bluuebunny/crossref_metadata_2025 huggingface dataset:
Simultaneous measurement of two major prostacyclin metabolites in urine
Authors: Zureick S , Nadler J , Yamamoto J , Horton R
Published: January 2020

We describe a combined HPLC-RIA technique to measure both major metabolites of prostacyclin (PGI2): 6-keto PGF1 alpha and 2,3-dinor-6 keto PGF1 alpha. The measurement of the former, which originates from renal blood vessels, and the latter, from systemic vessels and the liver, may provide a better overall evaluation of production than measurement of one metabolite. An aliquot of acidified urine with added 3H-labeled metabolites is adsorbed and then eluted from a C18 Bond-Elut column. The sample is then passed through an HPLC system by use of an isocratic solvent combination that separates the two metabolites from known prostaglandins. The purified metabolites are then quantified by RIA. Using a logit-log10 transform, one can measure between 12 and 250 pg of either metabolite, with high accuracy and precision (CVs of 12% for a low concentration and 7% for a high concentration). Reference values for apparently healthy subjects were, respectively, 107 (SD 45) and 171 (SD 69) ng/g creatinine for 6-keto PGF1 alpha and the dinor metabolite in men (n = 18) and 45 (SD 22) and 141 (SD 28) ng/g creatinine, respectively, in women (n = 15). Indomethacin in standard doses reduced both metabolite values by 50%. Intravenous administration of angiotensin II (5 ng/kg of body wt per minute) did not alter excretion rates, but equipressor doses of norepinephrine (0.1 microgram/kg per minute) increased the production of both metabolites (6-keto greater than dinor).
Determination of beta-carotene and its cis isomers in serum
Authors: Rushin W G , Catignani G L , Schwartz S J
Published: January 2020

All-trans-beta-carotene was resolved from its cis isomers in human serum by reversed-phase "high-performance" liquid chromatography. Absorption spectra of the cis peak suggested that 13-cis-beta-carotene was the predominant cis isomer. Analyses and recovery studies of fresh and stored sera eliminated the possibility that isomerization had occurred in samples during handling or storage. The average analytical recovery was 101.9% for standards of the all-trans-, 9-cis-, and 13-cis-beta-carotene compounds in pooled serum samples. We also demonstrated that cis isomers had not formed after the blood was drawn and that cis isomers of beta-carotene are present at significant concentrations in the human circulation.
Excess serum osmolality gap after ingestion of methanol
Authors: Meatherall R , Krahn J
Published: January 2020

Two cases of methanol ingestion and one case of combined methanol and ethylene glycol ingestion are presented to illustrate the large differences that exist between the serum osmolality gap and the measured methanol (plus ethylene glycol) concentration(s) before treatment of these poisonings. After treatment with intravenous ethanol and hemodialysis was initiated, the differences disappeared in all three cases. We speculate that one or more metabolites with osmotic activity are formed in cases of methanol intoxication where no ethanol has also been consumed. The possible identity of these compounds is discussed.
Clinical laboratory regulation under the Clinical Laboratory Improvement Amendments of 1988: can it be done?
Authors: Peddecord K M , Hammond H C
Published: January 2020

This report examines logical but not yet widely recognized ramifications of the Clinical Laboratory Improvement Amendments of 1988 (CLIA'88), federal legislation that will require certification of all laboratories examining human specimens. Examination of the CLIA'88 committee reports and committee hearings suggest that more than the conventional approach to laboratory standards will be needed to meet the public's expectations as articulated by our elected representatives. The conventional approach to clinical testing standards seeks to assure quality by regulating the laboratory analytical process. However, little empirical evidence is available to support or refute this model, which has been used during the past 25 years. One alternative paradigm for laboratory standards is an approach that examines the total laboratory testing process, including the selection, ordering, and interpretation of the test as well as the laboratory analysis per se. The history of controversy over laboratory standards--especially personnel standards, the glacial federal regulatory rulemaking process, public expectations of fail-safe technology, among other factors--suggests the implementation of CLIA'88 will be a lengthy and vigorously debated contest. The risk of a test is seldom inherent in the test itself, but rather is a function of the context in which the test is being used to provide information for medical decision making. Our premise is that diagnostic tests must be examined in the context of the laboratory testing situation. We suggest that now is the appropriate time for laboratory professionals, practicing physicians, and the public to abandon conventional thinking regarding clinical laboratory standards. We believe that CLIA'88 reflects a shift in public expectations toward fail-safe laboratory testing and the need for additional government oversight in laboratory test quality. If these new expectations persist, CLIA'88 represents a potential landmark in the course of federal authority and the practice of medicine in the United States.
Direct chemiluminescence immunoassay of estradiol in saliva
Authors: De Boever J , Kohen F , Bouve J , Leyseele D , VandeKerckhove D
Published: January 2020

A sensitive and simple direct solid-phase chemiluminescence immunoassay is described for estradiol in saliva. In this assay, a second antibody is bound to the wells of microtiter plates. Either buffer with standards or saliva (100 microL) is incubated in these wells with monoclonal anti-estradiol antibody and with estradiol-isoluminol conjugate. Incubation time is 2 h. Chemiluminescence of the bound fraction is measured in a manually operated luminometer (Biocounter). The assay has a detection limit of 3.8 pmol/L; analytical recovery of added estradiol is 96.8% (SD 7.0%); within- and between-assay CVs range between 2.5% and 12.7%. Forty unknown saliva samples can be assayed and results calculated within 4.5 h. Results of a slightly modified procedure-with black microtiter plates and a prototype of an automated plate reader (Luminoskan)--compare well with those of the described method (r = 0.97). Because steroid-binding globulins have been found in saliva, the effect of displacing agents on the results of the direct chemiluminescence assay is described.
Concentrations of apoprotein CII, CIII, and E in total serum and in the apoprotein B-containing lipoproteins, determined by a new enzyme-linked immunosorbent assay
Authors: Alsayed N , Rebourcet R , Chapman J
Published: January 2020

