A.C. electrokinetic bioassays : development of electrorotation assay for analytes in water
The work described is primarily concerned with the understanding of the induced AC electrokinetic properties of the transmissive stages of two genera of waterborne protozoan pathogens, namely Cryptosporidium and Giardia. The assessment of viability through the use of electrorotation (ROT) has been investigated in comparison with conventional techniques. The optimum conditions for detennining C. parvum viability using ROT are described and discussed. Two Giardia and three Cryptosporidium species were investigated, as were a total of ten Cryptosporidium parvum isolates. With all species investigated a good correlation was found between the ROT response of clean particles and conventional vital dye techniques. Adherent bacteria on the particles surface have been identified as a major problem for subsequent ROT analysis. This is the first description of the effect of adherent bacteria on the dielectric response of biological particles . . A novel single layer electrode array was designed and successfully tested to overcome the problems of low particle concentration, interfering debris and particle position in the electrorotational electrodes. The device was shown to selectively concentrate particles into a central region whereupon their physiological state was then assessed through their ROT response. Modification of the device enabled the isolation of one or more particles onto a membrane, providing a suitable collection method for low particle number handling. The clearest demonstration to date of the effect of membrane integrity on the ROT response is described from spectra obtained for an oocyst before and after excystation in vitro. Storage time effects for oocysts of C. baileyi are also described as are isolate differences for C. parvum oocysts which are apparent in the low frequency region of the spectra. The implications of these results to the water industry and potential diagnostic applications of electrorotation are discussed.