Melvyn F. Greaves

A method is described by which an immunoaffinity matrix was constructed by binding antibody directly or indirectly to protein A-Sepharose 4B followed by cross-linking of the complex with dimethyl pimelimidate. This allows optimal spatial orientation of antibodies and, thus, maximum antigen binding efficiency. The affinity matrices were stable to high and low pH buffers without any significant antibody loss. The optimal conditions of antibody saturation, cross-linker concentration, and elution system were established and affinity columns made with the monoclonal antibodies J5, W6/32, and OKT9 for one-step isolation of the common acute lymphoblastic leukemia-associated antigen, HLA-AB antigens, and transferrin receptor, respectively, from cell lysates. The same methodology was also applied to immobilize transferrin with polyvalent anti-transferrin antibodies. This was then used to isolate the transferrin receptor from cell lysates.

A murine monoclonal antibody (OKT9) raised against human leukemic cells binds to a wide variety of leukemia and tumor cell lines and to a minority of leukemia cells taken directly from patients. Fetal thymus and liver are strongly reactive as are some normal, immature hemopoietic cells and activated lymphocytes. Reactivity with OKT9 appears to correlate with proliferation status in both normal and malignant populations. Biochemical analysis indicates that this structure is a approximately equal to 180,000-dalton glycoprotein with two disulfide-bonded subunits of approximately equal to 90,000-daltons. Isolation of the transferrin receptor from a T-cell line (MOLT-4) indicates that it also has a dimeric approximately equal to 180,000-dalton structure. Radio-labeled transferrin bound to its receptors can be specifically precipitated by the monoclonal OKT9, although the latter does not bind transferrin itself, indicating that the antigenic structure defined by this antibody is likely to be the transferrin receptor.

An increasing number of reports document instances in which individual leukemic cells coexpress markers normally believed to be restricted to a single lineage. This has been interpreted by McCulloch and colleagues as aberrant programming or lineage infidelity and contrasts with earlier suggestions that lineage fidelity of gene expression was usually maintained in leukemia. We argue that several examples of infidelity are suspect on technical grounds, whereas others are bona fide and require explanation, eg, partial rearrangements and expression of Ig heavy-chain and/or T cell receptor genes in inappropriate cells and terminal deoxynucleotidyl transferase in leukemic myeloblasts. Individual examples of truly aberrant gene expression may well occur in leukemia but with insufficient regularity to be of general significance. We suggest that verifiable and consistent examples of apparent lineage infidelity do not reflect genetic misprogramming but rather the existence of a transient phase of limited promiscuity of gene expression occurring in normal biopotential or multipotential progenitors and able to be preserved as a relic in leukemic blast cell populations that are in maturation arrest. This alternative explanation has interesting implications for mechanisms of hematopoietic differentiation and leads to some testable predictions.