We describe a new enzyme-linked immunosorbent assay (ELISA) that permits direct determination of apoprotein (apo) CII, CIII, and E in total serum as well as in apo B-containing lipoprotein particles. To validate this ELISA technique, we studied several aspects of the assay: its specificity, the influence of the conditions of conservation of plasma and of lipoprotein fractions, the effect of delipidation, and its reproducibility. We measured the concentrations of apo CII, CIII, and E in total serum and in apo B-containing lipoproteins from a pool of normal sera and in sera from 75 healthy subjects. After sequential ultracentrifugation, the content of apo CII, CIII, and E in the major lipoprotein fractions was also determined. Total serum or plasma could be stored at -20 or -50 degrees C for at least six weeks and the isolated lipoprotein fractions for as long as four weeks, which suggests a protective effect of total serum on lipoprotein particle structure. Advantages of this ELISA include (a) its specificity, sensitivity, and reliability; (b) better discrimination than determination of total serum apoprotein; (c) easier application and greater rapidity; and (d) the possibility of application to population screening.
Development and comparative evaluation of immunoblot assays for detecting autoantibodies to Scl 70 and Jo 1 antigens in serum
Authors: Fonong T , Evans S M , Homburger H A
Published: January 2020

We developed rapid 24-h immunoblot assays for detecting autoantibodies to Scl 70 and Jo 1 antigens in serum. In comparative studies, we evaluated the analytical sensitivity of the immunoblot assays and commercial immunodiffusion assays for anti-Scl 70 and anti-Jo 1 autoantibodies with the use of positive control sera, and compared the frequencies of positive and negative results in a group of 116 sera, including specimens from 34 healthy controls and 82 patients with various connective-tissue diseases. The immunoblot assays were greater than 100-fold more sensitive than immunodiffusion for detecting both autoantibodies. Despite greater analytical sensitivity, there were no false-positive results by the immunoblot assay for anti-Scl 70 or anti-Jo 1 autoantibodies in sera from either the controls or the patients. The diagnostic sensitivity of the immunoblot assay for anti-Scl 70 autoantibodies in patients with scleroderma was greater than that of the immunodiffusion assay, 70% vs 20%, and was equivalent in patients with polymyositis, 43%. We conclude that rapid immunoblot assays for anti-Scl 70 and anti-Jo 1 autoantibodies are superior to immunodiffusion assays for clinical use and are suitable for routine use in the clinical laboratory.
Automation of specific human gene detection
Authors: Mayrand P E , Hoff L B , McBride L J , Bridgham J A , Cathcart R , Corcoran K P , Golda G S , Keith D H , Lachenmeier E W , Madden D E
Published: January 2020

An instrument/chemistry system is described that automates a new chemical procedure functionally equivalent to Southern blotting. A fluorescence gel scanner that detects migrating DNA fragments in real-time analyzes the samples produced by a prototype liquid-handling instrument that automates a solution-phase hybridization/solid-phase capture chemistry for DNA analysis. The combination of this chemistry, the gel scanner, and robotic automation eliminates the tedium encountered in traditional manual methods for specific gene detection and reduces analysis time from days to hours. Restriction fragment lengths are measured with high precision by comparison with in-lane standards to minimize effects attributable to migration anomalies. The utility of this automated system is demonstrated by executing a clinical research application involving hybridization to a multi-copy repeat sequence on the Y chromosome and its detection.
Apolipoprotein E polymorphism determined by restriction enzyme analysis of DNA amplified by polymerase chain reaction: convenient alternative to phenotyping by isoelectric focusing
Authors: Kontula K , Aalto-Setälä K , Kuusi T , Hämäläinen L , Syvänen A C
Published: January 2020

Three common alleles determine six apolipoprotein E (apo E) phenotypes that are associated with variations in serum cholesterol in the population. This genetic variation results from single nucleotide alterations at two DNA loci encoding the amino acid residues 112 and 158 of apo E. We compared results of apo E phenotyping carried out by isoelectric focusing with those of apo E genotyping accomplished by direct DNA analysis. In the latter, the target DNA was amplified by the polymerase chain reaction (PCR) and subsequently analyzed by digestion with the restriction enzyme Hha I, followed by polyacrylamide gel electrophoresis of the cleavage products. With one exception, these two techniques yielded similar results from all 40 samples tested. In addition, a rare variant form of apo E (phenotype E1) was analyzed separately and incorrectly diagnosed as E2 by the Hha I digestion method; the anticipated mutation in the codon 127 was, however, confirmed by demonstration of a new Taq I restriction site in this variant gene. These data confirm that the common isoforms of apo E usually arise from genetic variation of the codons 112 and 158 and demonstrate the feasibility of the PCR technique in apo E genotyping.
Determination of reference values for serotonin concentration in platelets of healthy newborns, children, adults, and elderly subjects by HPLC with electrochemical detection
Authors: Flachaire E , Beney C , Berthier A , Salandre J , Quincy C , Renaud B
Published: January 2020

