Recent evidence indicates that different forms of stress, including hypoxia, can induce specific proteins called heat-shock or stress proteins in various types of mammalian cells. These studies examined whether myocardial ischemia can result in increased levels of proteins with molecular weight and isoelectric point characteristics similar to those described for heat-shock or stress proteins. The left anterior descending coronary artery of the dog heart was completely occluded; normal and ischemic myocardial samples were obtained 6 hours after occlusion; and total cardiac proteins and RNA were prepared. Ribonucleic acid was translated in vitro in a modified rabbit reticulocyte lysate system, and [35S]-methionine-labelled translational products as well as unlabelled cardiac proteins were separated by two-dimensional gel electrophoresis. Total proteins were visualized by silver staining and in vitro translation products quantified by fluorometry. A translatable mRNA coding for a 71,000 dalton peptide with an isoelectric point of 5.8 was markedly increased in the ischemic myocardium after 6 hours of ischemia. A protein with similar migration characteristics was detected in ischemic myocardium but not in normal myocardium. These results indicate that an mRNA coding for a translational product with similar migration characteristics of heat-shock protein 71 is induced by ischemia in the dog heart.

A simple, fast, efficient, and accurate automated method is described for determining the quantity of glucose in 50 µl of serum or plasma. A hexokinase/glucose-6-phosphate dehydrogenase reaction requiring incubation at room temperature is used. The method is specific for glucose, and is not affected by increased concentrations of endogenous chemical substances in uremic sera or by several common drugs that interfere with other glucose methods. Results of parallel assays, in which glucose oxidase and o-toluidine are used, are compared.

This is an automated, cost-efficient enzymatic method for determining ammonia in plasma. In most assays for ammonia the reagents are stable for only one or two days at 4 degrees C, but the reagents in our method are stable for at least 14 days. Our method was developed for use with a Cobas-Fara analyzer but can be easily adapted to other automated analyzers. It is essential to use closed sample vials. Our results, obtained at much less cost, correlate well with those by the Du Pont aca method. Precision studies gave the following results: within-run (n = 20 each), for x = 46 mumol/L, SD = 0.7 mumol/L, CV = 1.6%, and for x = 593 mumol/L, SD = 4.0 mumol/L, and CV = 0.7%; day-to-day precision during a year: for x = 47.7 mumol/L, SD = 2.4 mumol/L, CV = 5.0% (n = 360); and for x = 643 mumol/L, SD = 20.5 mumol/L, and CV = 3.2% (n = 365). The standard curve was linear to 1000 mumol of NH3 per liter.

Alkali denaturation is the most commonly used technic to estimate fetal hemoglobin in red blood cells. The Betke and colleagues method (Nature 184: 1959; 1877-78) using cyanmethemoglobin (HiCN) was recommended by an International Committee for Standardization in Hematology for fetal hemoglobin levels between 2 and 40%. We showed that precision with samples containing up to 5% fetal hemoglobin can be considerably improved by measuring the absorbance of HiCN at 420 nm rather than at 540 nm because the molar absorptivity is 10 times greater in the Soret band. To determine optimum conditions for the assay, we studied the kinetics and stoichiometry of conversion of hemoglobin to HiCN with various concentrations and mixtures of ferricyanide, kinetics of alkali denaturation, ammonium sulfate precipitation of denatured HiCN, filtration as opposed to centrifugation for separating denatured HiCN, stability of HiCn in water and KCN solutions, and linearity of absorbance in the Soret band. With our modified procedure, the CV with normal amounts (less than 1%) of fetal hemoglobin improved from 28% to 8.7%. The CVs with 6.8% and 55% fetal hemoglobin were 7.0-4.4% respectively, and linear estimates were obtained with cells containing up to 50% fetal hemoglobin. We conclude that our modified alkali denaturation procedure yields reliable and reproducible estimates of fetal hemoglobin over a wide range of concentrations.