bluuebunny/crossref_metadata_2025 huggingface dataset:
Improved liquid-chromatographic determination of cyclosporine, with concomitant detection of a cell-bound metabolite.
Authors: Lensmeyer G L , Fields B L
Published: January 2020

This unique extraction and isocratic "high-performance" liquid chromatographic method for measuring cyclosporine (CsA) in blood involves a Zorbax cyanopropyl analytical column maintained at 58 degrees C, with detection at 214 nm, and recycling of the water:acetonitrile mobile phase for improved long-term column stability and efficiency. Routinely, 1.0 mL of serum, plasma, or whole blood is diluted with water:acetonitrile (70:30) and applied to a disposable solid-phase cyanopropyl column to rapidly extract the drug and the internal standard cyclosporin D (CsD). Analytical recovery for this step averages 90% with whole blood and 98% with serum and plasma. Between-run CVs were 6.5 and 2.6% for means of 104 and 1128 micrograms/L, respectively. The standard curve is linear up to 1600 micrograms/L. The minimum detection limit is 10 to 15 micrograms/L. No interferences from endogenous substances or other drugs were found. In addition, a compound cross reacting with the Sandoz radioimmunoassay antibody was isolated from patients' samples with the present procedure and was tentatively identified as a CsA metabolite(s). It appears to be highly partitioned on blood cells, very little being detected in the serum or plasma. In a comparison with RIA, correlation coefficients were 0.828 and 0.652 for serum and whole blood, respectively. Results from a 12-h pharmacokinetic study in which different sample types were analyzed by RIA and liquid chromatography further exemplified major discrepancies between types of CsA determinations.
Aminophenylboronic acid affinity chromatography and thiobarbituric acid colorimetry compared for measuring glycated albumin.
Authors: Rendell M , Kao G , Mecherikunnel P , Petersen B , Duhaney R , Nierenberg J , Rasbold K , Klenk D , Smith P K
Published: January 2020

Two techniques originally developed for measurement of glycated ("glycosylated") hemoglobin but also applicable to determination of glycated albumin are the thiobarbituric acid colorimetric technique (I) and the aminophenylboronic acid affinity chromatographic procedure (II). The latter reliably distinguishes diabetics from nondiabetics, and concentrations of glycated hemoglobin and glycated albumin are linearly correlated. I is nonspecific; it neither correlates with diabetic status nor with values derived via the affinity technique. Most of the chromogenic material is present in the fraction of albumin that does not bind to aminophenylboronic acid. Glucose interferes significantly with I but only slightly with II. Prolonged incubation of plasma with glucose dramatically increases the II-determined glycated albumin. Reactivity with thiobarbituric acid increases much less, and mainly in the II-bound fraction. This fraction contains a high proportion of nonspecifically reactive material. The percentage of glycated albumin determined in crude plasma samples by II differs only slightly from the value determined by purifying the albumin from the plasma. This technique appears more promising than I for eventual clinical applications.
Mitochondrial enzymes in human serum: comparative determinations of glutamate dehydrogenase and mitochondrial aspartate aminotransferase in healthy persons and patients with chronic liver diseases.
Authors: Jung K , Pergande M , Rej R , Schreiber G , Schimmelpfennig W
Published: January 2020

We measured the activities of two mitochondrial enzymes, the mitochondrial form of aspartate aminotransferase (EC 2.6.1.1) and glutamate dehydrogenase (EC 1.4.1.2), in the serum of apparently healthy persons (n = 84) and patients suffering from chronic liver diseases (n = 43). The distribution of activities for glutamate dehydrogenase, but not mitochondrial aspartate aminotransferase, was sex-dependent. The upper limits of the reference intervals (99th percentile) at 37 degrees C were 3.2 U/L for mitochondrial aspartate aminotransferase, 6.4 U/L for glutamate dehydrogenase (women), and 11.0 U/L for glutamate dehydrogenase (men); there was a weak correlation between the activities of both mitochondrial enzymes (r = 0.439). In patients with chronic liver diseases we found a greater increase in the activity of glutamate dehydrogenase than of mitochondrial aspartate aminotransferase and the correlation between the two mitochondrial enzymes was stronger. The diagnostic sensitivity and specificity of either mitochondrial enzyme was less than that of total aspartate aminotransferase, alanine aminotransferase (EC 2.6.1.2), or gamma-glutamyltransferase (EC 2.3.2.2).
Quantification of human serum apolipoprotein AI by enzyme immunoassay.
Authors: Bury J , Rosseneu M
Published: January 2020

We developed a quantitative assay for apolipoprotein AI (apo AI) in human serum, using a "sandwich"-type enzyme-linked immunosorbent assay. Diluted serum samples were pipetted into the wells of polystyrene microtiter plates that had been previously coated with purified rabbit anti-human apo AI antibodies. After incubation for 2 h and washing, antibodies conjugated to horseradish peroxidase (EC 1.11.1.7) were added and incubated for 2 h; after further washing, the bound enzyme was assayed by oxidation of o-phenylenediamine. Assay conditions were optimized for the incubation time and the amounts of coating antibodies and conjugate. Assay sensitivity is about 0.5 ng of apo AI, with a working range of 1 to 14 ng, similar to that of radioimmunoassays for human apo AI. The standard curves for apo AI in serum or HDL and for purified apo AI were parallel. Delipidation, heat treatment, or addition of detergents did not affect the amount of immunoassayable apo AI in human serum. The intra- and interassay CVs were 4 and 8%, respectively. Results for 100 serum samples compared well with those by immunonephelometry (r = 0.94).
Measurement of lipase activity by a differential pH technique.
Authors: Ceriotti F , Bonini P A , Murone M , Barenghi L , Luzzana M , Mosca A , Ripamonti M , Rossi-Bernardi L
Published: January 2020