We adapted a high-performance liquid chromatographic method with electrochemical detection (Clin Chim Acta 1984;139:1-12) to the determination of platelet serotonin. We used this method to determine platelet serotonin reference values in a healthy population, measuring platelet serotonin concentration in the following subjects: 31 newborns (16 girls, 15 boys); 41 children (11 girls, 30 boys), ages 20 months to 15 years; 56 adults (26 women, 30 men), ages 20 to 58 years; and 20 elderly subjects (16 women, four men), ages 65 to 94 years. There was no significant difference in platelet serotonin concentration between sexes in each age group. However, significant changes (P less than 0.001) were observed between the newborns (mean +/- SD: 1.67 +/- 0.74 nmol/10(9) platelets) and the children (4.09 +/- 1.04) or the adults (3.81 +/- 0.87). Moreover, the platelet serotonin concentration in the elderly subjects (2.57 +/- 1.12) was significantly (P less than 0.001) lower than in the adults and children and significantly higher (P less than 0.01) than in the newborns. Such age-related differences must be taken into consideration when data from neurological or psychiatric patients and control subjects are compared.
Relationship between delta checks for selected chemistry tests
Authors: Lacher D A
Published: January 2020

The correlation between delta differences for 20 serum chemistry tests was calculated for 2400 samples from 288 patients. There were 12 pairs of chemistry tests for which correlation coefficients of the delta checks exceeded 0.25; aspartate aminotransferase and alanine aminotransferase had the highest Pearson correlation coefficient, 0.915. The highest negative, indirect, correlation was between the delta checks of bicarbonate and chloride (-0.219). The relationship between delta differences may be used as a quality-control technique to detect analytical errors.
A monoclonal-antibody-based enzyme-linked immunosorbent assay of lipoprotein(a)
Authors: Wong W L , Eaton D L , Berloui A , Fendly B , Hass P E
Published: January 2020

Lipoprotein(a) [Lp(a)] is a low-density lipoprotein (LDL)-like lipoprotein particle recently described as a risk factor for premature coronary heart disease, stroke, and atherosclerosis. Structurally, Lp(a) is similar to LDL in that it has comparable lipid composition and contains apolipoprotein B-100 (apo B-100). In addition, Lp(a) contains the glycoprotein apolipoprotein(a) [apo(a)], which is disulfide-linked to apo B-100. The recent awareness of a striking correlation between atherosclerosis and concentrations of Lp(a) in plasma prompted our development of an accurate quantitative assay for plasma Lp(a), a monoclonal-antibody-based enzyme-linked immunosorbent assay for Lp(a) that is shown to be sensitive, precise, and highly specific. The response to several isoforms of Lp(a) is linear, and as many as 80 samples can be quantified on one plate. This easily performed assay is suitable for use in the clinical laboratory and for screening large populations.
Activity of phospholipase A2 in plasma increases in uremia
Authors: Costello J , Franson R C , Landwehr K , Landwehr D M
Published: January 2020

We measured phospholipase A2 (PLA2; EC 3.1.1.4) activity in normal and uremic plasma, using [1-14C]oleate-labeled autoclaved Escherichia coli as substrate. Hydrolysis of bacterial phospholipid by crude plasma from both groups was optimal at pH 5.5, was specific for the 2-acyl position of phospholipids, and had an absolute requirement for calcium. Activity was greatest in the presence of added Ca2+, 5 mmol/L, but this increase was inhibited by several divalent cations (Mg2+, Zn2+, Cu2+, Ba2+, Co2+, Pb2+, Fe2+) and by Fe3+. PLA2 activity was also inhibited by heparin at acid and alkaline pH, normal plasma being more sensitive than uremic plasma to this inhibition. Enzyme activity in undiluted plasma was eightfold greater in uremic than in normal plasma. Dilution of plasma by two to fourfold increased the total activity of both normal and uremic plasma. However, the relative differences in total activity between the groups remained constant (eight- to 11-fold). The cause and consequences of the increased PLA2 activity in uremia remain to be established.
A unitized enzyme-labeled immunometric digoxin assay suitable for rapid testing
Authors: Sommer R G , Belchak T L , Bloczynski M L , Boguslawski S J , Clay D L , Corey P F , Foltz M M , Fredrickson R A , Halmo B L , Johnson R D
Published: January 2020

An enzyme-labeled immunometric assay has been developed for measuring digoxin concentrations in serum or plasma. Unitized, compartmentalized reagents are used with an automated sample-processing instrument. The enzyme activity of the processed sample, which is directly proportional to the digoxin concentration, is measured by using a reagent strip and the Ames Seralyzer reflectance photometer. The test takes less than 15 min, and digoxin concentrations are calculated from a two-point calibration line stored in the instrument. Within-run CVs for controls at four concentrations ranged from 2.3% to 3.8%; between-run CVs were from 1.5% to 2.6%. Results obtained with clinical serum samples correlated well (r greater than 0.96) with those obtained by fluorescent polarization immunoassay (Abbott TDx) and RIA (Clinical Assays and NML). This rapid and convenient method for monitoring digoxin concentrations in serum or plasma is particularly well suited for decentralized sites such as emergency rooms, urgent-care centers, and physicians' offices.
Automated measurement of two indicators of human selenium status, and applicability to population studies
Authors: McMaster D , Bell N , Anderson P , Love A H
Published: January 2020

