bluuebunny/crossref_metadata_2025 huggingface dataset:
Adaptation of "EMIT" technique for serum phenobarbital and diphenylhydantoin assays to the miniature centrifugal analyzer.
Authors: Brunk S D , Hadjiioannou T P , Hadjiioannou S I , Malmstadt H V
Published: January 2020

We used a miniature centrifugal analyzer in a spectrophotometric rate-measurement mode to determine the anticonvulsant drugs phenobarbital and diphenylhydantoin in serum, by use of a modified enzyme immunoassay ("EMIT", Syva Corp.) We decreased reagent cost per determination by at least sixfold by means of microscale techniques. Also, the analysis rate is increased by measuring multiple samples simultaneously. Our method requires only 3 mul of serum for duplicate determinations. Replicate analyses of sera containing phenobarbital and diphenylhydantoin gave reaction rates with a CV of 1.5%. Run-to-run CV was 15%. Analytical recovery for drug-supplemented serum samples was 98%, and results for a series of samples compared well with results obtained by gas chromatography (for phenobarbital, r = 0.95; for diphenylhydantoin, r = 0.91).
Monitoring drug concentrations in a case of combined overdosage with primidone and methsuximide.
Authors: Johnson G F , Least C J , Serum J W , Solow E B , Solomon H M
Published: January 2020

We describe a case of fatal overdosage with primidone and methsuximide. During the early phase of the patient's hospital course we found concentrations of methsuximide, N-desmethylmethsuximide, and primidone in serum that far exceeded the usual therapeutic concentrations, as determined by gas-liquid chromatography. Determination of N-desmethylmethsuximide in peritoneal fluid demonstrated concentrations comparable to those in serum. This led to the therapeutic decision to manage the patient by dialysis. Subsequently, serum samples collected during the course of hospitalization were analyzed quantitatively by gas-liquid chromatography for methsuximide, N-desmethylmethsuximide, primidone, phenobarbital, and diphenylhydantoin. Selected serum specimens were also analyzed by gas chromatography-mass spectrometry, and N-methyl-2-hydroxymethyl-2-phenylsuccinimide, a metabolite of methsuximide not previously described in human serum, was identified by analysis of its mass spectrum.
Immobilized glucose oxidase used in the continuous-flow determination of serum glucose.
Authors: León L P , Narayanan S , Dellenbach R , Horvath C
Published: January 2020

We used a tubular glucose-oxidase wall reactor in the "AutoAnalyzer II" continuous-flow analytical system to determine glucose in blood serum. Sensitivity was high and wash characteristics were satisfactory with use of a 30-cm tube containing immobilized glucose oxidase. Results compared favorably with those of the conventional free-enzyme method. More than 25000 such assays can be performed with a single enzyme tube, which also shows long-term storage stability. Because of the steady-state chemistry 60 samples can be analyzed per hour. The linearity of the method is excellent and sample interaction from 5.0 to 1.0 g/liter is less than 5%. Results correlate well (greater than 0.993) with those obtained with both the neocuproine method used in the "SMA 12/60" multichannel analytical system and the free glucose oxidase method used in the AutoAnalyzer.
Separation of folate binding protein from human serum by DEAE-cellulose column chromatography.
Authors: Zettner A , Duly P E
Published: January 2020

On diethylaminoethyl-cellulose column chromatography, the folate binding protein in the serum of 21 patients eluted in the early effluents as a single sharply defined peak. The chromatographic behavior of the folate binder remained unchanged whether or not the serum was, before chromatography, complexed with tritium-labeled pteroylglutamic acid ([3H]PGA), dialyzed, or charcoal-adsorbed. Heating to 100 degrees C for 10 min dissociated the [3H]PGA-binder complex while destroying the folate binding property. The presence or appearance of this folate binder in increased amounts in the serum of patients with various diseases may be related to conditions of increased tissue turnover.
Methotrexate in blood, urine, and cerebrospinal fluid of children receiving high doses by infusion.
Authors: Wang Y , Lantin E , Sutow W W
Published: January 2020