This is a new electrochemical method for determination of lipase activity in biological fluids, including serum, plasma, and duodenal juice. Advantages of turbidimetric methods--short reaction time, and small sample and reagent volumes--are combined with those of titrimetric methods: measurement of absolute activity (i.e., no standardization required), saturated substrate conditions, and direct measurement of reaction products. The proposed method is easy, inexpensive, and takes only 3 min. Precision is good: CV = 3.74% within day and 7.3% between days at the clinical-decision concentration, CV = 1.86% within day and 4.65% between days for above-normal lipase activities. The standard curve is linear up to 4500 U/L. Results (y) correlate well with those by turbidimetry (x): y = 0.9287x - 65.3 (r = 0.9719). Reference values are between 0 and 130 U/L.
Two automated methods for measuring plasma thiocyanate compared.
Authors: Vesey C J , Kirk C J
Published: January 2020

In an interlaboratory comparison of two continuous-flow analytical procedures for measuring thiocyanate, we used ferric nitrate (y) and p-phenylenediamine/pyridine (x) as colorimetric reagents to measure its concentrations in plasma of 100 consecutive patients attending a peripheral vascular disease clinic. The results correlated well (r = 0.987, p less than 0.001; y = 0.938x + 1.2 mumol/L). However, there were small, systematic, positive differences between the phenylenediamine values and the corresponding ferric nitrate values (paired t = 5.4, p less than 0.001). These differences were linearly related to the means of the pairs of results (r = 0.42, p less than 0.001; y = 0.0739x - 2). Nevertheless, when we used previously determined cutoff points the two sets of SCN concentrations concurred completely in classifying the 100 patients as smokers or nonsmokers. On the basis of self-classification by 71 of these subjects, the measurement techniques had a sensitivity of 91% and a specificity of 75%; when five patients claiming to be nonsmokers but found to have abnormally high values for carboxyhemoglobin (2.7 - 6.9%) were reclassified as smokers, specificity increased to 89%.
Laboratory evaluation of the Greiner G-400 discrete selective multichannel analyzer.
Authors: Degiampietro P , Peheim E , Colombo J P
Published: January 2020

We evaluated the new Greiner G-400 selective multichannel analyzer. Thirty different tests can be run in combination profile and single-test mode at a rate between 150 and 300 tests per hour. The new photometer allows kinetic and equilibrium (endpoint) measurements. The G-400 analyzer demonstrated excellent precision, linearity, accuracy, ease of operation, and no carryover. Results for 16 different analytes as determined with the Hitachi 705, GSA II, and Corning 940 correlated favorably with those obtained with the G-400. The G-400 analyzer is suitable for both emergency and routine use.
How certain drugs interfere negatively with the Jaffé reaction for creatinine.
Authors: Kroll M H , Nealon L , Vogel M A , Elin R J
Published: January 2020

Phenacemide and cephalothin may interfere either positively or negatively with the determination of creatinine by the Jaffé reaction, depending on the analytical system. The drug-picrate chromophores maximally absorb within the first minute of reaction (21 s for phenacemide, 45 s for cephalothin), after which the absorbances decrease. Thus, these drugs negatively interfere in analytical systems involving kinetic measurements when the time interval of measurement includes the period of decreasing absorbance.
Magnesium content of mononuclear blood cells.
Authors: Elin R J , Hosseini J M
Published: January 2020

The magnesium content of mononuclear blood cells may provide a better assessment of intracellular magnesium or total body magnesium status than does measurement of magnesium in plasma or erythrocytes. We describe a method for determining this, and report results for 20 normal volunteers. The mononuclear cells are separated with a discontinuous Ficoll-Hypaque gradient, washed, centrifuged at 400 X g, and lysed by sonication in 10 mmol/L NaCl. The magnesium in the cell lysate, with 2.93 g of added lanthanum oxide per liter, is determined by atomic absorption spectrometry. The mean mononuclear cell magnesium content in our volunteers was 70.7 (SD 14.1) fg per cell and 9.29 (SD 1.85) mg/g of DNA. The CVs for the determinations of magnesium, DNA, and cell count were 3.0%, 5.0%, and 8.7%, respectively. There was a significant correlation (r = 0.67, p less than 0.001) between results by the two methods of expressing magnesium content of mononuclear cells. However, by either method there were no significant correlations among results for magnesium concentration of mononuclear cells, plasma, or erythrocytes.
Isolation and preliminary characterization of a monoclonal antibody that interacts preferentially with the liver isoenzyme of human alkaline phosphatase.
Authors: Lawson G M , Katzmann J A , Kimlinger T K , O'Brien J F
Published: January 2020

We have prepared murine monoclonal antibodies against isolated human bone alkaline phosphatase (ALP, EC 3.1.3.1). Hybridoma supernates were separately screened for reactivity against both human liver and bone ALP. Although most antibody-positive hybrids showed similar reactivity against both isoenzymes, one hybridoma produced an antibody that interacted preferentially with liver ALP. This antibody was purified and used to establish an immunoassay to differentiate liver ALP from bone ALP. When equal activities of the two isoenzymes (as determined by a conventional enzymic assay) were measured by the immunoassay, a fivefold greater response was obtained with liver than with bone ALP. The immunoassay can be used to measure the proportions of the bone and liver isoenzymes in mixtures of them. Cross reactivity with human placental and intestinal ALP is less than 3% relative to liver ALP. These findings support the feasibility of developing immunological methods to differentiate these isoenzymes in the clinical laboratory.
Effect of hematocrit on cyclosporine (cyclosporin A) in whole blood and plasma of renal-transplant patients.
Authors: Rosano T G
Published: January 2020

