In 1957, Erlanger1 first achieved a
successful preparation of steroid antigens. Until 1970, methods to
obtain haptens were virtually unchanged; namely, a functional group,
hydroxyl or ketone of the steroid hormone was converted to the
hemisuccinate or carboxymethyoxime.
In a second step, the hapten was
coupled to an immunologic carrier generally bovine serum albumin (BSA).
However, when the antibodies to these antigens are specific to the
hapten part in the particular steroid, the resulting antibodies lose
specificity at the functional group and its immediate vicinity.
Generally the antigens available today do not have free functional
groups.
In 1970, Midgley and Niswender2
reported that the antibodies prepared from antigens in which
progesterone was linked to the antigenic carrier through C-11 had a
greater specificity than those obtained from antigens in which
progesterone was linked through C-3 or C-20. For instance, progesterone
conjugated through position 20 to BSA generates antibodies that react
almost equally with progesterone its 20-dihydro and 17-hydroxy
derivatives, testosterone and even 11-dexoy
corticosterone2,3. These authors attributed the increased
specificity of this conjugation of the steroid antigen to the fact that
both ends of the steroid retained their pre-existing functions.
At the same time, Lindner, Perel
and Friedlander4 obtained an estradiol antigen linked at C-6.
Numerous studies have since then confirmed the Midgley hypothesis with
steroid linked at C-6, C-7, C-15 and C-1. The resulting antibodies had
greater specificity, with minimal cross-reactions. It is likely that
functional groups of the steroid which remain free act as antigenic
determinants.
Our antigens are synthesized in
this perspective and antibodies elicited have good specificity. Midgley
and Niswender5 observed significant antibody titre in rabbits
immunized with bovine serum albumin conjugates containing 20 or more
steroid molecules per molecule of protein. When the ratio was less than
10 the antibody was low6. The antigens below have 20 or more
molecules incorporated per mole of BSA as measured according to
Tamaoki7.
1. Erlanger, B.F., et al., Biol. Chem.,
228, 713 (1957)