Methotrexate was determined in plasma (31 patients), urine (eight patients), and cerebrospinal fluid (two patients) after high doses (35 to 150 mg/kg) by infusion, with citrovorum factor rescue. Concentrations in plasma were proportional to dose at 6, 24, 72 h after beginning treatment, but this trend was very minimal for samples obtained at 48 h. Clinical toxicity probably will not be serious if the methotrexate concentration in the serum is less than 4.5 X 10(-6) mol/liter at 48 h after the start of a 6-h infusion. In both of the two patients so examined, therapeutic concentrations appeared in cerebrospinal fluid after intravenous infusion of the drug. A kinetic enzymic method, radioassay, and radioimmunoassay all yielded similar results for drug concentrations.
In vitro changes in blood P50 and erythrocyte 2,3-diphosphoglycerate concentration.
Authors: Aberman A , Cavanilles J M , Michaels S , Shubin S H , Weil M H
Published: January 2020

We studied in vitro changes in P50 and erythrocyte 2,3-diphosphoglycerate concentration occurring in blood 2 to 8 h after venipuncture. When blood was incubated at 37 degrees C, significant decreases in P50 were observed at 2, 4, and 8 hr. Such a change was significantly less when blood was kept at 4 degrees C. The rate of decrease in P50 was not changed when pH was altered by adding either lactic acid or sodium bicarbonate to the blood before incubation at 37 degrees C for 2 h. The erythrocyte 2,3-diphosphoglycerate concentration of blood incubated at 37 degrees C did not change by 2 h, but had significantly decreased by 4 h. To avoid in vitro changes, we recommend that P50 be determined as soon as possible for blood sampling.
Ultramicromethod for determination of plasma uric acid.
Authors: Kamoun P , Lafourcade G , Jerome H
Published: January 2020

We describe an ultramicromethod for determination of uric acid by a manual procedure. Hydrogen peroxide, formed when uric acid is oxidized in the presence of uricase, oxidatively couples two molecules of p-hydroxyphenylacetic acid in the presence of peroxidase to produce a highly fluorescent compound. The specificity of the uricase reaction is retained, and the fluorometry results in a higher sensitivity (but a lower precision) than that obtained by applying spectrophotometric methods. As little as 50 ng of uric acid can be determined, and only 10 mul of plasma is required. Values obtained for human plasma are higher than those obtained by the spectrophotometric method.
Enzyme-immunoassay.
Authors: Wisdom G B
Published: January 2020

The bases of the different enzyme-immunoassay techniques are described and their applications to the identification and quantification of antigens, haptens, and antibodies in biological fluids are discussed. Substances for which enzyme-immunoassays have been developed are listed. The enzymes used as labels, the methods of linking them to proteins, and the methods for separating free and bound labeled material are described. The reliability and practicality of the enzyme-immunoassay methods are reviewed.
Some theory of reference values. II. Comparison of some statistical models of intraindividual variation in blood constituents.
Authors: Harris E K
Published: January 2020

Three models of intraindividual variation are reviewed, and statistical methods for distinguishing among them are discussed. Application of these methods to short series of observations from healthy individuals indicates that, in the large majority of cases, a strictly homeostatic model is appropriate for such constituents as serum calcium and magnesium. In less closely controlled variables, e.g., serum cholesterol and uric acid, a nonstationary, "rndom walk" model appears moresuitable in most cases. A more general autoregressive model, which includes the other models as extreme cases, could be used to describe all degrees of homeostatic control. This model is more complex, however, and requires at least 10 observations to yield estimates of acceptable precision. Moreover, it is sensitive to fluctuations in within-batch analytical variance. When biological variance is small relative to analytical variance, all three models yield essentially the same predicated values. To illustrate their use, these models have been applied to four short individual series of cholesterol observations showing increasing amounts of intrapersonal variation over long periods of time. I suggest that when less than 10 observations over time are available, the strictly homeostatic model and the nonstationary model be used to derive a "critical range" for assessing future changes. When longer series are available, the more general model might replace the other two for this purpose, if analytical variation has remained reasonably stable (within +/- 20% of its average value) during the period of observation. Much more experience with the use of all three models in health monitoring programs would be highly desirable.
Protein-bound carbohydrates in breast cancer. Liquid-chromatographic analysis for mannose, galactose, fucose, and sialic acid in serum.
Authors: Mrochek J E , Dinsmore S R , Tormey D C , Waalkes T P
Published: January 2020