Cyclosporine concentrations in whole blood and plasma were determined in 25 samples from renal-transplant patients. The portion of cyclosporine carried in plasma varied widely, from 9 to 58%. Radioimmunoassay and liquid chromatography both revealed a statistically significant inverse correlation between hematocrit and the plasma fraction of cyclosporine in the circulating blood. Regression analysis indicates that a 10% increase in hematocrit would decrease the portion of cyclosporine in plasma by 12 to 14%. Studies with cyclosporine-supplemented samples further demonstrated an effect of hematocrit on the drug concentration in plasma without a corresponding effect on the concentration in whole blood. The distribution of cyclosporine between plasma and erythrocytes was unaffected by plasma cholesterol and triglycerides for these transplant patients.
The Technicon RA-1000 evaluated for measuring sodium, potassium, chloride, and carbon dioxide.
Authors: Ng R H , Altaffer M , Ito R , Statland B E
Published: January 2020

We evaluated the Technicon RA-1000 "random-access" analyzer for the measurements of sodium, potassium, and carbon dioxide by an indirect potentiometric method (ion-selective electrode) and for chloride by a colorimetric method (mercuric thiocyanate). For various concentrations of control materials the total precision (CV) ranged from 0.9 to 1.2% for sodium, 1.1 to 1.3% for potassium, 1.0 to 1.2% for chloride, and 2.8 to 3.8% for carbon dioxide. The system demonstrated acceptable performance in linearity and carryover. Patients' results from the RA-1000 correlated well with those from the Beckman ASTRA-8. In a study on potential interferences, we found that high concentrations of salicylate and bromide significantly affected measurements of carbon dioxide and chloride, respectively. The RA-1000 requires only 30 microL of sample for all four tests and it offers a high throughout (30 specimens analyzed for the four tests in 25 min). This precise, easy-to-use, random-access analyzer requires minimal maintenance.
Procedure for characterization of creatine kinase variants on agarose electrophoretograms.
Authors: Abbott L B , Van Lente F
Published: January 2020

Electrophoretograms of atypical creatine kinase (EC 2.7.3.2, CK) isoenzymes may be difficult to interpret. In addition, their presence may cause discrepancies between results by immunological determinations for mass or activity and those by electrophoresis. Here we outline procedures to use in characterizing apparently atypical CK isoenzymes observed after electrophoresis, as illustrated by a CK isoenzyme variant that migrated as CK-MB.
125I radioimmunoassay for 17 alpha-hydroxyprogesterone in plasma, for diagnosing and managing congenital adrenal hyperplasia.
Authors: Yeo K H , Whitau W J
Published: January 2020

This rapid, inexpensive, and sensitive radioimmunoassay (RIA) for plasma 17 alpha-hydroxyprogesterone involves radioiodination. A single extraction with toluene/hexane removes an average 93% of the hormone from 0.1 mL of plasma. The extract is evaporated and the hormone is estimated by a simple, precise, and accurate 125I RIA involving a specific rabbit antiserum. A suspension of dextran-coated charcoal is used to separate free and bound steroid. Inter- and intra-assay CVs were less than 15 and less than 10%, respectively, and the sensitivity was 3 pg per assay tube. The regression equation for data on 17 alpha-hydroxyprogesterone added to steroid-free plasmas was y = 0.94x + 2.2 (r = 0.99). However, the turnaround time is only one-half to one-tenth that for most 3H RIA (3 h vs 6 to 30 h). The ranges of values found for plasma from normal subjects, treated and untreated patients with congenital adrenal hyperplasia, and infants with newly detected congenital adrenal hyperplasia were, respectively, 1 to 11, 0 to 20, 30 to 620, and 270 to 4900 nmol/L.
Derivatives of cyclosporin compatible with antibody-based assays: I. The generation of [125I]-labeled cyclosporin.
Authors: Mahoney W C , Orf J W
Published: January 2020

The immunosuppressive drug cyclosporin A, has been successfully iodinated to a specific activity of 300 Ci per gram. 125I-labeled cyclosporin and [3H]cyclosporin are nearly equivalent as tracers in a radioimmunoassay in producing standard lines (suppression by unlabeled cyclosporin) and in assigning values to clinical samples. In addition, the [125I]-labeled cyclosporin has greater than twice the sensitivity, and it is stable to long-term storage. Use of a [125I]-labeled cyclosporin tracer is more convenient, more reproducible, more precise, and easier than the tritiated-cyclosporin alternative in radioimmunoassay of this compound.
Probably myocardial infarction in a 14-year-old boy.
Authors: Feld R D
Published: January 2020

After vigorous physical activity a 14-year-old boy from a family with a strong history of cardiac disease developed symptoms consistent with a myocardial infarction, a relatively rare event in this age group. Isoenzyme patterns of creatine kinase and lactate dehydrogenase were consistent with the occurrence of an infarction.
Ion-selective electrodes for sodium and potassium: a new problem of what is measured and what should be reported.
Authors: Maas A H , Siggaard-Andersen O , Weisberg H F , Zijlstra W G
Published: January 2020

