Binding properties of a polyclonal antibody directed towards lead complexes

A previously described conjugate of 8-hydroxyquinoline and bovine serum albumin was complexed with lead(II), (8-hydroxyquinoline to metal ion ratio 2:1) and used as an immunogen to produce polyclonal antibodies against lead in chickens. Antibodies obtained from a first blood sample during a standard immunisation procedure showed very promising features (dynamic range of the assay was 1 to 1000 ng l(-1)). Nevertheless, proceeding with the immunisation caused a complete loss of the recognition of the complex. A modified brief immunisation procedure was carried out and, in this case, the immunogen proved to be sufficiently stable in vivo to produce antibodies that selectively bound to the lead(II) complex (in the same 2:1 ratio used as an immunogen). Since the antiserum obtained cannot reach the same performance levels as the first one, standard curves were obtained by adding the free 8-hydroxyquinoline to the solution, which enables 2:1 complexes to be more easily formed. Cross-reactivity and dependence from buffer were investigated, showing at least 10-fold lower binding to non-target divalent metal ions compared to lead(II). MES buffer (pH = 6.0) gave more sensitive but very imprecise curves, whereas Tris (pH = 8.5) allows higher precision but lower sensitivity to be observed.