We describr high-resolution chromatographic analysis for protein-bound sialic acid in serum, with use of a cerate oxidimetric detector. Values for sera from normal women averaged 680.5 mg/liter, with a coefficient of variation of 23%. Including data obtained by previously developed chromatographic procedures for protein-bound mannose, galactose, and fucsoe, we assessed sera from breast-cancer patients whose malignancy had been categorized as either stable, responsive, or progressive (based on clinical observations spaced from two to five months apart). All of 12 responsive patients had decreases of protein-bound fucose averaging 34.5% (SD, 16.1) and all of 10 patients with progressive disease had increases averaging 38.3% (SD 21.5). Changes in fucose averaged less than 6.7% (SD, 4.9) for eight patients with clinically stable breast cancer. Changes in protein-bound mannose, galactose, and sialic acid did not correlate as well as did fucose with the clinical disease status of the patients.
Double spike in the electropherogram of a myeloma serum, from Bence Jones protein.
Authors: Vladutiu A O , Kohli R K
Published: January 2020

Serum from a 71-year-old man with multiple myeloma complicated with renal failure showed a monoclonal IgG lambda. A second spike appearing in the serum protein electropherogram, suggesting a biclonal gammopathy, was found to be due to lambda Bence Jones protein (29.9 g/liter). Lambda Bence Jones protein was also found in smaller concentration (3.8 g/liter) in the urine. Tetramers of Bence Jones protein were not demonstrable by gel filtration and ultracentrifugation, and renal failure was probably the main reason for this very high concentration of Bence Jones protein in the serum.
Clinical relevance of polyamines as biochemical markers of tumor kinetics.
Authors: Russell D H
Published: January 2020

The polyamines, spermidine and spermine, and their diamine precursor, putrescine, constitute a unidirectional biosynthetic pathway whose biosynthetic enzymes and accumulation patterns appear to play important roles in the regulation of growth processes. Concentrations of these compounds in physiological fluids are low or undetectable under normal conditions, are elevated in patients with metastatic cancer, and are thought to reflect growth (putrescine concentrations) and cell turnover (spermidine concentrations) of the organism. Cancers are a broad spectrum of diseases in which there are altered growth fractions and cell-turnover fractions, and therefore cancer chemotherapeutic agents have been developed to take tumor kinetics into account. Because changes in polyamines in physiological fluids reflect cell kinetics, this review compiles evidence of their efficacy as biochemical markers of cancer and suggests their possible usefulness to clinicians in rapidly assessing tumor response to chemotherapy or to multimodality therapy.
Combined gel filtration, biuret/copper method compared with an immunochemical method for urinary protein measurement.
Authors: Peele J D , Gadsden R H , Loadholt C B
Published: January 2020

We compared an immunochemical method specific for plasma proteins with a chemical method, in which interfering substances are separated by gel filtration and "total" urinary protein is determined by the biuret reaction followed by reaction of protein-bound copper with diethyldithiocarbamate after a second gel filtration to remove nonprotein-bound copper. More than 250 24-h urine samples were analyzed by each method. There was linear agreement and a correlation of 0.96 between the two methods, but urinary protein values determined by the immunochemical method, especially for patients with multiple myeloma, were lower than by the chemical method.
Biochemical characterization of the tartrate-resistant acid phosphatase of human spleen with leukemic reticuloendotheliosis as a pyrophosphatase.
Authors: Lam K W , Yam L T
Published: January 2020

A tartrate-resistant acid phosphatase was isolated from a human leukemic spleen by freeze-thawing in saline and purified by repeated chromatography on carboxymethyl-cellulose. The purified enzyme has a molecular weight of 64 000. It catalyzes the hydrolysis of inorganic and organic pyrophosphate as well as the phenolic ester of monoorthophosphate, with optimal activity between pH 5 and 6. However, there is no activity toward mono-orthophosphate esters of aliphatic alcohols. The present data have identified its catalytic function as a pyrophosphatase. However, it has properties different from the pyrophosphatase previously observed in normal animal tissues.
A semi-automated fluorometric method for total estrogens in pregnancy urine.
Authors: Frye R , Fong P F , Johnson G F , Rock R C
Published: January 2020

