(2) Ka = 1
(Ab)
In other words. The concentration of free antibody that must be
maintained in order to bind 50% of the hapten is inversely
proportional to K . In a competitive radioimmunoassay, it is not
a
always necessary to achieve 50% binding of marker (although this
amount is often what is used), but binding cannot be much less than
25% or the level of bound radioactivity will be too small. As an
initial approximation, then, the K of the antibody determines the
a
maximal dilution of antiserum than can be used to obtain adequate
binding of the marker.
Thus, the practical sensitivity relates to a rule of thumb for determining maximal dilution which will
give 50% antibody binding and insuring that there will be sufficient marker to bind to the antigen for
detection.
Langone did not use Parker’s equation to determine the practical sensitivity. Rather he used
the equation in reverse to determine the affinity constant. From the data in Engvall’s example 1,
Langone selected a concentration of 12.5 µg/l and calculated a K of 5.2 x 10 liters per mole. ER9
a
3515-19, E131. However, it is not apparent that a person having ordinary skill in the art would make
the same selection in estimating the affinity constant. As indicated by the Parker text, the equation
applies to the situation in which 50% of the antigen is bound to the antibody. E130, p.111. Langone
did not establish or explain how he, or a person of ordinary skill in the art, would determine from the
data in Engvall’s specification that the value of 12.5 µg/ml in example 1 corresponds to 50% binding.
Without this information, one having ordinary skill in the art would not have any basis to select 12.5
µg/ml or any of the other values in example 1, to give a reasonable estimate of the affinity constant.
In our view, the person of ordinary skill in the art would not necessarily arrive at a value of
5.2x10 liters/mole using Langone’s technique. Thus, there is insufficient basis to find that the person9
of ordinary skill in the art would reach a reasonably similar determination of the affinity constant. In
other words, Langone’s determination is speculative and not the necessary and only reasonable
construction to be given to the data in Engvall’s example 1. Kennecott, 835 F.2d at 1423, 5 USPQ2d
at 1198.
34
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