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A ring test was organized for the detection of animal proteins in animal feed by microscopy in the framework of the annual ring tests of the IAG - International Association for Feeding stuff Analysis, Section Feeding stuff Microscopy. The organizer of the ring test was RIKILT - Wageningen UR, The Netherlands. The aim of the ring study was to provide the participants information on the performance of the local implementation of the detection method for their local quality systems. A further aim was to gather information about the application of the microscopic method. All four samples used in the ring test were based on an artificial feed with a formulation comparable to that of an average cattle feed. A mix of minerals was included at a level of 1%. The contaminations were: no animal proteins (blank), 2% of fish meal, 0.05% of land animal material, and 0.1% of tricalcium phosphate (TCP). Most of the specificity and sensitivity scores were at good levels. The specificity score for incorrect detection of meat and bone meal (MBM) in the blank is acceptable (0.94). The detection of 0.05% of MBM in feed appeared to be perfect (1.0). The TCP, which is a legal ingredient for non-ruminant feeds, was detected in a vast majority of cases as animal proteins in the sense of the legislation. Four participants reported fish in the presence of TCP. For the first time in the ring test of IAG section Microscopy participants were requested to report on their examination of either the flotate or the raw material of the sample. The results show a rather diverse view. The results for the blank were at a reasonable level (0.91). The detection of animal material (0.05%) in either flotate or raw material needs further improvement (0.34). According to the new procedure the results for examination of sediment and of flotate/raw material need to be combined in one result. Therefore, the sensitivity for the examination of flotate or raw material does not have a direct effect on the performance of the method. The share of the sediment used for examination differed between 2% and 100%. This difference showed to have an effect on the sensitivity of the method.
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IAG ring test animal proteins 2013
L. van Raamsdonk, V. Pinckaers, J. Vliege
Goals
● Quality support of participants
● Method performance
Ring test animal proteins
Ring test animal proteins
Procedure
● Preparation of samples; send 5 March
● Examination; return 12 April
● Preparation of report; draft ready 6 June
Ring test animal proteins
Materials, based on an artificial feed
● Blank sample
● Sample with fish meal 2%
● Sample with 0.05% ruminant MBM (Uruguay)
● Sample with 0.1% of tricalcium phosphate (TCP)
Ring test animal proteins
Microscopic method
● New amended version of R (EC) 152/2009
● Effective from 12 February, but no official SOPs at the time of the ring test
● New elements in procedure
● Implemented in ring test: examination of flotate or raw material
● Not implemented: repetition of examination
● Not implemented: official report texts
Ring test animal proteins
microscopy 53 participants; 53 sets
PCR 7 participants; 10 targets
Immunochemistry none
Response
Ring test animal proteins
Results
● Background of materials
● Accuracy fish
● Accuracy terrestrial animals
● Accuracy flotate or raw material
● Quantification
● Application of method parameters
● Results of PCR
● Evaluation
Ring test animal proteins
0.96
0.98
0.92
A: blank
B: 2 % fish
C: 0.05 % MBM
D: 0.1 % TCP
specificity sensitivity
0.98
Results fish
Ring test animal proteins
0.94
0.98
specificity sensitivity
1.0
0.92 *
Results terrestrial animals
A: blank
B: 2 % fish
C: 0.05 % MBM
D: 0.1 % TCP
*: EU member states only: 0.98
Ring test animal proteins
TCP identification
Ring test animal proteins
0.91
specificity sensitivity
0.83
0.34
0.68
Results flotate or raw sample material (target: animal material in general)
A: blank
B: 2 % fish
C: 0.05 % MBM
D: 0.1 % TCP
Ring test animal proteins
Method application
● Tendency to make smaller slides (small cover glass: n=38)
● Increasing use of special glass with cock
● Decreasing use of staining of sediment
● Unchanged frequency of using a binocular
● Amount of sediment used: 1 – 100 %
Ring test animal proteins
10 gram of material
200 milligram sediment
20 mg
Examination:
20 mg
1
gr
Example: 0.05% 20 particles
20 p
Result: positive
2 p
Result: suspect
Amount of sediment used
Ring test animal proteins
1.0
1.0
0.81
< 50% >= 50%
n=14 n=24
1.0
1.0
1.0
A: blank
B: 2 % fish
C: 0.05 % MBM
D: 0.1 % TCP *
*: p = 0.053
Amount of sediment used: sensitivity
Ring test animal proteins
Recommendations
● Improvement of examination of flotate or raw
material
● Method: full application
● Harmonisation of the method
● Further attention to detection of TCP
Ring test animal proteins
Thanks to:
● All participants of the study
● IAG board for cooperation
Thank you for
your attention
Questions?