We developed an automated system consisting of a centrifugal analyzer and an atomic absorption spectrophotometer with Zeeman background correction to produce a profile of the concentrations of selenium and the selenium-dependent enzyme glutathione peroxidase (EC 1.11.1.9) in serum and whole blood, for use in epidemiological surveys in Northern Ireland. No pretreatment of samples other than dilution is required, and at least 35 subjects can be screened within 24 h of venesection. For selenium in serum the between-run CV was 5.7% and 4.4% within run. For selenium in whole blood the CV was 5.9% between runs, 5.7% within run. The within-run CV for glutathione peroxidase was 0.92% for serum, 1.18% for blood; the between-run CV was 8.3% for serum. To illustrate the use of the system, we present the profile of selenium and glutathione peroxidase in serum and whole blood from 100 adults 25 to 64 years old.
Establishment of an external quality-assessment scheme for amino acid analyses: results from assays of samples distributed during two years
Authors: Rattenbury J M , Townsend J C
Published: January 2020

Ten different samples of lyophilized plasma and two of liquid urine were distributed during two years to 26 laboratories performing quantitative amino acid analyses in a scheme designed to provide external quality assessment. After each distribution, statistical summaries and performance scores based on delta standard deviations and percentage biases from the all-laboratory trimmed means were returned to participants, who also received annual performance summaries based on their accumulated results. Coefficients of variation calculated from returns across all the samples ranged from 13% for glycine to 65% for methionine. Automated ion-exchange amino acid analyzers with ninhydrin detection appeared to perform better than other methods, although there was no clearly superior method and no model of analyzer clearly outperformed the others. These exercises demonstrate that there is room for improvement in the performance of quantitative amino acid analyses and that individual expertise may be more important in maintaining good performance than the choice of method or analyzer.
Immunosuppressive activity of cyclosporine metabolites compared and characterized by mass spectroscopy and nuclear magnetic resonance
Authors: Copeland K R , Yatscoff R W , McKenna R M
Published: January 2020

Eight cyclosporine (CsA) metabolites were isolated from the urine of renal-transplant patients by high-pressure liquid chromatography. Structure and purity of the metabolites were assessed by fast atomic bombardment/mass spectroscopy, by proton nuclear magnetic resonance (NMR), and, when the quantity of metabolites permitted, by 13C-NMR. The immunosuppressive activities (I) of the metabolites were tested in three separate in vitro systems: primary and secondary mixed lymphocyte reactions as well as by a mitogen-stimulated system. The I, as measured by comparing the concentration of each metabolite required for 50% inhibition of incorporation of [3H] thymidine, varied among the assay systems, as did the ranking of I among the test systems. In general, the I of most metabolites in all assay systems were less than 10% of that for CsA. Metabolites with single modifications exhibited the greatest I; e.g., that of M-17 was congruent to 16% of that of CsA (potency ratio 0.16) in a secondary mixed lymphocyte reaction. The significance of these findings in relation to therapeutic monitoring of CsA is discussed.
Selection of medically useful quality-control procedures for individual tests done in a multitest analytical system
Authors: Koch D D , Oryall J J , Quam E F , Feldbruegge D H , Dowd D E , Barry P L , Westgard J O
Published: January 2020

Quality-control (QC) procedures (i.e., decision rules used, numbers of control measurements collected per run) have been selected for individual tests of a multitest analyzer, to see that clinical or "medical usefulness" requirements for quality are met. The approach for designing appropriate QC procedures includes the following steps: (a) defining requirements for quality in the form of the "total allowable analytical error" for each test, (b) determining the imprecision of each measurement procedure, (c) calculating the medically important systematic and random errors for each test, and (d) assessing the probabilities for error detection and false rejection for candidate control procedures. In applying this approach to the Hitachi 737 analyzer, a design objective of 90% (or greater) detection of systematic errors was met for most tests (sodium, potassium, glucose, urea nitrogen, creatinine, phosphorus, uric acid, cholesterol, total protein, total bilirubin, gamma-glutamyltransferase, alkaline phosphatase, aspartate aminotransferase, lactate dehydrogenase) by use of 3.5s control limits with two control measurements per run (N). For the remaining tests (albumin, chloride, total CO2, calcium), requirements for QC procedures were more stringent, and 2.5s limits (with N = 2) were selected.
Liver- and bone-derived isoenzymes of alkaline phosphatase in serum as determined by high-performance affinity chromatography
Authors: Anderson D J , Branum E L , O'Brien J F
Published: January 2020

To separate liver and bone alkaline phosphatase (ALP) isoenzymes in human serum, we used high-performance affinity chromatography (HPAC) on a column of wheat-germ lectin conjugated to 7-microns-diameter silica particles and an eluent containing N-acetyl-D-glucosamine (NAG). On-line spectrophotometric detection of ALP involved pumping diethanolamine-buffered p-nitrophenyl phosphate solution post-column. Bone and liver isoenzymes could be separated into two peaks with only 10% overlap when an exponential gradient was used. A linear-step gradient separated 80.9% of liver ALP and 91.6% of bone ALP in two distinct peaks. True bone and liver ALP peak areas for the linear-step gradient were determined by using correction factors, because each peak contained a co-eluted portion of the other ALP isoenzyme. The detection limit improved 10-fold over those of other techniques for ALP isoenzymes, owing to the relatively large sample that could be applied to the column. Correlation with a urea-inactivation procedure was reasonable for patients' serum samples (r = 0.98 and 0.79 for liver ALP and bone ALP, respectively).
Screening trace elements and electrolytes in serum by inductively-coupled plasma emission spectrometry
Authors: Melton L A , Tracy M L , Möller G
Published: January 2020

