Radioisotope Dilution Methods
An important group of analytical methods is based on measurements of the change in isotopic ratio
when active and non-active isotopes are mixed. In the simplest case, a known amount w 1 of labelled
analyte of known specific activity a 1 is added to the sample. After isotopic mixing has been established
sufficient of the analyte is separated (nor normally 100%) to allow the new specific activity a 2 to be
measured. Measurements of activity and the amount of the analyte separated are thus required.
Subsequently the amount w 2 of analyte in the sample may be calculated from equation (10.17).
This method has been applied to analyses as different as the determination of vitamin B 12 in biological
tissue (using a^60 Co label), and the amount of hydrogen absorbed into the lattice of a transition metal.
As the method does not require quantitative separations, and involves the acknowledged sensitivity
accompanying radioactive measurements it is attractive for trace element determination. Limitations are
however imposed by the problems of determining the chemical yield. A method known as
substoichiometric yield determination has been developed to overcome this. Typically, a known
(substoichiometric), amount of a complexing agent is used under conditions guaranteeing quantitative
reaction with the analyte. The amount of analyte separated is then known from the amount of
complexing agent used and does not have to be determined. For example, using^59 Fe as a tracer,
substoichiometric amounts of Cupferron, and chloroform extraction, iron may be determined at levels
of 10–^9 g cm–^3 with a relative precision of 2%. Reverse isotope dilution analysis employs dilution with a
non-active 'carrier' to determine the total amount of a labelled compound in a sample.
Radioimmunoassay
A most important technique which has been developed as an extension of the isotope dilution principle
is that of radioimmunoassay (RIA). Analyses by this method employ substoichiometric amounts of
specific binding immuno-chemical reagents for the determination of a wide range of materials
(immunogens) which can be made to produce immunological responses in animals such as sheep or
rabbits. It is possible to combine the specificity of an immunochemical reaction with the extreme
sensitivity of radiotracer detection. Analytical methods based upon these principles have achieved wide
applicability in the determination of organic compounds at trace levels.
In order to establish an RIA method, it is first necessary to obtain the appropriate specific binding
agent. This is done by using the analyte to trigger the natural defence mechanism of an animal in which
antibodies are produced to combine with, and to neutralize the foreign molecules (immunogens or
antigens) entering the bloodstream. These antibodies are