Phage-derived monoclonal anti-Lua

BACKGROUND: Monoclonal antibodies (MoAbs) are gradually replacing human polyclonal sera as typing reagents. Many blood group specificities, however, are not amenable to classic hybridoma technology. The phage display technology, aimed at isolating peptides or antibody fragments, offers an alternative strategy. Recombinant antibodies derived from this technology would greatly facilitate phenotyping and decrease analysis cost. STUDY DESIGN AND METHODS: A human single-chain Fv (scFv) phage-displayed library was panned on red blood cells (RBCs) in an attempt to isolate clones recognizing human RBC specificities. Three rounds of biopanning were performed. Enrichment was monitored by phage titration, and selected phage populations were analyzed further. RESULTS: Three major clones were identified by clone diversity analysis. One of them showed a specificity for Lu-a. This scFv was reconstructed into a human IgG(1) by recombinant DNA methods. The reactivity of the reconstructed human IgG(1) toward Lu-a is indistinguishable from its parent scFv. Moreover, the specificity of the antibody was confirmed by serologic assays, flow cytometry, and biochemical analysis with RBCs of different Lu phenotypes and a recombinant cell line expressing Lu glycoproteins. CONCLUSION: With phage display and standard recombinant DNA methods, isolation of a scFv of Lu-a specificity was successful, from which a complete human IgG(1) MoAb of equivalent reactivity was reconstructed. To our knowledge, this is the first MoAb specific for Lu-a.