This rapid, accurate procedure for trace elements and electrolytes in serum requires little sample preparation: to 1 mL of serum a single reagent is added that contains trichloroacetic and hydrochloric acids for protein precipitation, hydroxylamine sulfate for iron reduction, and yttrium as the internal standard. After centrifugation, the supernates are directly analyzed for Na, K, Mg, Ca, Pi, Fe, Cu, and Zn by inductively-coupled plasma emission spectrometry. The CVs were respectively 7.9%, 8.4%, 8.6%, 10.0%, 9.0%, 9.4%, 9.0%, and 9.0% for five assays of National Institute of Standards and Technology Standard Reference Material (SRM) no. 1598, Bovine Serum. Analytical recoveries ranged from 92% to 107% for both SRM 1598 and commercial control serum.
Cholesterol screening: comparative evaluation of on-site and laboratory-based measurements
Authors: Bachorik P S , Rock R , Cloey T , Treciak E , Becker D , Sigmund W
Published: January 2020

We measured cholesterol in capillary blood samples from 9683 volunteers over a four-day on-site community screening program, using the "Reflotron" desk-top analyzer (Boehringer-Mannheim Diagnostics, Indianapolis, IN). We also measured cholesterol in venous blood samples from 3% of those screened (a) with the Reflotron at the screening sites, (b) in a qualified hospital clinical laboratory, and (c) in a Centers for Disease Control standardized lipoprotein research laboratory. The sensitivity (and specificity) of the Reflotron measurements, with use of the lipoprotein laboratory measurements as the point of reference, was 0.95 (0.73) in capillary blood samples and 0.88 (0.93) in venous blood samples, compared with 0.99 (0.87) in the hospital clinical laboratory. The Reflotron measurements correlated less well with the lipoprotein laboratory values in both venous blood (r = 0.91) and capillary blood (r = 0.89) samples than did the clinical laboratory values (r greater than 0.99). Furthermore, the capillary blood measurements averaged 7% higher than venous measurements when both kinds of samples were analyzed in the Reflotron.
Rapid purification of human DNA from whole blood for potential application in clinical chemistry laboratories
Authors: Adeli K , Ogbonna G
Published: January 2020

A simple, rapid method for isolating human DNA has been developed, which can be routinely used in clinical chemistry laboratories. The entire procedure takes less than 90 min, and as many as 12 blood samples can be handled in one cycle. One milliliter of EDTA-treated blood is lysed and centrifuged to yield a nuclear fraction. The nuclear pellet is treated with sodium dodecyl sulfate/urea and phenol/chloroform to remove contaminating proteins, then the crude DNA extract is purified by use of a Sephadex G-25 spin-column. Typical 260 nm/280 nm absorbance ratio (used to assess purity) and yield for DNA so purified were 1.84 and 24.5 micrograms/mL, respectively. Within- and between-day CVs for recovery of DNA from pooled blood were 8% and 11% respectively. Such DNA preparations were found quite suitable for digestion by a variety of restriction endonucleases and for restriction fragment length polymorphism analysis. We are using this method to isolate DNA from whole blood of myocardial infarction patients for studies on the apolipoprotein B gene.
Immunoradiometric assay of parathyrin with polyclonal and monoclonal region-specific antibodies
Authors: Bouillon R , Coopmans W , Degroote D E , Radoux D , Eliard P H
Published: January 2020

In this immunoradiometric assay (IRMA) of parathyrin (PTH) a polyclonal anti-amino-PTH(1-34) is the capture antibody and a radiolabeled monoclonal anti-hPTH(44-68) is the second antibody. Gel filtration of serum from a hyperparathyroid patient yielded only a single peak of PTH, corresponding to the elution position of synthetic PTH(1-84). Healthy elderly individuals (ages 78 +/- 5 y, mean +/- SD, n = 45) had PTH concentrations (21 +/- 13 ng/L) not significantly higher than those from healthy younger (38 +/- 11 y) adults (20 +/- 8 ng/L, n = 94). Assay results agreed well with those obtained with a carboxyl-terminal PTH assay both in normal subjects (r = 0.63, P less than 0.001) and in patients with primary hyperparathyroidism (r = 0.59, P less than 0.001). Both assays equally discriminated patients with surgically confirmed primary hyperparathyroidism from normal individuals, but the PTH(1-84) IRMA also allowed a nearly absolute discrimination between normal subjects and patients with primary hypoparathyroidism (undetectable serum PTH in 18 of 21 cases) and secondary hypoparathyroidism (caused by hypercalcemia that was caused by a malignant tumor, PTH 1.3 +/- 1.3 ng/L, n = 32). Moreover, the PTH(1-84) IRMA is more sensitive (detection limit in serum, 0.8 ng/L) and easier and quicker to perform than the carboxyl-terminal assay.
Lipoprotein lipids in women with androgen excess: independent associations with increased insulin and androgen
Authors: Wild R A , Applebaum-Bowden D , Demers L M , Bartholomew M , Landis J R , Hazzard W R , Santen R J
Published: January 2020

