2. Materials and methods
2.1. Study design
To determine the suitability of PAG-ELISA for the
detection of EM/FM, the following three experiments were
conducted.
1. A validation of the PAG-ELISA to verify that a decline in
the PAG concentration can be identified by measuring
the optical density (OD) using a semiquantitative PAGELISA.
2. A retrospective verification experiment to investigate a
previously described cut-off for the prediction/diagnosis
of EM/FM (PAG-OD¼0.4–1.5; (Meyerholz et al. 2013)).
Receiver-operating-characteristic (ROC) curve analysis
was conducted to calculate the following two cut-off
values: (1) high sensitivity (SEN) for prediction
(SEN490% and specificity [SPE]Z50%) and (2) high SPE
for the diagnosis of EM/FM (SPE490% and SENZ50%).
3. A field experiment to compare TRU results with the
PAG-ELISA results for the prediction/diagnosis of EM/
FM. Moreover, a combination of PAG-ELISA and TRU for
the prediction/diagnosis of EM/FM was tested.