Food Additives and Contaminants 715 Figure 3. SPR optical biosensor inhibition assay format. RU 650 600 550 500 450 e sn 400 o sep 350 R 300 250 200 150 100 0 1 2 3 4 5 6 7 Concentration 8 9 10 11 ng/ml Figure 4. Typical response versus PSP toxin concentration calibration curve using a polyclonal antibody binder. Ruberu et al. 2003). This microtitre plate format has recently undergone successful single-laboratory valida- tion (van Dolah et al. 2009). The fact that all STX analogues bind to SCRs with affinities that vary according to their toxic potency (Doucette et al. 1997) indicates that receptor-based competitive bind- ing assays provide a measure of a sample’s toxicity, irrespective of which PSP toxin(s) is present. The saxiphilin receptor, occurring naturally in various amphibians and terrestrial invertebrates, has also been integrated into a competitive binding assay using a microtitre plate format (Llewellyn and Doyle 2001). Llewellyn (2006b) studied the behaviour of mixtures of PSP toxins in these two unrelated compet- itive binding assays and found that the most potent toxins affect the toxicity of the mixture to the greatest extent with the less active toxins present required to be several orders of magnitude greater for the mixture to reflect their potency. Receptor ligand binding assays using SPR technol- ogy had previously been investigated (Gestwicki et al. 2001; De Jong et al. 2005). As receptor binders for PSP toxins reflect toxicity in a similar manner to the mouse bioassay, it was envisaged that they could be an ideal candidate for the development of an SPR assay to replace the mouse bioassay. Campbell et al. (2007) examined crude extracts of SCR from rat brains using SPR analysis, but technical difficulties were encoun- tered with the SCR in that this preparation appeared to be stable for only a short time at 4 C, which in effect was detrimental to reliability and reproducibility. Another issue with developing SPR assays for small molecular weight toxins is that when the toxin is immobilised onto the surface, the toxin may then be orientated in such a manner that it no longer locks into the binding site of the receptor pocket, making the toxin unrecognisable with minimal to no binding occurring. As SPR is dependent on changes in mass, for the receptor to be immobilised on the toxin surface the toxin binding in isolation is not of a large enough mass to achieve a change in response and the signal would need to be amplified. Amplification could be performed in a manner similar to a sandwich assay by using a secondary antibody. For small molecular weight compounds this is difficult to achieve due to a