For clinical purposes the activities of Na+ and K+ obtained with ion-selective electrodes in undiluted whole blood or serum should be multiplied by an appropriate factor to obtain the same values as the substance concentrations obtained by flame photometry. The factor is primarily dependent on the mass concentration of water in normal plasma divided by the molal activity coefficient of Na+ (or K+) of normal plasma. We discuss the value of the molal activity coefficient of Na+ obtained by theoretical calculations and by direct measurement. The discrepancies between theory and measurement (gamma Na+ of 0.747 and 0.73, respectively) may be due to some binding of Na+ (protein binding or ion pair formation), a small and variable residual liquid-junction potential, or certainty about the appropriate value for the ionic strength of normal plasma (0.16 mol/kg or somewhat higher).
Hereditary dysfibrinogenemia.
Authors: Bithell T C
Published: January 2020

Inherited qualitative abnormalities of fibrinogen have been documented in more than 100 families. These dysfibrinogenemias usually are clinically silent, but in some cases are associated with bleeding, thrombosis, or defective wound healing. Abnormalities of the fibrinogen molecule may impair any of the major steps involved in the conversion of fibrinogen into stabilized fibrin; i.e., cleavage of the fibrinopeptides by thrombin, polymerization, and cross-linking of fibrin. Biochemical studies of several abnormal fibrinogens have demonstrated that the functional defects are the result of single amino acid substitutions. The hereditary dysfibrinogenemias are the first coagulation disorder in which the pathophysiology has been elucidated on a molecular level. Studies of these "experiments of nature" have important implications in such diverse processes as wound healing and thrombosis.
Simultaneous determination of trace elements in lavage fluids from human bronchial alveoli by energy dispersive x-ray fluorescence. 1: Technique and determination of the normal reference interval.
Authors: Maier E A , Rastegar F , Heimburger R , Ruch C , Pelletier A , Maier A , Leroy M J
Published: January 2020

We have applied energy dispersive x-ray fluorescence to the analysis of broncho-alveolar lavage fluids, to determine the concentration of several elements simultaneously with manipulation of the sample minimized. A 20-microL sample of a homogenized lavage fluid, to which two internal standards (containing Y and V) have been added, is deposited on a polypropylene film, 4 microns in thickness, and analyzed by x-ray fluorescence. We report the analytical procedure, the detection limits, and the reproducibility. The normal concentration range for trace elements is reported for a selected population. The simplicity and rapidity of the method, and the possibility of automating measurements, make this procedure suitable for screening large numbers of lavage specimens and to establish the diagnosis of some pneumoconioses.
Nickel concentrations in serum of patients with acute myocardial infarction or unstable angina pectoris.
Authors: Leach C N , Linden J V , Hopfer S M , Crisostomo M C , Sunderman F W
Published: January 2020

Nickel was measured, by electrothermal atomic absorption spectrophotometry, in sera from (a) 30 healthy adults, (b) 54 patients with acute myocardial infarction, (c) 33 patients with unstable angina pectoris without infarction, and (d) five patients with coronary atherosclerosis who developed cardiac ischemia during treadmill exercise. Mean (and SD) concentrations in Group a were 0.3 (0.3) microgram/L (range less than 0.05-1.1 microgram/L). Within 72 h after hospital admission, hypernickelemia (Ni greater than or equal to 1.2 microgram/L) was found in 41 patients of group b (76%) and in 16 patients of group c (48%). Hypernickelemia was found before and after exercise in one patient of Group d (20%). Peak values averaged 3.0 micrograms/L (range 0.4-21 micrograms/L) in Group b, 1.5 microgram/L (range less than 0.05-3.3 micrograms/L) in Group c. In Group b, the mean time interval between the peak values for creatine kinase activity and for nickel was 18 h. Serum nickel concentrations were unrelated to age, sex, time of day, cigarette smoking, medications, clinical complications, or outcome. Mechanisms and sources of release of nickel into the serum of patients with acute myocardial infarction or unstable angina pectoris are conjectural, but hypernickelemia may be related to the pathogenesis of ischemic myocardial injury.
An experimental study of the effect of zinc on the activity of angiotensin converting enzyme in serum.
Authors: Reeves P G , O'Dell B L
Published: January 2020

The activity in serum of zinc-dependent angiotensin converting enzyme (ACE), is measured to aid in diagnosis and monitor treatment of certain diseases. This report shows the effect of dietary zinc deprivation on ACE activity in the serum of rats. The mean (and SE) of the zinc concentration (mumol/L) in serum was 3.5 (0.3) in rats deprived of dietary zinc for four days, 16.3 (0.2) in control rats, and 19.8 (0.9) in rats deprived of zinc for four days, then repleted with zinc for 12 h. The respective mean (and SE) of ACE activities (nmol/mL per min) in serum were 390 (15), 543 (13), and 545 (20). Serum ACE activity was restored also by adding zinc to the assay mixture in vitro. The Vmax for ACE was 1.4 times greater when serum was diluted 40-fold as compared with twofold dilution. There was a small effect on the Km for the substrate, but the Km for zinc was decreased by 22-fold when serum was diluted 40-fold. The Vmax under these conditions was decreased by only 9%.
Determination of cyanide in whole blood, erythrocytes, and plasma.
Authors: Lundquist P , Rosling H , Sörbo B
Published: January 2020

We describe a method for determination of cyanide in whole blood, erythrocytes, and plasma after stabilization of cyanide by addition of silver ions. The cyanide is then transferred from the acidified sample, by aeration, into sodium hydroxide and quantified by the König reaction, with sodium hypochlorite as the chlorinating agent. A rapid loss of measurable cyanide found when cyanide was added to plasma in the absence of silver ions was attributed to a reaction with serum albumin. Cyanide added to whole blood was bound to a saturable component in erythrocytes, which we identified as methemoglobin.
Determination of theophylline in serum with the Seralyzer Aris reagent strip test evaluated.
Authors: Lindberg R , Ivaska K , Irjala K , Vanto T
Published: January 2020

