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Multi residue EIA

Development of a multi-antibiotic EIA for the detection of virginiamycin, spiramycin, tylosin, bacitracin, and olaquindox.

After preparation of four separate prototype EIAs for the detection of virginiamycin, bacitracin, spiramycin/tylosin and olaquindox, a single multi-antibiotic prototype EIA was constructed.

Multi residue kit for the detection of 5 antibiotics.
Introduction:
Strips of a microtiter plate are coated with 4 different antibodies directed against 5 different antibiotics (virginiamycin, spiramycin/tylosin, bacitracin and olaquindox. For each antibiotic to be tested a zero standard (B max) and a cut-off standard has to be used in duplicate. Sample, standard solution and conjugate are added to the wells of the microtiter plate and incubated overnight at 4C. The conjugate contains the immunogens (virginiamycin, spiramycin/tylosin, bacitracin and olaquindox) individually conjugated to HRP. After incubation, the plate is washed followed by a substrate incubation of 30 minutes. The reaction is stopped by the addition of sulphuric acid. The colour intensity is measured in a spectrophotometer at 450 nm.

Interpretation of the results:
Results are calculated using the following equation:
Result = (Neg - Abs)/(Neg - Pos) x 100% (threshold)

Neg = absorbance of standard 0 (negative control)
Pos = absorbance of cut-off standard (positive control)
Abs = absorbance of sample

Sample is positive at a threshold > 80%
Sample is negative at a threshold < 80%

Ring test.
The ring test performed by 15 laboratories as part of the RADIUS project has shown the developed prototype ELISA test to be a very reliable guide for detecting the presence of a range of antibiotics present in animal feeds.

Reported by

EURO-DIAGNOSTICA BV
Beijerinckweg 18
6827 BN Arnhem
Netherlands
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