Concentrations of triglycerides are increased and concentrations of high-density lipoprotein (HDL) cholesterol are low in women with hyperandrogenism. These alterations could be related to excessive androgen or estrogen, to hyperinsulinism, or to a combination of these abnormalities. We examined their independent influences on lipids in 21 women with hyperandrogenism, subgrouped according to apparent source of androgen excess. Results for lipid, androgen, and insulin did not differ among subgroups, so these data were pooled. Free plus albumin-bound testosterone (uT) was correlated with triglycerides (r = 0.69, P less than 0.01) and HDL cholesterol (r = -0.56, P less than 0.01). Both triglycerides (r = 0.66, P less than 0.01) and HDL cholesterol (r = -0.48, P less than 0.05) were also correlated with insulin measured during fasting. Partial correlation revealed that, after adjusting for insulin, lipids were associated with uT. This suggests that androgen excess is independently related to lipid excess. Insulin also was correlated with lipids when adjusted for uT. Free plus albumin-bound estradiol was not associated with any of the lipids. We conclude that altered lipids in women with hyperandrogenism result from the independent effects of androgen and insulin.
Effect of stripping thyroxin from thyroxin-binding globulin on the measurement of free thyroxin in serum by equilibrium dialysis and by radioimmunoassay
Authors: Nakagawa T , Matsumura K , Takeda K , Shinoda N , Matsuda A , Matsushita T , Tagami T
Published: January 2020

In considering factors that might influence measurement of free thyroxin (T4), we evaluated the proportion (%) of T4 that could be stripped from thyroxin-binding globulin (TBG). The percentage of free T4 was measured in serially diluted sera from four normal subjects, four patients with hyperthyroidism or hypothyroidism, four pregnant women, and four malnourished subjects with low TBG. The critical percentage of stripping was determined by the product of the percentage free T4 and the critical dilution factor (the point where the percentage free T4 began to decrease). The mean values obtained for the respective patient groups--6.38%, 2.76%, 16.73%, 9.75%, and 4.28%--were proved to be related to the rate of saturation of TBG with T4. Values for percentage stripping determined with the "GammaCoat two-step RIA" and the "LiquiSol RIA" were well within the critical percentage stripping by equilibrium dialysis, except in the case of low-TBG serum as measured by LiquiSol RIA. Free T4 concentration as measured by LiquiSol RIA decreased as sample volume decreased. These findings were ascribed to the relatively high values for percentage stripping in the LiquiSol RIA, which led to erroneously low values for free T4.
Falsely low estimation of triglycerides in lipemic plasma by the enzymatic triglyceride method with modified Trinder's chromogen
Authors: Shephard M D , Whiting M J
Published: January 2020

The enzymatic assay of triglyceride, based on the use of L-glycerol-3-phosphate oxidase (EC 1.1.3.21) and a modified Trinder's chromogen involving 4-chlorophenol, is subject to strong negative interference at concentrations of triglyceride greater than 20 mmol/L, such as occur in grossly lipemic plasma. This interference is caused by the rapid utilization of oxygen, resulting in the reaction becoming transiently anaerobic. The dye product already formed may then be reduced ("bleached") by acting as an alternative electron acceptor for glycerol-3-phosphate oxidase. Reduction of the dye leads to a marked decrease in final absorbance at 505 nm. Grossly underestimated values for triglyceride concentrations, apparently within the linear range of the assay, may therefore be inadvertently obtained with equilibrium methods. We suggest that samples giving unexpectedly low results for lipemic plasma should be re-assayed after dilution or with use of a smaller volume of sample.
Discriminant functions in differential diagnosis of hypercalcemic patients
Authors: Gibb J A , Ogston S A , Paterson C R , Evans J R
Published: January 2020

Using readily available biochemical assays of plasma and urine constituents, we have defined discriminant functions useful as a guide to the differential diagnosis of patients with hypercalcemia. The decreasing rank order of contribution of the variables to the discriminant functions was as follows: plasma albumin, plasma phosphate, plasma chloride, log10 (calcium excretion per liter of glomerular filtrate), and log10 (plasma gamma-glutamyltransferase). Discriminant functions have been defined for patients with values for plasma creatinine above and below 185 mumol/L, and for practical conditions in which plasma and urine samples, or plasma samples only, are available.
Quantifying the MB isoenzyme of creatine kinase with the Abbott "IMx" immunoassay analyzer
Authors: Brandt D R , Gates R C , Eng K K , Forsythe C M , Korom G K , Nitro A S , Koffler P A , Ogunro E A
Published: January 2020

In this two-step automated assay of the MB isoenzyme of creatine kinase (CK-MB), developed for the Abbott "IMx" immunoassay analyzer, monoclonal anti-CK-MB antibody immobilized onto latex microparticles and polyclonal anti-CK-MM antibody coupled to alkaline phosphatase are used. Within-run CVs ranged from 3.9% to 9.0%, between-run CVs from 0.0% to 5.6%, and the sensitivity was 0.2 microgram/L. Twenty-four results can be obtained in about 37 min. Analytical recovery of CK-MB added to human serum or plasma ranged from 89% to 109%. Icteric, lipemic, or hemolyzed samples did not interfere with CK-MB recovery. Cross-reactivity with CK-MM and CK-BB was 0.012% and 0.001%, respectively. The normal reference interval was 0-5 micrograms/L. IMx CK-MB results correlated well with CK-MB enzyme activity as determined by electrophoresis (n = 203; r = 0.97; slope = 0.90; y-intercept = -4.29) and with commercial immunoassays. We think that this assay will be useful for confirmation of acute myocardial infarction, both in critical-care units and in the clinical laboratory.
Intracellular free amino acid patterns in duodenal and colonic mucosa
Authors: Ollenschläger G , Langer K , Steffen H M , Schrappe-Bächer M , Schmitt H , Allolio B , Roth E
Published: January 2020