We evaluated a new Seralyzer Aris reagent strip test (Ames Div., Miles Labs.) for the determination of theophylline in human serum. The method is based on the monoclonal enzyme immunoassay with dry reagent chemistry. The analysis is rapid and simple to perform: results are available only 5-10 min after receipt of the sample. Intra-assay precision (CV) was 2.2-3.3% (n = 15) for theophylline concentrations of 5-25 mg/L; interassay CV was 5.9% (n = 19) at 15 mg/L. The results (y) agreed well with those by liquid chromatography (x): r = 0.949 (p less than 0.001), and y = 0.967x + 0.214. We conclude the method is useful for rapid evaluation of theophylline concentrations in asthmatic patients.
A scheme for determining the correct activity of the kinetic angiotensin-converting enzyme.
Authors: Buttery J E , Chamberlain B R
Published: January 2020

The absorbance difference measured when angiotensin-converting enzyme (EC 3.4.15.1) hydrolyzes the substrate N-[3-(2-furyl)acryloyl]-L-phenylalanylglycylglycine is the basis for measuring its activity. We show this difference to be instrument dependent, and describe a method for deriving it that is applicable to manual or automated procedures.
Better laboratory evaluations of instruments and kits are required.
Authors: Fraser C G , Singer R
Published: January 2020

There are many published protocols for evaluating instruments and reagent-kit sets, and a plethora of papers describe the results of evaluations performed. It might be assumed, therefore, that this facet of clinical chemistry has no problems, but examination of evaluations of instruments published in the first half of 1984 shows flaws in some aspects of experimental design and execution, statistical analysis, and interpretation of data. We believe that clinical chemists, manufacturers and suppliers, and journal editors and referees can all contribute to improvement of this situation, and we suggest some guidelines for each of these groups.
Liquid chromatography of serotonin and adenine nucleotides in blood platelets, illustrated by evaluation of functional integrity of platelet preparations.
Authors: Ingebretsen O C , Bakken A M , Farstad M
Published: January 2020

In this relatively rapid liquid-chromatographic procedure the endogenous serotonin in blood platelets is quantified by fluorometry. We used this experimental procedure to estimate the platelet-release reaction as an indicator of the functional integrity of stored platelets. a decrease in the platelet-release reaction more sensitively indicates platelet changes during storage than do alterations in total ATP concentration or adenylate energy charge. The described method for quantifying serotonin is convenient enough for routine use in blood banks.
Urinary glycolate measured by use of (S)-2-hydroxy-acid oxidase.
Authors: Bais R , Nairn J M , Potezny N , Rofe A M , Conyers R A , Bär A
Published: January 2020

Glycolate can be determined in urine by using (S)-2-hydroxy-acid oxidase (EC 1.1.3.15; formerly called "glycolate oxidase"), either immobilized in a continuous-flow system or in a semiautomated procedure for the centrifugal analyzer. In the presence of peroxidase (EC 1.11.1.7), the hydrogen peroxide formed from glycolate is detected by use of a peroxide indicator reaction. Before the analysis, urine must be treated with charcoal to remove reducing substances such as ascorbic acid, which interfere with the assay by decreasing the color of the indicator reaction. Lactate also interferes with the determination of glycolate because it also is a substrate for this oxidase; thus a correction has to be made for the lactate content of urine. The system with (S)-2-hydroxy-acid oxidase immobilized to the inner surface of nylon tubing is accurate, precise, and sensitive but unsuitable for routine use because, even immobilized, the oxidase is unstable and can only be used for 12 days. We have used the semiautomated assay routinely: it has a mean analytical recovery of 96% (SD 4.2%), a within-batch CV less than 2%, and a between-batch CV less than 5%. The normal reference interval for urinary excretion of glycolate so measured is 0.13 to 1.31 mmol per day (n = 55).
Measurement of plasma fructosamine evaluated for monitoring diabetes.
Authors: Lim Y S , Staley M J
Published: January 2020

The mean fructosamine concentration in plasma of diabetics (n = 200) differed significantly (p less than 0.001) from those of a hospitalized nondiabetic population (n = 163)--the latter mean being essentially the same for ambulatory subjects (n = 145), expectant mothers (n = 58), and patients with renal failure (n = 31), regardless of sex. In the diabetic group, values for plasma fructosamine correlated with those for glycated hemoglobin (r = 0.767, p less than 0.01) and glycated protein (r = 0.817, p less than 0.01). Values for plasma fructosamine were stable from day to day in patients with controlled blood glucose, but fluctuated in certain patients receiving only parenteral nutrition, even when their concentrations of glucose were normal and stable. We conclude that measurement of plasma fructosamine is suitable for assessing intermediate-term control of blood glucose when the turnover of plasma proteins is normal.
Vitamin B12 assays compared by use of patients' sera with low vitamin B12 content.
Authors: Sheridan B L , Pearce L C
Published: January 2020

We compared four radioisotope dilution (RD) methods and a microbiological assay for measuring concentrations of vitamin B12 in a selected panel of serum samples from patients known to be deficient in the vitamin. Low (less than 100 ng/L) and borderline (100-180 ng/L) results were similar between methods, but use of the manufacturers' recommended ranges for borderline results would have changed the diagnostic classifications for 22 of 38 samples. Results of all the RD methods inter-correlated well, but less so with the microbiological assay. Borderline, nondiagnostic results were common to all methods, and no apparent advantage was gained from using the microbiological assay.
Enzyme immunoassay of free thyroxin in dried blood samples on filter paper.
Authors: Hata N , Miyai K , Ito M , Endo Y , Iijimi Y , Mizuta H , Amino N , Nose O , Harada T
Published: January 2020

