Temperature effects on free-thyroxine measurements: analytical and clinical consequences.

We compared the analytical and clinical performance of two free-thyroxine (FT4) assays--a solid-phase radioimmunoassay, Spectria, and a time-resolved fluoroimmunoassay, Delfia, both of them two-step methods--with the performance of a direct radioimmunoassay, Nichols, to measure FT4 concentration in equilibrium dialysate of undiluted serum. The three assays showed comparable analytical performance. We tested clinical utility in sera from 135 healthy subjects with and without thyroxine-binding abnormalities and in 61 patients with and without thyroidal illnesses. We found significant differences for FT4 measured by different assays in sera from the same euthyroid patients. To explain the differences, we studied the influence of temperature on performance and calibration. Most important was the neglected fact that the association constant for the binding of thyroxine to thyroxine-binding globulin decreases when the temperature rises from 20 to 37 degrees C, causing a doubling of FT4. The two-step assays, if performed at room temperature without a well-defined calibration, can give misleading FT4 concentrations. This is the case when sera from patients with thyroxine-binding abnormalities are measured against kit standards, made up in normal human sera. If an assay is to reflect the in vivo FT4 concentration at body temperature in all types of samples, it should be performed at body temperature. For practical reasons 37 degrees C is recommended, and reference values should be defined at 37 degrees C. The same might be valid for other free-hormone assays.

The use of human placental lactogen, oxytocinase and estriol in twin pregnancies with intrauterine growth retardation.

In 38 twin pregnancies human placental lactogen (HPL) and oxytocinase were measured in serum and estriol in urine. Eleven women carried light-for-dates (LFD) twins (birth weight less than P5), and 27 had appropriate-for-dates twins (birth weight greater than P5). We investigated if intrauterine growth retardation in these patients could be detected either by the biochemical measurements of by clinical examination. The values of oxytocinase and estriol were not related to fetal weight. A serum level of HPL of 12 mg/l after 34 wk of gestation proved to be helpful in the detection of LFD twins. The predictive value of a positive test was 64% (7/11) and of a negative test 84% (21/25). Eight LFD twins were suspected clinically. If clinical examination and HPL measurements were used in conjunction all LFD twins could have been suspected before birth.

Vitamin B-1, B-2 and B-6 deficiencies in geriatric patients, measured by coenzyme stimulation of enzyme activities.

The vitamin B-1, B-2 and B-6 nutritional status of 153 geriatric patients was determined by measurement of the activities of transketolase (TK), glutathione reductase (GR) and glutamic-oxaloacetic transaminase (GOT) from hemolyzed erythrocytes before and after in vitro stimulation with their respective coenzymes. The change in enzyme activity after incubation of the hemolysate with the coenzyme was used to determine the activation coefficient, which was taken as an index for the vitamin B-1, B-2 and B-6 nutritional status. Determination of the normal values in 54 healthy blood donors showed that activation coefficients of TK greater than 1.27 indicated a biochemical vitamin B-1 deficiency. Activation coefficients of GR greater than 1.29 and GOT greater than 1.86 indicated, respectively, deficiencies of vitamins B-2 and B-6. On the basis of these findings 22.9% of the geriatric patients appeared to suffer from vitamin B-1 deficiency, 11.7% from vitamin B-2 deficiency and 19.0% from vitamin B-6 deficiency. Of the total number of patients, 44% showed a deficiency of one or more of these three vitamins. Oral administration of vitamin B-1 (20 mg/day), vitamin B-2 (10 mg/day) and vitamin B-6 (20 mg/day) for twelve days normalized nearly all activation coefficients. Determination of enzyme activities without coenzyme stimulation revealed significantly lower values in the deficient patients as compared with the blood donors. However, the distribution of activities for both groups overlapped to a great extent. Oral administration of vitamins raised the enzyme activities to normal values.

Effects of calcium ions on pyruvate kinase from human erythrocytes.

Ca2+ ions have a biphasic effect on the allosteric pyruvate kinase (EC 2.7.1.40) from human erythrocytes: Ca2+ is an activator at low phosphoenolpyruvate (PEP) concentrations: at increased PEP concentrations Ca2+ behaves as an inhibitor. In the presence of ATP the same effect was observed and at low PEP concentrations Ca2+ ions can completely abolish the ATP inhibitory effect. At high Ca2+ concentrations there is a loss of the cooperativity towards PEP. The enzyme activated by fructose-1,6-diphosphate (FDP) is inhibited by Ca2+ ions at all concentrations of PEP tested. Mg2+ ions are not able to counteract the activation by Ca2+ ions at low PEP concentrations. The results are interpreted on the basis of the model of Monod.