We report for the first time the concentrations of free amino acids in human intestinal biopsies obtained by routinely performed endoscopy. We studied 15 medical patients with no changes of the mucosa and six HIV-infected persons with duodenitis. The mean (and SD) sum of all amino acids, taurine excepted, was 61.9 (5.4) mmol/kg dry weight in duodenal biopsies of HIV-negative subjects (n = 11) and 82.9 (0.6) mmol/kg in colonic specimens: 50% (44%) of the total (minus taurine) consisted of aspartate and glutamate and 14% (12%), of the essential amino acids. The relative amino acid pattern in duodenum and colon differed completely from that for muscle: aspartate was fourfold higher; glutamate, phenylalanine, glycine, valine, leucine, and isoleucine were about twofold higher. In contrast, glutamine amounted only to 4% (duodenum) to 14% (colon) of muscle glutamine. In duodenal biopsies of the HIV-infected persons, we found significantly (P less than 0.01, except glutamine: P less than 0.025) increased concentrations of glutamate (24.1 vs 17 mmol/kg dry weight), ornithine (1.4 vs 0.4), valine (2.2 vs 1.7), and glutamine.
Growth-hormone-releasing hormone
Authors: Vance M L
Published: January 2020

Growth-hormone-releasing hormone (GHRH, somatoliberin) is the hypothalamic peptide hormone that specifically stimulates synthesis and release of growth hormone (GH, somatotropin) by somatotrope cells of the anterior pituitary gland. GHRH is the last of the classically postulated hypothalamic hormones to be characterized, synthesized, and used in clinical medicine. In this review of GHRH, I discuss the discovery and characterization of the peptide, its role in the regulation of GH secretion, and its clinical use in pathological states of GH excess and GH deficiency. The two most clinically useful aspects of GHRH are to establish the etiology of GH deficiency, most commonly the result of a hypothalamic GHRH deficiency, and to treat GH-deficient children. Use of GHRH as therapy for GH deficiency currently is experimental and, to date, results encourage the idea of a therapeutic role for this peptide in promoting endogenous GH secretion with resulting acceleration of linear growth.
Muscle enzymes and isoenzymes in Emery-Dreifuss muscular dystrophy
Authors: Bialer M G , Bruns D E , Kelly T E
Published: January 2020

Emery-Dreifuss muscular dystrophy (EDMD) is a rare X-linked muscular dystrophy. Creatine kinase (CK) activity usually is increased in serum of affected males, but results for aldolase and lactate dehydrogenase (LD) in serum have been inconsistent, as have those for CK in carrier females. There have been few studies of CK-MB or LD isoenzyme-1 (LD-1) in EDMD. We measured CK, CK-MB, LD, LD-1, and aldolase activity in sera of 84 members of two large families with EDMD. DNA analysis had been carried out on all subjects. Although CK, LD, and aldolase activities were significantly increased in affected males, CK activity was the most consistently increased and was the least subject to artifactual increases. Mean CK-MB in serum was mildly increased, but LD-1 was within the normal reference interval, suggesting that CK-MB is increased in skeletal muscle in EDMD, as has been found in other forms of dystrophy. CK decreased with age in affected males. We saw no significant increases of muscle enzymes or isoenzymes in 33 EDMD carriers studied, of whom 19 were obligate carriers and 14 had been identified by DNA analysis.
Molecular probe protocol for determining carrier status in Duchenne and Becker muscular dystrophies
Authors: Prior T W , Friedman K J , Highsmith W E , Perry T R , Silverman L M
Published: January 2020

By use of cDNA probes, molecular deletions were identified in 66.6% of 42 patients with Duchenne muscular dystrophy (DMD) or Becker muscular dystrophy (BMD). Owing to this high deletion rate, a new strategy for detecting DMD/BMD carriers is feasible in which the polymerase chain reaction is used as an initial screen for detecting the deletions occurring in specific deletion-prone exons. Because the deletions do not occur randomly, specific cDNA probes are utilized first with Southern blot analysis. Identification of a deletion permits direct analysis for DMD carrier status and removes the inherent limitations of the conventional restriction fragment length polymorphism technique. Carrier status is determined by scanning the autoradiographs with a densitometric spectrophotometer or by detection of a junction fragment.
Manganese content of the cellular components of blood
Authors: Milne D B , Sims R L , Ralston N V
Published: January 2020

We measured the manganese content of whole blood, plasma, platelets, mononucleated cells, polymorphonucleated cells, and erythrocytes. Platelets and blood cells were separated from whole blood by use of discontinuous gradients of colloidal polyvinylpyrrolidone-coated silica (Percoll), and their manganese content was measured by Zeeman graphite furnace atomic absorption spectrophotometry, after digestion with nitric acid and hydrogen peroxide. Erythrocytes account for about 66% of the manganese in whole blood, whereas the "buffy coat"--platelets and leukocytes--accounts for about 30%. The "buffy coat" components turn over more rapidly than do erythrocytes, so their manganese content may better indicate the body's manganese status.
Differentiating various abnormalities of thyroxin binding to serum proteins by radioelectrophoresis of thyroxin and immunoassay of binding proteins
Authors: Pandian M R , Morgan C , Nelson J C , Fisher D A
Published: January 2020