We describe a double-antibody enzyme immunoassay for determination of free thyroxin (FT4) in dried blood samples on filter paper, with use of a T4-beta-D-galactosidase complex. The measurable range of FT4 concentration in two 3-mm blood discs, each of which contained about 2.7 microL of blood, was 1.9 to 93 ng/L, as determined by comparison with concentrations of FT4 in known serum standards. FT4 in blood samples dried on filter paper was stable for at least four weeks when kept dry at -20 degrees C, room temperature, or 37 degrees C. The mean coefficients of variation were 7.6% (within assay) and 6.4% (between assays). Results for FT4 by this method correlated well with those for serum determined by radioimmunoassay (r = 0.98). The proposed method can be used to differentiate persons with hyper- and hypothyroidism from normal subjects and those with abnormal concentrations of thyroxin-binding globulin. The procedure seems suited for screening studies.
Variations in total phospholipids and high-density lipoprotein phospholipids in plasma from a general population: reference intervals and influence of xenobiotics.
Authors: Cuvelier I , Steinmetz J , Mikstacki T , Siest G
Published: January 2020

The influence of different factors on variations in the concentrations of total phospholipids and high-density-lipoprotein (HDL) phospholipids in plasma was studied in a presumably healthy population of 2000 subjects, four to 70 years old. Age is the major factor associated with variation of total phospholipids. In females, this is due in part to age-related changes in hormonal status. The use of oral contraceptives affects only HDL phospholipid values. Use of tobacco does not influence plasma phospholipid values, but alcohol consumption increases values for both total and HDL phospholipids. We propose reference intervals for total plasma phospholipids and HDL phospholipids, adjusted for age and sex. Screening for lipid status can now include determinations of phospholipids as well as cholesterol and triglycerides.
Liquid-chromatographic determination of vanillylmandelic acid in urine.
Authors: Anderson G M , Feibel F C , Cohen D J
Published: January 2020

Urinary vanillylmandelic acid (VMA) was determined by "high-performance" liquid chromatography with fluorometric (LC-F) and amperometric (LC-EC) detection. Urine samples were first purified on a small, open-bed, reversed-phase preparatory column. VMA and the internal standard (iso-VMA) were then separated by reversed-phase ion-pair liquid chromatography. Analytical recovery of VMA was high (98.3%, SD 3.3%, n = 8), and concentrations measured by LC-F and LC-EC were in excellent agreement (r = 0.996). The LC-F chromatograms of urine samples had fewer late peaks; however, detection limits were lower (15 vs 120 micrograms/L) for the LC-EC method. Typical concentrations of 1-10 mg/L in urine can be measured easily with either method.
Liquid-chromatographic determination of total hydroxyproline in urine.
Authors: Turpeinen U , Pomoell U M
Published: January 2020

A reversed-phase "high-performance" liquid chromatographic assay for total hydroxyproline in urine is described. The urine samples are hydrolyzed overnight with acid, evaporated, solubilized, and derivatized with 4-dimethylaminoazobenzene-4'-sulfonyl chloride. The derivatives are chromatographed with a solvent gradient consisting of sodium acetate buffer and acetonitrile, the effluent being monitored at 436 nm. The useful lower limit of sensitivity for quantification is 13 pmol of hydroxyproline per 5-microL injection, corresponding to 33 mumol/L of urine. Either glutamine or cysteic acid is satisfactory as the internal standard. Peak heights and the amounts of hydroxyproline applied to the column are linearly related from 13.3 to 266 pmol. Mean analytical recovery was 83%. For four different concentrations the mean within-assay CV was 9.0% and the between-assay CV 12%. The normal reference interval found for 31 healthy adults was 31 to 177 mumol/24 h per square meter of body surface. We compared results for 10 samples from patients.
Increased serum sulfate in pregnancy: relationship to gestational age.
Authors: Cole D E , Baldwin L S , Stirk L J
Published: January 2020

Controlled-flow ion chromatography has significantly improved the precision with which inorganic sulfate (SO4) can be measured in serum. In this study, we have shown that serum SO4 is increased in pregnancy. The increase appears to follow gestational age, resulting in a 39% higher value by the middle of the third trimester. We suggest that this increase is a natural physiological process, which enhances SO4 availability to the growing fetus and placenta.
Immunoturbidimetric assay for alpha 1-acid glycoprotein in serum with a centrifugal analyzer.
Authors: Marteau F , Biou D , Thérond P , Demelier J F
Published: January 2020

This is a simple immunoturbidimetric method for the rapid assessment of alpha 1-acid glycoprotein in serum with the "Flexigem" microcentrifugal analyzer. Results obtained for a pediatric population (n = 95) agree with those by radial immunodiffusion (x) and yield the regression equation: y = 1.4x + 0.017 (r = 0.98). Results are printed automatically after a procedure that takes only about 15 min. Within-run precision (CV) ranged from 2.2 to 2.5% for concentrations of 0.35 to 2.4 g/L. Day-to-day CV was 4.25% for an average concentration of 0.9 g/L.
Possible role of lysolecithins and nonesterified fatty acids in the pathogenesis of Reye's syndrome, sudden infant death syndrome, acute pancreatitis, and diabetic ketoacidosis.
Authors: Shaw W
Published: January 2020

