Folding of an antibody variable domain in two functional conformations in vitro: calorimetric and spectroscopic study of the anti-ferritin antibody VL domain.

Understanding refolding pathways of recombinant antibody fragments is essential for efficient production of these proteins of high biomedical significance. The recombinant VL domain of mouse anti-human ferritin antibody F11 formed two distinct functional conformations obtained by refolding from bacterial inclusion bodies using two different procedures. Involvement of a dialysis step at pH 2-3 resulted in the VL-1 conformation with fluorescence of the highly conserved Trp-35 residue quenched by the spatially proximal disulfide bond.

Significance of serum immunoglobulin G for leukocytosis and prognosis in childhood B-lineage acute lymphoblastic leukemia.

Background: This study was conducted to evaluate the significance of serum level of immunoglobulins (Igs) and particularly IgG for leukemic cell persistence in peripheral blood (PB) and prognosis for childhood acute lymphoblastic leukemia (ALL).

Procedure: Human sera were obtained from 68 children with primary B-lineage ALL at diagnosis and 46 healthy children (control). Serum level of IgM, IgG, IgA, IgG1, IgG2, IgG3, IgG4, antitumor antibody, homogeneous IgG were quantified by turbidimetric or enzyme-linked immunosorbent assays.

Fusion of the antiferritin antibody VL domain to barnase results in enhanced solubility and altered pH stability.

Chimeric immunotoxins that combine antigen recognition domains of antibodies and cytotoxic RNases have attracted much attention in recent years as potential targeted agents for cancer immunotherapy. In an attempt to obtain a structurally minimized immunofusion for folding/stability studies, we constructed the chimeric protein VL-barnase. The chimera comprises a small cytotoxic enzyme barnase, ribonuclease from Bacillus amyloliquefaciens, fused to the C-terminus of the light chain variable domain (VL) of the anti-human ferritin monoclonal antibody F11.

Partially structured state of the functional VH domain of the mouse anti-ferritin antibody F11.

An antibody combining site generally involves the two variable domains, VH from the heavy and VL from the light chain. We expressed the individual VH domain of the mouse anti-human ferritin monoclonal antibody F11. The loss of affinity was not dramatic (K(a)=4.