The ability to respond to specific antigens develops in a programmed fashion. Although the antibody repertoire in adults is presumably generated by stochastic combinatorial joining of rearranged heavy variable, diversity, andjoining (VH-DH-JH) and light (VL-JL) chains, experimental evidence in the mouse has shown nonrandom utilization of variable gene segments during ontogeny and in response to specific antigens. In this study, we have performed sequence analysis of 104-day human fetal liver-derived, randomly isolated constant region C, transcripts and demonstrate a consistent preference during fetal life for a small subset of three highly conserved VH3 family gene segments. In addition, the data show that this preferential gene segment utilization extends to the DHQ52 and the JH3 and JH4 loci. Sequence analysis of two "sterile" DH-JH transcripts suggests that transcriptional activation of the JH-proximal DHQ52 element may precede initiation of DH-JH rearrangement and influence fetal Do utilization. Sequence comparisons reveal striking nucleotide polymorphism in allelic gene segments which is poorly reflected in the peptide sequence, implying considerable evolutionary selection pressure. Although vertebrate species utilize a variety of strategies to generate their antibody repertoire, preferential utilization of VH3 elements is consistently found during early development. These data support the hypothesis that VH3 gene segments play an essential role in the development of the immune response.Immunoglobulins are generated by combinatorial joining of rearranged gene segments of the heavy chain variable, diversity, and joining regions (VH, DH, and JH) and light chain regions (VL and JL) (1). Starting with less than 1000 of these germ-line elements, more than 109 different antigen binding sites can be generated even in the absence of eitherjunctional diversity or somatic mutation. In mice, only a small fraction of this potential repertoire seems to be expressed as functional antibody (2). Utilization of this repertoire appears to be developmentally regulated in a strain-specific fashion (3-5) with subsequent modification by environmental stimuli (7).The human neonate is relatively immunodeficient at birth (8). Controlled mobilization of germ-line variable gene segments has been postulated to underlie, in part, the maturation of humoral immunity (5, 9). With these observations in mind, we have concentrated on the use of molecular cloning strategies to dissect the development of the human heavy chain repertoire during fetal life. Because the extent of VH region polymorphism in the outbred human population is undefined, we have chosen to analyze individual fetal samples.Fetal B lymphopoiesis begins in the liver, with pre-B cells first detectable by 8 weeks of gestation (10). We isolated fetal liver mononuclear cells, which are rich in B-cell precursors, and generated oligo(dT)-primed cDNA libraries. We previously reported evidence of preferential usage of VH, DH, and JH gene segments by cloning and sequencing 1...