Cloning of Size-Selected Human Immunoglobulin Heavy-Chain Rearrangements from Third Complementarity-Determining Region Fingerprint Profiles

摘要

Methods have been developed to rapidly visualize the size distribution of third complementarity-determining regions (CDR3) in immunoglobulin. (Ig) and T-cell receptor (TCR) molecules. DNA fragments spanning the Ig or TCR CDR3 are generated by PCR using primers at fixed positions in the variable and constant segments. These fragments differ in length due to size variation of the CDR3s Visualization of the amplification products in polyacrylamide gels as a "CDR3 fingerprint profile" is a rough measurefor the complexity of the Ig and TCR antigen-binding specificities. We report an adaptation of this method for the analysis of human Ig heavy-chain genes that incorporates silver staining, which allows for the fine analysis of specific regions of the profiles. This is especially useful for the study of low-abundant transcripts.