We report a continuous-flow fluorometric method for total urinary estrogens that involves the Kober reaction, with extraction of the reaction product into dichloroethane containing trichloroacetic acid as described by Hahnel and Jones [Clin. Chim. Acta 16, 185 (1967)]. The dichloroethane extraction gives greater stability to the Kober color and sharper separation of aqueous and organic phases. The analytical system is adjusted to give maximum response to estriol 16alpha,beta-D-glucuronide, the principal estrogen conjugate in urine from late pregnancy, when calibrated with estriol standards. An initial 20-fold dilution of the sample with water increased analytical recovery of estriol conjugates from urine while maintaining adequate fluorescent response. Glucose interference was reduced by dilution and eliminated by treatment with sodium borohydride. Urinary protein up to 20 g/liter did not interfere. Total estriol, as determined by gas-liquid chromatography, comprises about 70% of total urinary estrogens in late pregnancy as measured by our continuous-flow fluorometric method. A reference range is presented based on 209 randomly collected urine specimens in which total urinary estrogens are expressed as a ratio to creatinine.
High-pressure liquid-chromatographic method for determination of gentamicin in plasma.
Authors: Peng G W , Gadalla M A , Peng A , Smith V , Chiou W L
Published: January 2020

A rapid, specific method for measuring gentamicin in plasma by high-pressure liquid chromatography was developed. After deproteinization, gentamicin in the supernate was dansylated and extracted into ethyl acetate. The organic extract was chromatographed on a microparticulate reversed-phase column, which was eluted with aqueous acetonitrile. Use of the dansyl derivative enables fluorometry, for more sensitive quantitation. Various factors that could affect the assay sensitivity were investigated. With 0.2-ml plasma samples, the method can accurately measure as little as 1 mg of gentamicin per liter. We encountered no interferences from plasma supplemented with various drugs or plasma of patients who were on therapy with other drugs. This method can also separate gentamicin C1 from C1a and C2, all of which are present in various ratios in commercial dosage forms. This method is also applicable to gentamicin determination in urine.
Effect of incubation with Mg2+ on the measurement of alkaline phosphatase activity.
Authors: Rej R
Published: January 2020

Human serum incubated with 2-amino-2-methyl-1-propanol (860 mmol/liter) and magnesium ion (270 mumol/liter) at pH 10.35 showed greater alkaline phosphatase (EC 3.1.3.1) activity than if an equal amount of magnesium ion was added at the time of measurement. The apparent increase is due in part to a slight lability of serum alkaline phosphatase in 2-amino-2-methyl-1-propanol, which is prevented by the inclusion of magnesium. For some sera, however, a portion of this increased activity is real rather than artifactual. Use of serum rather than 4-nitrophenylphosphate to initiate the reaction produced relatively low activities and, in some cases, markedly nonlinear (increasing) rate progress curves. The behavior of some commercial lyophilized control sera differed significantly from that of patients' sera, in particular exhibiting a marked lability in the presence of 2-amino-2-methyl-1-propanol. Incubation of these labile materials with Mg2+ slightly improved their stability; addition of Zn2+ plus Mg2+ markedly stimulated and completely protected their alkaline phosphatase activity.
Adaptation to the centrifugal analyzer of an enzymatic method for the measurement of lactate in plasma and cerebrospinal fluid.
Authors: Alderman J A , Cross R E
Published: January 2020

A fully enzymatic method for measurement of l(+)-lactate has been adapted for use with a centrifugal analyzer. The procedure is based on a two-point fixed-time kinetic approach for elimination of static interferences, and commercially available reagents are used. Specimen collection and preservation are simplified by the use of standard evacuated tubes containing sodium fluoride. Under these conditions lactate is stable for as long as seven days at 4 degrees C, and deproteinization is unnecessary. The method is extremely simple and ideally suited for emergency and routine use. Determinations may be performed on plasma or cerebrospinal fluid. Sensitivity and precision are excellent, linearity extends to 8.0 mmol/liter, and results compare well with those obtained using the manual enzymatic procedure.
Methaqualone metabolites in human urine after therapeutic doses.
Authors: Kazyak L , Kelley J A , Cella J A , Droege R E , Hilpert L R , Permisohn R C
Published: January 2020

