Creatine phosphokinase isoenzymes: A comparison of a kinetic and an electrophoretic method in the diagnosis of myocardial infarction

A comparison has been made of the results of the levels of the MB isoenzyme of creatine phosphokinase by a kinetic and an electrophoretic method performed on patients suspected of having myocardial infarction. Reviewed in depth are those cases in which discrepant results have occurred. On the basis of the combined clinical and laboratory data, the kinetic method is a more sensitive but less specific indicator of myocardial necrosis. Thus, the kinetic method may be used as a screening test, with confirmation of positive results by electrophoresis.     

Electroradioimmunoassay: a sensitive method for the quantitation of alpha-fetoprotein.

A sensitive technique, electroradioimmunoassay has been adapted for the estimation of alpha1-fetoprotein in normal adult human plasma and serum. Commercially available antisera were used and a 10-fold increase in sensitivity was obtained in comparison with two conventional radioimmunoassays. Plasma alpha 1-fetoprotein levels of 2.6 +/- 1.2 microgram/1 (mean and standard deviation) were found in a normal population as compared with levels of 4.1 +/- 2.6 microgram/l in a group of 49 patients with alcoholic liver disease.     

Glycosylated plasma protein: a simple method for the elimination of interference by glucose in its estimation

Interference by glucose in the colorimetric estimation of glycosylated plasma protein was effectively eliminated by the precipitation of the proteins with trichloroacetic acid. The procedure was rapid and the recovery of protein quantitative. Results were highly reproducible.Preliminary clinical data obtained using the modified procedure showed rapid reflection by glycosylated plasma protein of the short-term control of glucose levels in diabetics when glycosylated haemoglobin values failed to indicate changes.     

Mucolipidosis I: Studies of sialidase activity and a prenatal diagnosis

The characteristics of the sialidase (N-acetyl-α-neuraminidase) of human leukocytes, fibroblasts and amniotic fluid cell cultures were determined with a radioactive assay method utilizing neuramin-[³H]actitol as the enzyme substrate. Fibroblast cultures from patients with the inherited sialidase deficiency diseases including mucolipidosis I, sialidosis I and sialidosis II, juvenile type have less than 10% of normal sialidase activity using either this substrate, 2-(3′-methoxyphenyl)-N-acetyl-α-neuraminic acid, or 2′-(4-methylumbelliferyl)-Nacetyl-α-neuraminic acid. The total sialic acid content of fibroblasts and leukocytes from mucolipidosis I and sialidosis I patients is greatly elevated; this parameter is useful in establishing a diagnosis of sialidase deficiency. The sialic acid content of sialidosis II, juvenile type, with coexistent sialidase and β-galactosidase deficiencies, is only slightly elevated above normal levels. A patient with mucolipidosis I has 16% of normal neuramin-[³H]lactitol sialidase activity in his peripheral leukocytes. His parents were clearly distinguished from the normal range using leukocyte enzyme levels and a maternal aunt was identified as a possible carrier. The presence of this enzyme in amniotic fluid cell cultures, both fibroblastic and mixed cell type, makes possible the prenatal detection of these diseases. A pregnancy from a family at risk for having a child with mucolipidosis I was monitored by amniocentesis and subsequent sialidase measurement of the amniotic fluid cell culture.     

A rapid method for assay of branched-chain keto acid decarboxylation in cultured cells and its application to prenatal diagnosis of maple syrup urine disease

A sensitive assay for measurement of branched-chain keto acid decarboxylation in small numbers of fibroblasts or amniotic cells grown in the wells of a microtitre plate using [1-14C]leucine as substrate is described. The method was applied to the amniotic cells from a pregnancy at risk for maple syrup urine disease and a heterozygous fetus predicted.     

Determination of 3,4-dihydroxyphenylethyleneglycol in urine by gas chromatography with a flame ionization detector: a new rapid method.

3,4-Dihydroxyphenylethyleneglycol, a major metabolite of noradrenaline in rat brain, is estimated alone or with 4-hydroxy-3-methoxyphenylethyleneglycol in rat and human urine by gas chromatography with a flame ionization detector. The samples are hydrolyzed and extracted at pH 2 with ethyl acetate. Then, to analyze only 3,4-dihydroxyphenylethyleneglycol the reaction with n-butaneboronic acid is carried out directly; if 4-hydroxy-3-methoxyphenylethyleneglycol also has to be estimated, preliminary acetylation in alkaline aqueous solution is performed. The advantages of the specificity due to the reagents used is discussed.     

Determination of chromium sesquioxide in faeces by a spectrophotometric method

Chromium sesquioxide Cr2O3, present as a non-absorbable marker in faeces, may be determined spectrophotometrically as chromate ion in aqueous solution after ashing and alkaline fusion. Recovery of this substance is excellent. The method described is simpler, more suited to the clinical laboratory and less hazardous than previously reported methods.     