Using the simple method of protein analysis described here, we could identify thyroxin (T4)-binding-protein abnormalities in euthyroid patients with hyperthyroxinemia or hypothyroxinemia. Serum incubated with [125I]thyroxin was analyzed by agarose gel electrophoresis, with bromphenol blue staining of protein. The relative distribution of radioactive T4 was determined for each binding protein--thyroxin-binding globulin (TBG), transthyretin, albumin, and T4-binding immunoglobulin (when present)--and the mass of T4 bound to each was determined. We also used sensitive immunoassays to quantify TBG, transthyretin, and albumin concentrations, then calculated the mass of T4 (as determined by electrophoresis) bound per unit mass of the respective binding protein. When the concentration of binding proteins was altered (e.g., TBG excess or TBG deficiency), the T4 binding/mass ratio for each protein remained within the expected range; but when the functional affinity of a binding protein was altered--as in dysalbuminemic hyperthyroxinemia and in low-T4 nonthyroidal illness--this ratio was abnormal. This procedure can be used to help identify TBG excess, TBG deficiency, dysalbuminemic hyperthyroxinemia, prealbumin-associated hyperthyroxinemia, variant TBG with reduced affinity for T4, euthyroid sickness, and T4-binding autoantibodies.
A simulation study of the Westgard multi-rule quality-control system for clinical laboratories
Authors: Wood R
Published: January 2020

A particular version of the Westgard-Barry-Hunt-Groth (Westgard) multi-rule quality-control system is studied by Monte Carlo simulation. The principal focus is on the amount of distortion in the working power curve caused by skewness in the distribution of analyte measurements. The effect of using only the first two rejection modes of the Westgard system is also studied.
Suppression of endogenous cortisol for evaluating pharmacodynamics of prednisolone in early allograft rejection in renal transplantation
Authors: Oka K , Hirano T , Shimodaira H , Homma M , Sakurai E , Tamaki T , Kozaki M
Published: January 2020

Concentrations of endogenous cortisol were examined in 34 kidney-transplant recipients by improved "high-performance" liquid chromatography. Ten recipients were treated with prednisolone and azathioprine, the others with prednisolone and cyclosporine. Peripheral serum samples were collected just before transplantation, daily for two weeks after the transplant, weekly until discharge for about two months, and then monthly or occasionally. Mean (+/- SD) cortisol concentrations, initially 145 +/- 87 micrograms/L, decreased immediately to 5.93 +/- 5.11 micrograms/L after transplant, remained at almost these same values for two months, and then swiftly increased to 51 +/- 59 micrograms/L by 1000 days. Cortisol concentrations within the period characterized by a cumulative dose of prednisolone at 300-700 mg were correlated significantly with the presence or absence of acute allograft rejection; patients with cortisol greater than 4 micrograms/L had a higher risk of rejection. The majority of stable patients showed cortisol concentrations between 1 and 4 micrograms/L throughout the cumulative prednisolone period characterized above. Concentrations less than 1 microgram/L after high-dose administration of methylprednisolone were accompanied by severe lung infection. We conclude that suppressed concentrations of endogenous cortisol, as assessed by highly specific HPLC, might provide a basis for predicting the therapeutic efficacy and adverse effects of prednisolone.
Response of various indices of iron status to acute iron depletion produced in menstruating women by low iron intake and phlebotomy
Authors: Milne D B , Gallagher S K , Nielsen F H
Published: January 2020

We investigated response sensitivities of indices of iron status to controlled iron depletion and repletion in 11 premenopausal women. The women were depleted of storage iron (as reflected by serum ferritin) through a combination of a low-iron diet and phlebotomy. They then consumed a diet containing 13.7 mg of iron per 2000 kcal, supplemented with either ascorbic acid or placebo (for 5 1/2 weeks) and a daily 50-mg iron supplement (for the subsequent 17 days). The relative sensitivities of different indices for detecting iron depletion were as follows: ferritin greater than % transferrin saturation greater than plasma iron greater than hemoglobin greater than hematocrit greater than zinc protoporphyrin (ZnPP) and erythrocyte protoporphyrin (EP). Ascorbic acid treatment during repletion, before iron supplementation, significantly (P less than 0.05) affected changes in hemoglobin, ZnPP, ZnPP/heme, and EP/heme. Changes in heme synthesis evidently do not occur until iron stores are depleted and, conversely, during iron repletion hematopoiesis must be satisfied before iron stores, as reflected by ferritin, increase. Thus, the use of only one index of iron status is of limited value for detecting iron depletion.
Immobilization of immunoglobulins onto surface-treated and untreated polystyrene beads for radioimmunoassays
Authors: Kakabakos S E , Livanlou E , Evangelatos G P , Ithakissios D S
Published: January 2020

The immunoreactivity of anti-triiodothyronine (anti-T3) IgG, pretreated at acidic pH and adsorbed onto polystyrene beads, was significantly greater than that determined for native anti-T3 IgG immobilized at optimum pH 3.5. Acidic-pH-pretreated IgG, adsorbed at pH 7.0 onto ethanol-treated beads, had less immunoreactivity than that on untreated beads and gave values similar to those of native IgG adsorbed onto activated beads at pH 9.6. The rate of immobilization onto treated beads was significantly greater than onto untreated ones, and the binding was reproducible (intra- and inter-batch coating CV, 3.7-4.6% and 7.2%, respectively) and very resistant to successive washings. A post-washing incubation in 10 mg/L bovine serum albumin solution was required to eliminate the decreased immobilized immunoreactivity caused by some washing reagents. For solid-phase radioimmunoassays of T3, 40% less antibody was needed when acidic-pH-pretreated IgG was adsorbed onto untreated beads or when native IgG was adsorbed onto ethanol-treated beads, in comparison with native IgG adsorbed onto untreated beads. Results and assay characteristics with such beads were comparable with results for solid-phase and liquid-phase polyethylene glycol assay of T3.