Results and Discussion
ELISA Cross reactivity
The first step in carrying out this study was to interpret the diaminoatrazine ELISA cross
reactivity data provided by the manufacturers, to determine if compounds that may be in
Wisconsin's groundwater would also react in the assay.(Tables 1 and 2) Cross reactivity
is the ability of an ELISA to detect related compounds to varying degrees. It is a difficult
concept to quantify especially when more than one cross-reactant is present in a sample.
It should not be concluded from the cross-reactivity table that the cross reactants are
cumulative (for example, using the atrazine cross reactivity table (table 1), if there were
0. 1ppb atrazine and 0.1 ppb propazine present in a sample, the atrazine immunoassay
would not necessarily provide a result of 0.2ppb). From a public health standpoint cross-
reactivity can be a positive or a negative feature of the assay depending on the intended
use of the results.
Cross-reactivity to metabolites would be a positive attribute if screening samples for a
group or a certain class of compounds were the objective. Using the atrazine ELISA as an
example, if there is an ELISA result of <0.062 ppb, it can be concluded (from the cross
reactivity, Table 1) that atrazine, propazine, ametryn, prometryn, prometon, desethyl
atrazine, are all less than 0.062 ppb, as 0.062 ppb is the highest minimum detection level
(MDL) concentration of that group. That finding would rule out the occurrence (at levels
above 0.062 ppb) of several chemicals with one simple test. On the other hand, a positive
atrazine ELISA detection in a sample could be due to any one or a combination of the
cross-reactants and the results of the assay must be interpreted within the constraints of
that knowledge.
Both the diaminoatrazine ELISA, and the commercially available atrazine ELISA, have
some cross reactivity to other atrazine metabolites as well as to other triazine herbicides
and their metabolites. The manufacturers have tested cross reactivity to some compounds
for both test kits (Tables 1. and 2). Some of the samples used in this study had known
concentrations of deisopropylatrazine. Tables 1 and 2 do not indicate that
deisopropylatrazine is a significant cross reactant and therefore will not skew the results.
This lack of reactivity to deisopropylatrazine could hamper the effectiveness of either
assay in determining the safety of the rare drinking water sample that may have a high
concentration of deisopropylatrazine, but little or no significant concentrations of other
atrazine or atrazine metabolites.
Recovery of analyte would also be a factor affecting comparability of the methods.
Beacon Analytical Systems did not provide diaminoatrazine recovery data for the new
ELISA test. Subsequently, recovery data was acquired during this study by implementing
a 2.0 ppb diaminoatrazine spiked blank that was analyzed in five different analytical runs.
The average spike recovery over the five runs was 10000 (STD. DEV. =0.094074). This
recovery is very favorable when compared to the recoveries for the conventional DATCP
GC method, which varied between 50-100%o. (2)