Evaluation of automated glucose oxidase methods for serum glucose: Comparison to hexokinase of a colorimetric and an electrometric method

Two automated glucose oxidase methods have been evaluated with respect to accuracy, precision, recovery, linearity and various potential interferences.Trinder's method on an AutoAnalyzer II had a between-day coefficient of variation (C.V.) of 2.6% (mean 228 mg/dl), was linear to 500 mg/dl, and produced a mean recovery of 99.7%. Comparison of Trinder's method with a manual, blanked hexokinase method yielded the regression equation: TR = ? 1.95 + HEX (1.04); Spearman's rho correlation coefficient was: 0.974.The Beckman System I glucose method had a between-day C.V. of 1.6% (mean 198 mg/dl), was linear to 500 mg/dl, and recovered an average of 98.0% of added glucose. Comparison with the same hexokinase method yielded: SYI = 1.27 + HEX (1.02): Spearman's rho = 0.991.None of the possible interfering compounds tested caused significant deviation of results by either method within the range of concentrations encountered physiologically.Trinder's method on the AutoAnalyzer II and the System I method are accurate, precise methods and are highly recommended for routine use in the clinical laboratory.     

An alternative protein precipitation method for antipyrine estimation in plasma

The isoenzymes of aspartate transaminase differ in their kinetic properties in that the cytoplasmic isoenzyme is more readily inhibited by adipate and by 2-oxoglutarate (substrate) at low pH. A differential kinetic assay based on this phenomenon has been optimised for use in assays of serum samples. The new method agrees well with an immune absorption procedure. Methods based on Chromatographic separation of the isoenzymes fail in the presence of serum.     

Dihydropteridine reductase deficiency: diagnosis by leukocyte enzyme assay

An assay procedure for dihydropteridine reductase in peripheral leukocytes is described. The assay utilizes the tetrahydropterin-dependent reduction of ferri-cytochrome C in the presence of NADH and requires a smaller number of cells than assays described for cultured skin fibroblasts.Dihydropteridine reductase activity was not detectable in the peripheral leukocytes nor in the cultured skin fibroblasts from two adolescent patients with malignant hyperphenylalaninemia. The parents of the patients showed approximately 50% of normal dihydropteridine reductase activity in their peripheral leukocytes.Immunochemical experiments using antibodies against bovine liver dihydropteridine reductase suggest that normal leukocytes and skin fibroblasts contain dihydropteridine reductase which is immunologically similar to that of human liver.The present studies indicate that the determination of dihydropteridine reductase activity in peripheral leukocytes can be used to diagnose hyper-phenylalaninemia due to a defect in dihydropteridine reductase.     

A method for identification and follow-up of patients with a steroid-21-hydroxylase deficiency.

A method is described for the determination of 17 alpha-hydroxyprogesterone from blood samples obtained by heel prick and dried on filter paper. Discs (10 mm diameter) were cut from the filter paper and extracted in the assay tubes with 1 ml of a methanol/diethyl ether/ethyl acetate (50 :45 :5, v/v) solvent mixture. Antibody, tritium-labelled tracer and dextran-coated charcoal were added to assay tubes using a multichannel dispenser. The approach used permits one technician to analyze two series, each of 60 duplicate samples, within one working day. Thus the method is applicable for the centralized screening of suspected cases, while emergency samples may be analyzed at the same time within 4 h. Comparisons with a highly specific but more elaborate technique for the determination of blood 17 alpha-hydroxyprogesterone showed a correlation coefficient of 0.99, and the regression equation for the present method (y) against the established method (x) was y = 1.07x + 2.72. The calculated upper reference limit (mean +/- S.D.) for 17 alpha-hydroxyprogesterone in healthy infants from two days to eight years of age of 7.5 ng/ml of serum.     

Albumin I.M.V.S.: a variant detected by a Bromcresol Green dye binding method

A unique variant of human albumin was discovered in a family following routine multiple biochemical analysis. Their sera demonstrated increased Bromcresol Green (BCG) dye binding under certain reaction conditions (excess amounts of the detergent Pegosperse in the presence of Merthiolate). This effect was also seen with a structurally similar dye, Bromcresol Purple (BCP). The variant sera also showed an altered pattern of BCG dye binding (in the presence of Merthiolate and excess Pegosperse) compared to normal sera when the temperature or the pH of the dye reagent was changed. Decreased binding of the dye 2-(4'-hydroxybenzeneazo)benzoic acid (HABA) was also noted. Similar findings were observed in six other patients over a period of 21 months. Column chromatography of the variant sera on a DEAE-Sepharose column separated the albumin into two fractions, one showing normal dye binding properties, the other showing the more extreme pattern of dye binding seen for the whole variant serum.     

Inorganic phosphorus measurement: An improved method

An improved micromethod for the determination of inorganic phosphorus in serum and urine is outlined. The procedure requires no deproteinization and yields a stable colour. The new formulation proposed avoids the pitfalls of other techniques. The method is both accurate (recovery 99–100.4%) and precise (C.V. 2.19%).     

Estimation of cholesterol in bile: Assessment of an enzymatic method

Cholesterol in bile has been estimated using two variants of the cholesterol oxidase method and compared with gas-liquid chromatographic (GLC) assay. As published, the enzymatic method seriously underestimated biliary cholesterol. This was due to interference by other constituents of bile, particularly bilirubin and lecithin. However, by dilution of reagents and samples in water and isopropanol respectively a close correlation with the GLC method is achieved. It is concluded that the cholesterol oxidase method as published is unsuitable for the estimation of cholesterol in bile, but in modified form it is simple, accurate and economical.