In vitro denaturation and (or) alteration of protein function by detergents have been extensively documented. I suggest that similar biochemical and clinical features of Reye's syndrome, sudden infant death syndrome, acute pancreatitis, and diabetic ketoacidosis may be explained as sequelae of the toxic detergent effects of nonesterified fatty acids and lysolecithins. These diseases may be provoked by a drug-induced diminution of the detergent-buffering capacity of blood or tissue proteins; by excess detergents produced in vivo, consequent to lipase activity induced by viral infection or metabolic disease; or by some combination of these factors.
Interlaboratory evaluation of salicylate interference in colorimetric acetaminophen methods and its clinical significance.
Authors: Jenny R W
Published: January 2020

Serum specimens with concentrations simulating an overdose of salicylate and acetaminophen were submitted to laboratories participating in an external quality-control program, to evaluate both the magnitude of salicylate interference in colorimetric acetaminophen methods and the clinical significance of the interference. The apparent acetaminophen concentration determined by nitration methods was increased by about 0.70 mg/L per milligram of salicylate per deciliter. Of those laboratories using nitration procedures, 25% do not routinely correct for salicylate and 66% use the (incorrect) correction factor provided by a kit manufacturer. Laboratory data, as they would have been reported to physicians, were used to estimate the acetaminophen half-life and were also applied to a nomogram used to assess the probability of hepatotoxicity. Interference by salicylate in the simulated overdose of 10 g (total dose) of each drug falsely indicated impending hepatic necrosis unless the appropriate correction factor was used. Laboratories using nitration procedures should screen samples submitted for acetaminophen assay for the presence of salicylate and, if present, either use a method specific for acetaminophen or utilize a correction factor determined in-house.
"Total" acidic metabolites of catecholamines in urine as determined by hydrolysis with hydriodic acid and liquid chromatography: application to patients with neuroblastoma and melanoma.
Authors: Ito S , Kato T , Maruta K , Jimbow K , Fujita K
Published: January 2020

We describe a method for determining the "total" excretion of acidic metabolites of catecholamines by measuring 3,4-dihydroxyphenylacetic acid (DOPAC) formed by hydriodic acid hydrolysis of 4-hydroxy-3-methoxyphenylacetic acid (HVA), 4-hydroxy-3-methoxymandelic acid (VMA), and their conjugates. The DOPAC in the diluted hydrolysate is measured directly by liquid chromatography with electrochemical detection. Normal values, expressed in relation to excretion of creatinine, vary as a function of age. For healthy subjects, the mean DOPAC value after hydrolysis was 1.15 times that for unconjugated HVA, VMA, and DOPAC combined. Preliminary results for patients with neuroblastoma and melanoma indicate the potential usefulness of the method for diagnosis and prognosis of patients with neural crest tumors that produce dopa or catecholamines.
Determination of propranolol in plasma by radial compression liquid chromatography with fluorometric detection.
Authors: el-Yazigi A , Martin C R
Published: January 2020

This radial compression liquid-chromatographic assay for propranolol in plasma is rapid, reproducible, and suitable for use in routine monitoring. A 10-micron particle, 8 mm X 10 cm CN cartridge is used in conjunction with a radial compression separation system. The mobile phase is monobasic sodium phosphate (pH 3) solution/methanol/acetonitrile (760/84/156 by vol), the flow rate 6 mL/min. Propranolol was detected by use of a spectrofluorometer equipped with a 20-microL flow-through cell, at excitation and emission wavelengths of 250 and 336 nm. The retention times for propranolol and metoprolol (the internal standard) are 3.13 and 1.42 min, respectively. A one-step extraction with chloroform yields "clean" chromatograms, with greater than 90% of the drug being analytically accounted for. Under these conditions, results are precise and accurate. Currently we are using this method to monitor propranolol in hypertensive neonates. Data on changes in the concentrations of propranolol in plasma with time are presented for one such patient.
A cause of discrepancy between values for urinary protein as assayed by the Coomassie Brilliant Blue G-250 method and the sulfosalicylic acid method.
Authors: Shiba K S , Kanamori K , Harada T , Nakao M , Nakajima K , Kodaira T , Nakagawa H
Published: January 2020

In simultaneous assays of urinary proteins by the Coomassie Brilliant Blue G-250 (CBB) and the sulfosalicylic acid (SSA) methods, we noticed that about 18% of samples showed about twice higher protein values by the former method than by the latter. Some urinary proteins are soluble in SSA and react with CBB. Examinations with sodium dodecyl sulfate/polyacrylamide gel electrophoresis showed that these proteins migrated in 13 protein bands having relative molecular masses ranging from 15 000 to 230 000. The protein corresponding to the most intensely stained band in urine samples from the patients studied (with malignant tumors, renal disorders, etc.) had an Mr of 45 000; that in the pattern for healthy subjects had an Mr of 94 000. The former was identified as alpha 1-acid glycoprotein, the latter as Tamm-Horsfall mucoprotein.
Guidelines for immunoassay data processing.
Authors: Dudley R A , Edwards P , Ekins R P , Finney D J , McKenzie I G , Raab G M , Rodbard D , Rodgers R P
Published: January 2020

These guidelines outline the minimum requirements for a data-processing package to be used in the immunoassay laboratory. They include recommendations on hardware, software, and program design. We outline the statistical analyses that should be performed to obtain the analyte concentrations of unknown specimens and to ensure adequate monitoring of within- and between-assay errors of measurement.
Assessment of corpus luteum function by direct radioimmunoassay for progesterone in blood spotted on filter paper.
Authors: Petsos P , Ratcliffe W A , Anderson D C
Published: January 2020

