RIDASCREEN® Ei/Egg Protein Art. No. R6402 General Egg white is recognized as being more allergenic than egg yolk. The major allergenic proteins in egg white are Ovomucoid (11 %), Ovalbumin (54 %), Ovotransferrin (12 %) and Lysozyme (3.4 %). In many countries egg and egg ingredients are covered by allergen legislation and have to be declared on the food label.
Test Principle The basis of the sandwich ELISA is an antigen-antibody reaction. Detailed information is contained in the test kit insert.
Test Validation RIDASCREEN®FAST Ei/Egg Protein is a sandwich enzyme immunoassay for the quantitative analysis of hen’s egg protein in food like salad dressings, baking mixtures, ice cream and wine.
Sample preparation The sample extraction is carried out using the RIDASCREEN® Allergen Extraction Buffer as described in the test kit insert.
Calibration Curve A typical calibration curve for RIDASCREEN®FAST Ei/Egg Protein is shown in appendix 1. The result is expressed as mg/kg (ppm) whole egg powder. The assay is calibrated against NIST RM8445 whole egg powder. To create the calibration curve the cubic spline function is recommended.
Specificity
03/2012
The antibodies detect ovalbumin and ovomucoid of hen’s egg.
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Allergens
RIDASCREEN® Ei/Egg Protein Art. No. R6402 Sensitivity Limit of Detection (LOD) The Limit of Detection or the lowest detectable concentration of whole egg powder in the RIDASCREEN®FAST Ei/Egg Protein ELISA that can be distinguished from zero matrices was determined to be 0.101 mg/kg (ppm) whole egg powder (see table 1). This is equivalent to 0.049 mg/kg whole egg protein and 0.027 mg/kg egg white protein. Table 1 Determination of the Limit of Detection (LOD) Sample
Standard 1 (OD*)
Standard 2 (OD)
Mean of samples* (OD)
Standard deviation (SD)
Mean + 3x SD (OD)
LOD (mg/kg)
White wine
0.176
0.384
0.254
0.027
0.081
0.148
Salad dressing
0.171
0.314
0.170
0.028
0.084
0.116
Lemon ice cream
0.173
0.372
0.123
0.029
0.086
0.039
Mean value
0.101
* mean of 10 extractions, each measured in duplicates OD = optical density
Limit of Quantification (LOQ) The Limit of Quantification or the lowest concentration that can be determined in a sample with acceptable precision (repeatability) and accuracy under the stated conditions of the test was found to be 0.5 mg/kg (ppm) whole egg powder (dilution factor of 20 included).
03/2012
Precision (scatter of replicate readings around their mean value) To determine the reproducibility and repeatability of the RIDASCREEN®FAST Ei/Egg Protein ELISA intra-assay and inter-assay coefficients were calculated.
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Allergens
RIDASCREEN® Ei/Egg Protein Art. No. R6402 Variance of the ELISA Intra-Assay Variation (repeatability) Within run variation was calculated by measuring the calibrators in one assay run (n=6). The within assay variation over the calibration curve is 8.5 % (data not shown). The average intra-assay variation of samples has been found at 16.8 % (see table 2) with an average recovery of 90.4 % (see table 2). Table 2 Determination of the Intra-Assay Variation (repeatability) of spiked samples (n=6) Sample
Target concentration whole egg powder (mg/kg)
Concentration measured (mg/kg)
Standard deviation
CV* (%)
Recovery (%)
Salad dressing
0
< 0.5
Salad dressing
1.5
1.5
0.3
19.8
100.0
Salad dressing
13.5
10.9
1.4
13.7
80.7
16.8
90.4
0.0
Mean value *CV coefficient of variation
Inter-Assay Variation (reproducibility) Between run variation was determined by repeated measurements of the calibration curves from different test kits of one production batch performed by three technicians (n=3 runs). In each run duplicates of samples and calibrators have been measured. The mean between assay variation for the RIDASCREEN®FAST Ei/Egg Protein is 7.2 % over the calibration curve (data not shown). The inter-assay variance of one negative and two spiked food samples is 10 % (see table 3). Table 3 Determination of the Inter-Assay Variation (reproducibility) of spiked samples (n=3) Sample
Target concentration whole egg powder (mg/kg)
Concentration measured (mg/kg)
Salad dressing
0
< LOQ
Salad dressing
1.5
Salad dressing
13.5
Mean value
03/2012
* CV coefficient of variation
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Standard deviation
CV* (%)
Recovery (%)
2.5
0.2
6.1
168.9
13.2
1.8
13.9
97.9
10
133.4
Allergens
RIDASCREEN® Ei/Egg Protein Art. No. R6402 Samples from the market Different food samples were bought in a supermarket and analyzed (see table 4). In several cases the food label did not declare egg although the ELISA RIDASCREEN®FAST Ei/Egg Protein gave a clearly positive result. Even in some heat processed food egg traces could be determined. Table 4 Measurement of egg containing samples Sample
