB/c mice. However, murine strains differing in immunoglobulin allotype did not produce these idiotypes in an anti-DNP serum response although similar idiotypes were observed in low frequency at the B-lymphocyte level among neonates of these strains. These results are compatible with a reproducible generation of antibody specificities which predicts a large overlap of both the antibody clonotypes present and their temporal expression among genetically identical individuals