Capillary electrophoresis procedures for serum protein analysis: comparison with established techniques.

Methods using automated capillary electrophoresis (CE) instrumentation are available for serum protein electrophoresis with monoclonal band quantitation, isoelectric focusing and sodium dodecyl sulphate-polyacrylamide gel electrophoresis separations. The advantages of CE over previous gel methods relate to the time and labour saved by the automated instrumentation. High pI monoclonal bands and cryoglobulin specimens can be successfully analysed by CE. However, if the CE application uses a standard company supplied kit, then the cost savings are often negated by the high cost of the kit. Improvements such as the inclusion of both a UV-Vis as well as a fluorescence detector as standard within the one commercial instrument, the production of coated IEF capillaries with a useful life of at least 100 samples, and the introduction of a capillary array into all commercial instrumentation would ensure greater use of CE within both the clinical and other protein laboratories.

Capillary electrophoresis as a clinical tool.

Clinical laboratories must produce accurate results for patients with a minimum turn-around time. Automated commercial capillary electrophoresis instrumentation has been available to the clinical laboratory for the past five years. Our laboratory has utilised capillary electrophoresis (CE) to automate serum protein electrophoresis. We have used the technique of CE to produce clinical results for nearly two years. CE methods are also available for the quantitation of haemoglobin variants, by both isoelectric focusing and free solution techniques. Micellar electrokinetic separations by CE have been developed for some specialised drug assays and for B-group vitamin analysis, while gel-filled capillaries have the capability to separate DNA fragments, such as PCR products. Isoenzyme analysis has shown possibilities by CE, but quantitative results are needed to be clinically useful. Analysis of amino acids for newborn screening programs and as an arterial clotting indicator are being developed. The next five years should see a proliferation of clinical laboratory methods using automated CE.

Evaluation of serum protein separation by capillary electrophoresis: prospective analysis of 1000 specimens.

To critically assess the method of capillary electrophoresis (CE) we examined 1000 prospective serum samples submitted for protein electrophoresis by both high-resolution agarose gel electrophoresis (HRAGE) and CE. CE was performed using a 72 cm (50 cm to detector) x 50 microns I.D. fused-silica capillary with detection of absorbance at 200 nm. The 1000 samples examined contained 362 monoclonal paraproteins with concentrations ranging from 1 to 71 g/l. We evaluated the individual paraprotein correlations, the overall correlation between the two methods being 0.96. We found that HRAGE gave slightly higher values for the monoclonal bands than CE and the difference was statistically significant. We conclude that CE is a viable alternative to HRAGE for the determination of protein dyscrasias in a routine clinical laboratory.

Quantification of serum proteins using capillary electrophoresis.

Capillary electrophoresis is a technique that can be automated for the separation of charged particles. By investigating suitable sample dilution and injection time and adhering to a strict washing procedure we have been able to quantify paraproteins in serum samples. This has enabled us to use the technique of capillary electrophoresis for the provision of serum protein electrophoresis in a routine clinical laboratory. We present our findings of 260 serum samples, which included 76 samples with paraproteins analysed by both capillary electrophoresis (EC) and high resolution agarose gel electrophoresis (HRAGE). CE was able to detect all the monoclonal bands detected by HRAGE, and in particular, better able to detect IgA monoclonal bands occurring in the beta region. The major advantages of CE over HRAGE relate to the automated nature of CE with the elimination of the need for a densitometer.

Education and clinical biochemistry in Australia.

Education of clinical biochemists within Australia occurs primarily after the individual has attained a primary degree (Science or Applied Science) and is employed within a clinical laboratory, public or private. In the case of medical graduates, professional education is conducted under the auspices of the Royal College of Pathologists of Australia and results in the successful candidate obtaining the status of Fellow of that professional body. While scientists have a number of means of obtaining postgraduate qualifications in clinical biochemistry it is the Membership and Fellowship examinations of the Australian Association of Clinical Biochemists (AACB) which have greatest recognition within the profession. The majority of the continuing education of, and training programmes for, clinical biochemists are undertaken by the AACB. Currently, there are no formal registration requirements for laboratory scientists within Australia.

Identification and quantitation of human urine proteins by capillary electrophoresis.

Capillary electrophoresis, as a fully automatable, powerful analytical technique, has the potential for use in clinical laboratories for separation and identification of protein types in normal and abnormal human urine specimens. Abnormal urine specimens containing previously identified proteins, anion-exchange resin treatment and addition of known components were used to identify and determine migration times of the peaks found in the electropherogram of human urine. The correlation coefficients between capillary electrophoresis and the currently used method of high-resolution agarose gel electrophoresis for Bence Jones protein and albumin were found to be 0.95 and 0.93, respectively.

The effects of substituting frusemide for a thiazide diuretic in the drug regimens of patients with essential hypertension.

Eighteen patients with mild to moderate hypertension on a drug regimen which included a thiazide diuretic had the latter substituted by frusemide for twelve weeks after an initial two-week placebo wash-out period. Blood pressure and heart rate and a number of plasma and urinary biochemical indices were measured. Significant findings included a reduction in standing blood pressure and an elevation of plasma sodium, potassium, chloride, osmolarity, creatinine and alkaline phosphatase levels at the end of the twelve week frusemide phase relative to the values on the thiazide. However the means for all the biochemical indices remained within the normal laboratory reference limits. In the 24-hour urinary studies, no significant findings emerged, apart from an elevated calcium. The foregoing suggest that frusemide is an effective component of an anti-hypertensive drug regimen and that in a dose of 40 mg/day it produces no detectable perturbations of plasma electrolytes. The significance of the enhanced levels of urinary calcium excretion in conjunction with the augmented plasma alkaline phosphatase is unclear.

The tubular maximum for calcium reabsorption: normal range and correction for sodium excretion.

Increased tubular reabsorption of calcium is one of the three variables which can contribute to the pathogenesis of hypercalcaemia. It is therefore important to establish the normal range for this variable in a manner which allows for its variation with the plasma calcium concentration. Graphic methods depicting the relationship between urinary calcium excretion and plasma calcium concentration are valid but cumbersome and imprecise. The notional tubular maximum for calcium reabsorption (TmCa) has therefore been calculated in 130 healthy young subjects and a normal range of 1.75-2.61 mmol/l of glomerular filtrate established. Owing to the dependence of urinary calcium on urinary sodium, TmCa was negatively related to sodium excretion. Because the latter was higher in the males than the females, mean TmCa was slightly (but not significantly) lower in our male than our female subjects. The normal range of TmCa, corrected to zero sodium excretion, is 1.98-2.76 mmol/l of glomerular filtrate. The TmCa was also calculated using plasma calcium values corrected for albumin concentration. The range of TmCa using both corrections is 1.98-2.71 mmol/l of glomerular filtrate.