The whole electrophoretic procedure, including polymerization of the gels, was carried out at 15 C in a cold room.
Fraction Collection and Analysis
For elution, a peristaltic pump was set to a flow rate of 1ml/min. The absorption profiles of the eluates were continuously monitored at 280 nm. A total of 100 fractions were collected at 3 min intervals. The time for the complete isotachophoretic procedure was approximately 24 h.
For platinum detection, fractions were completely mineralized by an open, wet digestion step with nitric acid, perchloric acid, and sulfuric acid. Platinum was determined by our sensitive absorptive voltammetric method that has detection limits in the sub-pg range.5
These investigations were performed with the aim of developing analytical methods for the separation and detection of platinum species in plant materials. It was shown that by an approach combining gel chromatography and isotachophoresis, platinum-containing species in grass samples could be separated with high resolution.4,5 Figure 1 shows the UV profile and platinum distribution of the isotachophoresis step. Very sharp and concentrated platinum peaks were obtained. The platinum of the corresponding peaks eluated in only one or two fractions as a result of isotachophoresis.
The molecular mass of these platinum-containing fractions were
determined by native horizontal flat-bed electrophoresis (PAGE).
By using our sensitive determination method it was possible to
detect platinum in separated and stained protein bands after
PAGE, followed by digestion of the excised gel segments.5