Micropreparative isoelectric focusing protein separation in a suspended drop

IEF protein binary separations were performed in a 12-μL drop suspended between two palladium electrodes, using pH gradients created by electrolysis of simple buffers at low voltages (1.5-5 V). The dynamics of pH gradient formation and protein separation were investigated by computer simulation and experimentally via digital video microscope imaging in the presence and absence of pH indicator solution. Albumin, ferritin, myoglobin, and cytochrome c were used as model proteins. A drop containing 2.4 μg of each protein was applied, electrophoresed, and allowed to evaporate until it splits to produce two fractions that were recovered by rinsing the electrodes with a few microliters of buffer. Analysis by gel electrophoresis revealed that anode and cathode fractions were depleted from high pI and low pI proteins, respectively, whereas proteins with intermediate pI values were recovered in both fractions. Comparable data were obtained with diluted bovine serum that was fortified with myoglobin and cytochrome c.

[Rumenocentesis: a suitable technique for analysis of rumen juice pH in cattle?]

In the United States, rumenocentesis has been recommended especially for early diagnosis of subacute rumen acidosis (SARA). The objective of the current study was to evaluate health risks due to the technique ofrumenocentesis and to measure pH in ruminal juice using a commercial indicator paper (Pehanon) and a pH electrode (reference method). After 11 dairy cows underwent rumenocentesis, the clinical status of those animals was evaluated daily, and cows were slaughtered as well as pathologically--anatomically examined on day 7. During the observation period, the following pathological clinical signs were evident: forced inspiration (3 cows), transient episode of hyperthermia (2 cows), increased tension of the abdominal wall (8 cows) and positive foreign body tests (3 cows). One cow had to be culled on day 7 because of severe generalised septic peritonitis spreading from the site of rumenocentesis. At slaughter, hematoma formation in the area of the puncture site was found in 9 out of 10 cows. It was concluded that the severe complications encountered with this technique do not legitimate rumenocentesis as a routine procedure for collection of rumen juice samples in cows under Swiss conditions. The correlation between the pH reference method and the commercial indicator paper was the high (r = 0.926).