In this specific, direct RIA for progesterone in capillary blood dried on filter paper, progesterone is eluted, with phosphate buffer containing bovine serum albumin, from 5.9 microL of blood dried on 5.0-mm (diameter) discs of filter paper. The eluate is assayed, with 125I-labeled progesterone-11 alpha-glucuronyl-tyramine as tracer, with separation by a double-antibody solid-phase technique. The sensitivity of the assay is 4.7 pg per tube, corresponding to 2.5 nmol per liter of blood. Within- and between-batch CVs averaged 7.0 and 9.2%, respectively, over the working range of the assay (4.5-64 nmol/L). Concentrations of progesterone in blood spots (y) correlated well with those in serum (x) as measured by an established direct RIA (Clin Chem 28:1314, 1982): y = 0.430x - 2.44 (r = 0.972, n = 104). Progesterone is stable in the blood spots for at least 15 weeks at 25 degrees C. The convenience of multiple sampling of blood by finger prick and the simplicity of the assay make this approach useful in investigating serial progesterone concentrations in outpatients.
Epitope-selective, monoclonal-antibody-based immunoradiometric assays of predictable specificity for differential measurement of choriogonadotropin and its subunits.
Authors: Schwarz S , Berger P , Wick G
Published: January 2020

Knowing the epitope specificities of our monoclonal antibodies (MCA) to human choriogonadotropin (hCG), we could design three classes of two-site immunoradiometric assays (IRMA): a combination of two MCA recognizing two separate alpha-epitopes (alpha-MCA) provides a system (i.e., an alpha-IRMA) that measures holo-hCG plus free alpha-subunits plus follitropin, lutropin, and thyrotropin, whereas a beta-IRMA, consisting of two beta-MCA, quantifies holo-hCG plus free beta-subunits. The amount of either of the two subunits can be calculated by subtracting the amount of holo-hCG determined in parallel in a holo-hCG-IRMA. In the latter, one of the alpha- or beta-MCA may be either cross-combined or, preferably, paired with an MCA specific for a conformational epitope. These analytical specificities, predicted from our previously established epitope map of hCG, could be experimentally verified. With these IRMAS we could demonstrate that in certain choriocarcinoma cell lines the earliest and quantitatively predominant tumor marker is the free alpha-subunit. Similar results showing an unbalanced secretion of hCG and its subunits were obtained for patients with related tumors. These findings challenge the present diagnostic practice of relying solely on "beta-hCG" radioimmunoassays and at the same time offer a novel analytical strategy.
An evaluation of a fluorescence polarization immunoassay of thyroxin and thyroxin-uptake.
Authors: Symons R G , Vining R F
Published: January 2020

We evaluated the fluorescence polarization immunoassay for total thyroxin (T4) and thyroxin-uptake (T-U) in the Abbott "TDx" Analyzer. Between-assay precision was good when we did once-fortnightly calibration and assayed samples in singleton. Measured T4 concentration was decreased in hemolyzed samples with obvious red coloration and undetectable in severely hemolyzed samples. The T-U assay was unaffected by hemolysis. Unlike the triiodothyronine-uptake methods, the T-U assay utilizes labeled T4 and measures a variable related to serum thyroxin-binding capacity rather than the concentration of unoccupied binding sites in serum. The T4 and T-U values of 422 samples correlated highly with a T4 (in-house) radioimmunoassay and a commercial assay for thyroxin-binding globulin, respectively. The free thyroxin ratio (ratio of T4 to T-U, FTI) correlated highly with free T4 concentration as measured by T4-analog-tracer radioimmunoassay (fT4 analog RIA). FTI and fT4 values were discordant in late pregnancy (normal FTI and low fT4) and euthyroid sick patients (above-normal or normal FTI and low fT4), suggesting that the FTI gives fewer misleading results in these patients.
Preparation and use of serum-based material as control and calibrator in evaluating ion-selective electrodes for calcium.
Authors: Toffaletti J , Lee K M
Published: January 2020

We analyzed 93 sera and 81 whole-blood samples, plus several aqueous materials and one serum-based material as controls, with two analyzers for ionized calcium (Radiometer ICA 1 and AVL 980). On 16 days, comparisons of results for patients' samples were good, with nearly all between-instrument differences (delta Ca2+) of samples being within 0.04 mmol/L. On 12 days, comparisons were mediocre, with the delta Ca2+ of samples usually 0.04 to 0.10 mmol/L. The control material that most consistently indicated the direction and magnitude of the delta Ca2+ of patients' samples was the serum-based control. Adjustment of results for samples from patients, based on the delta Ca2+ of the serum control on the same day, substantially improved comparisons between instruments. Our findings suggest that the use of a serum-based calibrator provides the most valid comparison between different calcium-ion analyzers and reliably indicates when electrode replacement is needed.
Adequacy of interlaboratory precision criteria in measuring intralaboratory performance.
Authors: Ehrmeyer S S , Laessig R H
Published: January 2020

We compared the predictive value of the various criteria used for grading pH and blood-gas measurements in interlaboratory proficiency testing programs with performance as determined from actual intralaboratory quality-assurance data. The evaluation criteria were the two-standard-deviation interval (2 SDI) proposed by the College of American Pathologists (CAP); CAP's proposed fixed criteria; and the fixed criteria of the American Thoracic Society (ATS). These were compared with 95% confidence limits derived from the individual laboratories' actual intralaboratory data. We found that the CAP's most-stringent criterion (2 SDI) overestimated the number of outliers (unacceptable results) for PCO2 and PO2, whereas the proposed fixed limits underestimated them. For pH and PCO2 the ATS's limits, which are more stringent, more closely match the individual laboratory's actual performance as measured by conventional (mean +/- 2 SD) intralaboratory quality-assurance practices.