A sensitive bioassay for the mycotoxin aflatoxin B1, which also responds to the mycotoxins aflatoxin G1 and T-2 toxin, using engineered baker's yeast

A novel aflatoxin B(1) bioassay was created by introducing a Lipomyces kononenkoae alpha-amylase gene into a strain of S. cerevisiae capable of expressing the human cytochrome P450 3A4 (CYP3A4), and the cognate human CYP450 reductase. This strain and a dextranase-expressing strain were used in the development of a microtitre plate mycotoxin bioassay, which employed methanol as the solvent and polymyxin B nonapeptide as a permeation enhancer. Stable co-expression of the CYP3A4 gene system and of the dextranase and amylase genes in the two bioassay strains was demonstrated. The bioassay signalled toxicity as inhibition of secreted carbohydrase activity, using sensitive fluorimetric assays. The amylase-expressing strain could detect aflatoxin B(1) at 2 ng/ml, and was more sensitive than the dextranase-expressing strain. Aflatoxin G(1) could be detected at 2 microg/ml, and the trichothecene mycotoxin T-2 toxin was detectable at 100 ng/ml.

Elemental analysis of soil samples for forensic purposes by inductively coupled plasma spectrometry - precision considerations

Major and trace elemental composition provides a powerful basis for forensic comparison of soils, sediments and rocks. However, it is important that the potential 'errors' associated with the procedures are fully understood and quantified, and that standard protocols are applied for sample preparation and analysis. This paper describes such a standard procedure and reports results both for instrumental measurement precision (repeatability) and overall 'method' precision (reproducibility). Results obtained both for certified reference materials and example soils show that the instrumental measurement precision (defined by the coefficient of variation, CV) for most elements is better than 2-3%. When different solutions were prepared from the same sample powder, and from different sub-sample powders prepared from the same parent sample, the CV increased to c. 5-6% for many elements. The largest variation was found in results for certified reference materials generated from 23 instrument runs over an 18 month period (mean CV=c. 11%). Some elements were more variable than others. W was found to be the most variable and the elements V, Cr, Co, Cu, Ni and Pb also showed higher than average variability. SiO2, CaO, Al2O3 and Fe2O3, Rb, Sr, La, Ce, Nd and Sm generally showed lower than average variability, and therefore provided the most reliable basis for inter-sample comparison. It is recommended that, whenever possible, samples relating to the same investigation should be analysed in the same sample run, or at least sequential runs.