Sample Processing - Waterborne Pathogens: Detection Methods and Applications - page 93

found that a glass channel flow system performed better than a disposable
plastic molded cartridge, which they speculate was due to rougher walls in
the cartridge offering sites where pathogens could become trapped. 72 The
authors report that their work is a step toward creating automated sample
processing for protozoa, reducing the number of steps and human interven-
tion required. See Chapter 10, Section 10.2.1, for further discussion of how
microfluidics is applied to sample processing.
4.4.5. Ultrafilter centrifugation
Ultrafilter centrifugation is an alternative approach for concentration of
microbes from water that requires no preconditioning of the sample. Ultra-
filter centrifugation is mainly used for secondary concentration of viruses,
but will also concentrate bacteria and parasites if present in the sample.
Spin filters are available with different pore sizes, for examples Centricon
Plus-70 (Millipore, 5-100 kDa) and Jumbsep™ Centrifugal Devices (Pall,
3-300 kDa). During centrifugation, water and molecules smaller than the
cut-off will pass through the filter, and viruses and remaining particles/
microorganisms/molecules are concentrated. Several parameters influence
the efficiency of the ultrafiltration including membrane modules and mate-
rial used, operational conditions, microbe types, as well as water source and
quality. Due to the small pore size, more turbid surface water samples may
take a long time to process, and filters are likely to clog during concentrat-
ing waters rich in solid matter. Therefore, such samples are often subjected
to a centrifugation step prior to loading of the supernatant onto the filter.
The filters mentioned above have loading capacities of 70 and 60mL,
respectively and concentrate down to 0.35 mL and 3.5-4 mL according to
the manufacturers.
4.4.6. PEG precipitation
As mentioned previously, PEG is commonly used for the precipitation of
viruses. PEG is a chemically inert, water soluble, synthetic polymer. PEG
reduces the solubility of protein compounds by excluding proteins from the
solvent, effectively increasing their concentration until solubility is exceeded
and protein precipitation or crystallization occurs. 73 More highly charged
or hydrophobic proteins should be more easily precipitated than those of
lesser charge. 74 As enteric viruses are charged due to their surface proteins,
their precipitation will be favored although not exclusively in this manner.
PEG is nontoxic, but the disadvantage of using PEG upstream of molecular
detection is the risk to co-precipitate substances that can act as inhibitors.