Phage display‐based molecular methods in immunohematology

Abstract

T he goal of the current Food and Drug Administration-sponsored workshop was to explore the feasibility of utilizing newly developed molecular approaches in the area of immunohematology. As stated by the workshop's organizers, conventional methods for blood grouping and associated compatibility testing have served us well for many decades, yet blood incompatibility remains a significant problem in transfusion medicine. This is due, in part, to the variability in the performance characteristics of preparations of polyclonal or monoclonal antibody reagents as well as to certain inherent limitations in a detection scheme that is agglutination-based. Furthermore, in comparison with diagnostic testing performed in other areas of laboratory medicine, current pretransfusion-testing methods are extraordinarily labor-intensive, an order of magnitude more expensive, subjective with respect to the interpretation of assay results, and tend to defy the development of high-throughput automation.Another drawback, as expressed by the Food and Drug Administration, is that current blood bank testing is a "single 'analyte' per test kind of test," in other words, one antibody/agglutination result per tube. The ability to get more than one result per test, that is, "multiplexing" blood group antigen identifications, would not only save time and expense, but could conceivably lead to a paradigm shift in the practice of transfusion medicine. As is the current standard of practice, red cells (RBCs) are only matched for the A, B, and D antigens unless "unexpected" alloantibodies to other red cell antigens are identified before a transfusion. Cost-effective methods for providing extended phenotyping and the matching of the blood supply to patients beyond the abbreviated three-antigen set could have significant medical and fiscal benefits. For example, the benefits of prospective RBC phenotype matching to three additional antigens (E, C, K1) in certain defined transfusion-dependent patient populations have already been seen as measured by a decrease in alloimmunization rate and reduction in the occurrence of delayed hemolytic transfusion reactions. 1 By the additional matching to other clinically significant antigens and by extending the practice of prospective antigen matching to all patients in need of transfusion, not only would the incidence of delayed hemolytic transfusion reactions lower dramatically, 2 but also would the development of positive antibody screens markedly decrease. This would obviate the need for the majority of time-consuming and expensive antibody identification procedures which currently comprise more than 50 percent of pretransfusiontesting costs.One approach to addressing these issues is a rapidly developing technology collectively referred to as "genotyping" in which the apparent phenotype of an RBC is deduced through the analysis of the donor's (patient's) genomic DNA. As described in several of the accompanying reports in this issue of TRANSFUSION, the need for reagent-grade serological reagents is essentially...

Translational applications of antibody phage display

“…The insets in (b) show corresponding microplate agglutination patterns with the anti-Rh(D)/anti-B phage mixtures using anti-M13 phage antibody as "Coombs reagent" (adapted from Ref. [32]) …”

Red cell blood grouping in the molecular era

“…The future of blood grouping reagents could lie with recombinant DNA and phage display technology [28]. This topic, however, is outside the scope of this review.…”

Delivery of Drugs Bound to erythrocytes: New Avenues for an Old Intravascular Carrier

“…Antibody phage display has been the subject of a number of recent reviews [279–281] and several excellent laboratory manuals are available that provide step-by-step protocols [282–285]. Typically, M13 filamentous bacteriophage, virions approximately 7 nm in diameter and up to 1 μm in length, are employed.…”