Lennart Waldenlind

Affinity electrophoresis of human serum alkaline phosphatase isoenzymes in agarose gel containing lectin

Separation of alkaline phosphatase isoenzymes using affinity electrophoresis in agarose gel containing lectin is described. The bone and biliary isoenzymes precipitate during electrophoresis and are clearly separated from the liver isoenzyme. The liver, intestinal and placental alkaline phosphatases are essentially not affected by the lectin. The migration distances of the precipitating bone and biliary fractions vary with their alkaline phosphatase activity. The bone isoenzyme is more heterogeneous than the biliary isoenzyme with respect to interaction with lectin forming both insoluble and soluble complexes. Affinity electrophoresis in agarose gel containing lectin can be used for quantitation by densitometry of liver and bone isoenzymes in sera containing only these two fractions but must be combined with conventional electrophoresis, preferably in agar gel, if biliary, intestinal, or placental isoenzymes are also present.

Characterization of serum alkaline phosphatase isoenzymes by affinity electrophoresis in agarose gel containing lectin combined with agar gel electrophoresis

The combined use of affinity electrophoresis in agarose gel containing lectin and of agar gel electrophoresis for the quantitation of liver, bone, biliary and intestinal alkaline phosphatase isoenzymes is described. Sera from patients with various diseases and from normal subjects (blood donors) have been analyzed. Data from normal subjects show that the bone isoenzyme is the predominant fraction (about 62%) in adults. The relative proportions of the alkaline phosphatase isoenzymes are similar in both sexes in adulthood (21-50 years). The higher alkaline phosphatase activity found in men than in women (ages 21-50 years) is due to higher values for both liver and bone isoenzymes. The difference between men and women tends to decrease after the age of 50 mainly due to an increase of the bone isoenzyme in women.

An improved method for the determination of endogenous thiamine and its phosphate esters in biological material.

Nonphosphorylated thiamine was extracted from tissue homogenates in saline or sucrose into ethylbutylketone-acetonitrile using counter-ion extraction and reextracted into hydrochloric acid. The dried samples were then analyzed by the fluorimetric thiochrome method. Phosphorylated thiamine was treated with Takadiastase to split off the phosphate groups and was thereafter extracted and measured. No column chromatography was needed for the separation of thiamine before the thiochrome assay. A higher accuracy and sensitivity for the thiamine assay was thus achieved.

Creatine kinase‐MB mass concentration versus creatine kinase‐B activity for the detection of acute myocardial infarction in patients with slightly elevated total creatine kinase activity in serum

Abstract. Objective. The diagnostic value of creatine kinase‐MB mass concentration (CK‐MB mass) was compared with that of creatine kinase‐B (CK‐B) activity in patients with suspected acute myocardial infarction (AMI) but with total serum CK activity only slightly above the reference range.

Inherited occurrence of a heat stable alkaline phosphatase in the absence of malignant disease

We describe a family with an inherited persistent elevation of serum alkaline phosphatase activity in the absence of malignant disease, observed for at least 15 yr. Isoenzyme studies revealed that this increased activity was due to an enzyme which showed similarities to serum placental alkaline phosphatase from pregnant women having the following properties: high heat stability; reactivity to anti-placental alkaline phosphatase antiserum; lack of inhibition by L-homoarginine; moderate inhibition by EDTA; and lack of interaction with wheat germ lectin. The enzyme was less sensitive than placental alkaline phosphatase to inhibition by L-phenylalanine, L-tryptophan, L-leucine, L-leucyl-glycyl-glycine and L-phenylalanyl-glycyl-glycine. The enzyme also differed from the placental alkaline phosphatase in its electrophoretic mobility, isoelectric heterogeneity and apparent molecular mass. We conclude that the enzyme is an inherited heat stable alkaline phosphatase variant which might correspond to a rare phenotype of placental alkaline phosphatase.

Radio-enzymatic assay of acetylcholine in tissues

Abstract A method for the determination of ACh in tissues and subcellular fractions is described. The ACh is extracted by an ion pair exchange method, purified by paper chromatography and determined by enzymatic phosphorylation with choline kinase after enzymatic hydrolysis with AChE.

FURTHER CHARACTERIZATION OF A HEAT-STABLE ALKALINE PHOSPHATASE WITH LOW SENSITIVITY TO L-PHENYLALANINE

A heat-stable alkaline phosphatase, hitherto found in two families with inherited hyperphosphatasemia, was further characterized. The enzyme was similar to serum placental alkaline phosphatase from pregnant women concerning its apparent affinity constant (Km) for 4-nitrophenyl phosphate and its reactivity with H7 monoclonal anti-placental alkaline phosphatase (PLAP) antibodies, but different in the following respects: it exhibited greater heat stability, a higher pH optimum, lower sensitivity to inhibition by L-phenylalanine, and no reactivity with C2 monoclonal anti-PLAP antibodies. The low sensitivity to L-phenylalanine suggests that the enzyme might correspond to a rare phenotype of placental alkaline phosphatase found in human term placenta.

Evidence for the formation of 2-methylthiamine in nerve tissue

Abstract Rats and cats were injected with [ 3 H]choline, [ 14 C]choline or [ 35 S]thiamine. Choline and thiamine metabolites were extracted from different tissues by ion-pair extraction. Different Chromatographic techniques and the use of two separate precursors-choline and thiamine-showed that a fraction of the radioactivity was incorporated into a [ 3 H]2-methylthiamine-like substance. A relationship was found between the amounts of radioactive acetylcholine and the 2-methylthiamine-like substance formed.