Evaluation of an inhibitor assay to determine serum isoamylase distribution.

We tested a technique to distinguish salivary from pancreatic isoamylase using a wheat protein which inhibits salivary isoamylase. The inhibitor technique accurately reflected the preponderance of pancreatic or salivary isoamylase in sera which had been "spiked" with human pancreatic or salivary isoamylase. Comparison of the results of either cellulose acetate electrophoresis or isoelectric focusing showed an excellent correlation (r = 0.99) in 43 hyperamylasemic sera which did not contain macroamylase. Normal values obtained in 160 healthy subjects indicated that pancreatic isoamylase comprised 41 +/- 21% (1 SD) of total serum amylase activity and the upper limit of normal for serum pancreatic isoamylase was 166 IU/liter. The inhibitor assay provides a simple and accurate means of differentiating salivary from pancreatic hyperamylasemia.

Diagnostic value of routine isoamylase assay of hyperamylasemic serum.

The purpose of this study was to determine if routine isoamylase assay would provide valuable diagnostic information in patients with hyperamylasemia. Isoamylase distribution was determined in sera of 37 consecutive hyperamylasemic patients. The attending physicians (without knowledge of the isoamylase level) had considered acute pancreatitis to be "probable" in 19, "possible" in 4, and "unlikely" in 14 of these 37 patients. Three of the patients considered probably to have pancreatitis and 3 thought possibly to have pancreatitis had normal serum pancreatic isoamylase levels. Knowledge of the normal pancreatic isoamylase level in these 6 patients probably would have changed the clinical diagnosis to some condition other than pancreatitis. Of the 14 hyperamylasemic patients thought "unlikely" to have pancreatitis, 7 had an elevation of just pancreatic isoamylase and 3 additional patients had elevation of both pancreatic and salivary isoamylases. It seems likely that knowledge of these elevated pancreatic isoamylase levels would have changed the clinical diagnosis to "probable" pancreatitis for many of these patients. We conclude that routine isoamylase assay provides diagnostic information that might change the clinical diagnosis in 20%--40% of hyperamylasemic patients.

Measurement of ultrafiltrable calcium in serum with use of the "Worthington Ultrafree Anticonvulsant Drug Filter".

We describe a method for ultrafiltrable calcium involving the use of Worthington Ultrafree Anticonvulsant Drug Filters. When measured at 37 degrees C, with mineral oil covering the sample to prevent loss of CO2, values for ultrafiltrable calcium correlated reasonably well (r = 0.91) with those for ionized calcium as measured with an ion-selective electrode. All patients' samples with significantly high or low values for ionized calcium were identified by the ultrafiltration method, including one specimen for which the McLean-Hastings protein correction could not explain the discrepancy between ionized and total calcium. The method requires about 2 mL of serum, yields about 100 microL of protein-free ultrafiltrate, and with it any laboratory with a semi-micro calcium method can measure ultrafiltrable calcium.

An enzymatic approach to lipoprotein quantification.

Lipoprotein cholesterol levels were determined without ultracentrifugation by using modified enzymatic methods for cholesterol, high-density-lipoprotein (HDL) cholesterol, and triglyceride and the formula, low-density-lipoprotein (LDL) cholesterol = total cholesterol-HDL cholesterol-triglycerides/5. The methods for cholesterol and triglyceride determinations were standardized for accuracy and precision by the Center for Disease Control's Lipid Standardization Laboratory, which monitored this laboratory for 16 months. The lipoprotein cholesterol values obtained correlated well with lipoprotein cholesterol values determined at the Minnesota Lipid Research Clinic Laboratory using ultracentrifugation. LDL cholesterol determined at the Minneapolis Veterans Administration Hospital Laboratories (Y axis) produced a curve with an intercept of 9.38 mg/dl, a slope of .977, standard error of the estimate (Sy.x) of 8.8 mg/dl, and a correlation coefficient (r) of .983 (n = 32). HDL cholesterol was Y = 0.998 X + .89 mg/dl, Sy.x = 1.6 mg/dl (r = .984, n = 53), and very-low-density-lipoprotein (VLDL) cholesterol was Y = 1.010 X -1.32 mg/dl, Sy.x = 1.3 mg/dl (r = .996, n = 54).

An enzymatic triglyceride method that is suitable for long-term population studies.

An enzymatic triglyceride method has been shown to be a suitable alternative to the Lipid Research Clinics' extraction/fluorometry method in long-term population studies. Correlation of results obtained with this method by this laboratory (y-axis) and by the Minneapolis Lipid Research Clinic Laboratory (x-axis) during a nine-week standardization period produced a curve with an intercept of -72 mg/liter, a slope of 1.019, and a correlation coefficient of r=0.996 (n=47). During this standardization period certain methodological problems were observed and corrected. An increase in background in certain clinical specimens, caused by spontaneous degradation of NADH, was observed, accurately measured, and taken into account when appropriate.

Enzymatic determinations of cholesterol in high-density-lipoprotein fractions prepared by a precipitation technique.

An enzymatic method for cholesterol in serum [Clin. Chem. 20, 470 (1974)] was initially found to be unsatisfactory for measuring cholesterol in high-density-lipoprotein fractions prepared by precipitation with Mn2+. A fine precipitate formed in the cuvette and cholesterol values were falsely increased. We describe a simple, convenient method for circumventing these problems. An ethylenediaminetetraacetate solution is used to reconstitute the enzymatic reagent. Cholesterol values by this procedure correlated with those obtained by the Lipid Research Clinic's procedure for the same lipoprotein fraction preparations (regression slope, .998; Y-intercept, 8.9 mg/liter; correlation coefficient, .984; standard error of the estimate, 16.8 mg/liter). Precision of the assay, including the precipitation step, was calculated. The SDwithin day was 9.7 mg/liter and SDoverall was 23.7 mg/liter. Results for total cholesterol with the modified reagent were linearly related to concentrations exceeding 4 g/liter, thereby permitting determination of high-density-lipoproteins and total cholesterol in a single run.

An example of lyophilized protein-based materials not simulating patient sera.

Use of reconstituted lyophilized protein-based materials in the clinical laboratory is partly based on the assumption that these materials adequately simulate patients' sera. We examined several of these materials and found that certain ones do not have the same adsorbancies at 340 and 380 nm as do most sera. The implication of this is examined with respect to glucose determination by the hexokinase method on a dual-wave-length blank-subtraction instrument.