Aflatoxin test kits

Figure 3. Operating characteristic curves for USDA/FGIS collaborative study of aflatoxin test kits.

The time required for complete analysis using any of the above kits was about 30 minutes depending on the amount of time required for the filtration of the sample. In summary, any of the above immunoassay tests could be use for screening of animal feeds down to a level of 20 ppb. Since the above study, the USDA recently completed a comparative study on six commercially available aflatoxin test kits including the above four with... 

Table I. Commercially Available Immunoassay Test Kits for Aflatoxins...

A third collaborative study (10) was conducted examining the Immuno Dot Screen Cup (Afla-20, IDS Cup), a test kit containing antibodies with considerable cross reactivity with aflatoxins B2 and G,. The manufacturer s procedure was also modified to increase the reliability of detection at 20 ng/g total aflatoxins, and to broaden the applicability to include peanut butter samples. 

Zheng Z, Humprey CW, King RS, Richard JL (2005) Validation of an ELISA test kit for the detection of total aflatoxins in grain and grain products. Mycopathologia 159 255-263... 

This study focused on the applicabili of the commercially availaUe immunochemical test kits to adequately screen for aflatoxin in cottonseed undw non-laboratoiy conditions by comparing their performance to laboratoiy results. On a obal e, TLC and immunochemical methods umuld have highest potential use. TLC was used to conflrm the presence and identity of aflatoxins in the test samples. 

Aflatoxin Analysis. Immimoassays were performed at the various locations where samples were collected and repeated approximately 24 hours later in the laboratoiy (Food Toxicology Research Laboratory, Department of Nutrition and Food Science, University of Arizona). The four commercially available (US market) test kits, listed below, were used ... 

Ammoniated cottonseed samples were also analyzed in this study. Ammoniation is a process Aat has successfully reduced (>99%) aflatoxin levels in com, peanut and cottonseed products (1. Arizona uses ffiis process to treat aflamzin-contaminated, udiole cottonseed and cononsccd meal. This process had no apparent effect on the fiinction of any of die immunochemical test kits. 

Determination test kits are based on direct but more often on indirect-type ELISA techniques. They use 96-well microtitration plates or beads or sticks. Analysis for mycotoxins can be completed within 3 h. This kind of test needs a photometric reader for the quantitation by plotting sample value against standard-curve derived values. Nowadays, a number of companies are developing such tests for aflatoxins, OTA, patulin, zearalenone, T-2 toxin, fumonisins, DON, etc. with limits of quantification as low as 1 pgperkg or even less. 

A conventional microtiter plate, ELISA requires equilibrium of the antibody-antigen reaction that would require an incubation time of approximately 1-2 h. Currently, most of the commercially available ELISA test kits for aflatoxins are working in the kinetics phase of antibody-antigen binding, which reduces the incubation time to minutes. Although reduction of incubation time may lead to some loss of assay sensitivity, the test kit can provide accurate and reproducible results [75]. 

This classic biological methodology is widely used in research, diagnosis, and testing because it is affordable and sensitive to tiny amounts of material. It is applied to domains such as the environment and agriculture (pesticides, anabolic steroids, aflatoxins, HPA) for which numerous assay kits exist. However only compounds for which there exist adapted antibodies and the conjugated enzymes can be measured. 

The evaluation of a number of immunoassay diagnostic kits was undertaken to determine their usefulness in a regulatory analytical laboratory environment in the food, feed and pesticide areas. Four rapid enzyme immunoassay tests for the detection of aflatoxin residues at the 20 ppb level in animal feeds were compared to the official HPLC procedure. In the pesticide area, a commercial pentachlorophenol competitive inhibition assay for residues in water was investigated as to its applicability to poultry and pork liver matrices. In addition, an ELISA screening procedure for the herbicide fusilade was developed. Modifications were incorporated into the rapid immunoband 1-2 Test procedure for the detection of motile Salmonella in various food and animal feed products resulting in quicker analysis than the standard culture method. Also, a comparative evaluation of a Quik-Card Test for sulphamethazine drug residues in pork urine, liver and muscle tissue, is described. 

E-Z Screen. Manufactured with a proposed use for aflatoxins B1, B2, G1, G2 at 10 ppb, was found to be effective for B1 and G1 in spiked corn at the 10 ppb level. Sample preparation was as prescribed by the kit manufacturer except the sample was shaken for one hour, instead of blending for one minute. A 50 g sample was shaken for one hour with 100 ml 80% methanol and allowed to settle. Then a 100 uL of extract was diluted with 200 uL buffer and 50 uL of this mixture was applied to the sample port of the EZ card. Aflatoxin B2 was detected at the 15 ppb level and G2 at the 20 ppb level. This card test was less sensitive for mixed feeds which contain more pigments- Due to the lack of demand, the manufacturer has stopped producing cards sentitive at the 10 ppb level, but control cards at the 20 ppb are available. In one study 22 samples were tested along with standards and formulated product. No false negatives were encountered and only one false positive. No false positives were encountered on 40 samples tested in another laboratory. Recently cards sensitive to 5 ppb were introduced. Preliminary research indicates the cards will perform to specifications. As the presently used HPIC procedure processes 6 samples in a 2-day period and 99% of the samples were negative, the aflatoxin E-Z Screen could provide the necessary screening capability. 

Goto, T. Manabe, M. Application of ELISA to aflatoxin analysis Results of preliminary tests of analysis kits on the market Rep. Natl. Food Res. Inst, 52 53-9. 1988. Japanese. 

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