We measured five principal metabolites of methaqualone in the urine of seven volunteers after single and multiple doses of the drug. Urine, collected for up to 72 hours after the last dose, was analyzed for methaqualone and its principal metabolites by high-resolution capillary-column gas chromatography. The major biotransformation of methaqualone under therapeutic conditions occurred through benzylic and para-hydroxylation of the o-tolyl moiety. Methaqualone itself was present in concentrations of no more than 1 mg/liter, if it could be detected at all. The observed physiological effects ant total urinary excretion of metabolites reflected the cumulative nature of the parent drug when it was administered in multiple doses. No clear relationship was found between appearance of a specific metabolite and time after ingestion of the drug, although higher amounts of 2-methyl-3-(2'-hydroxymethylphenyl)-4(3H)-quinazolinone were noted in those individuals who tolerated the drug less well.
Improved colorimetry of urinary 3-methoxy-4-hydroxyphenylacetic acid (homovanillic acid).
Authors: Knight J A , Haymond R E
Published: January 2020

Homovanillic acid is an important metabolite of dopamine, and a high proportion of patients with neuroblastoma excrete increased amounts of it in their urine. When this diagnosis is suspected, both homovanillic acid and vanilmandelic acid should be measured, because such a combined measurement reportedly leads to detection of 95% of cases. Although rapid and reliable chemical methods are available for vanilmandelic acid, the same is not true for homovanillic acid. We report here a colorimetric method for homovanillic acid that reasonably fills this void and that can be used in most clinical laboratories. In addition, we present normal values determined for children of various age groups.
Removal of an endogenous antigen from an antibody to increase its effective affinity constant, as illustrated by triiodothyronine assay.
Authors: Oliver L K , Cano C
Published: January 2020

Antisera to triiodothyronine were shown to contain large quantities of the hormone, well in excess of normal circulating concentrations. Extracting the triiodothyronine from the antiserum with alkaline ethanol yielded an antibody of increased affinity. Use of this antibody in a radioimmunoassay resulted in a fourfold increase in sensitivity as compared with the unextracted material.
Semi-automated vs. visual reading of urinalysis dipsticks.
Authors: Peele J D , Gadsden R H , Crews R
Published: January 2020

Semi-automated reflectance readings of urinalysis dipsticks showed better precision than visual readings in the case of artificially prepared urine samples containing glucose, ketone, and protein. Actual pathological specimens containing glucose, protein, and nitrite also showed that instrumental readings were better than visual. Results of repeated visual readings spread over three different color blocks for certain concentrations of glucose, ketone, and protein, whereas repeated instrumental readings were never spread greater than two color blocks. Subjectivity can be reduced by semi-automated dipstick urinalysis.
Measurement of carbamazepine and its epoxide metabolite by high-performance liquid chromatography, and a comparison of assay techniques for the analysis of carbamazepine.
Authors: Mihaly G W , Phillips J A , Louis W J , Vajda F J
Published: January 2020

We describe a modified high-performance liquid-chromatographic method for the simultaneous analysis of carbamazepine andits biologically active metabolite, carbamazepine-10, 11-epoxide. Concentrations of both these compounds in the plasma of 35 epileptic patients receiving chronic carbamazepine therapy are presented. Concentrations of carbamazepine in plasma were related to those of carbamazepine-10, 11-epoxide (r - 0.495, P less than 0.05). Total daily doses of carbamazepine were better correlated with plasma concentrations of carbamazepine-10, 11-epoxide (r = 0.714, P less than 0.001) than of carbamazepine (r = 0.269, P greater than 0.05). Close correlations were found between results of the three assay procedures we used to measure plasma carbamazepine concentrations: high-performance liquid chromatography, gas-liquid chromatography, and enzyme immunoassay. Correlation coefficients exceeded 0.97 and regression slopes were near unity, indicating that all three procedures were individually specific for the quantification of plasma carbamazepine.
Use of gel filtration to examine the distribution of calcium among serum proteins.
Authors: Toffaletti J , Savory J , Gitelman H J
Published: January 2020

Gel filtration of serum by use of dextran bisacrylamide beads ("Sephacryl S-200," fractionation range: 5000-250 000 daltons), separates the serum proteins into three distinct peaks. We used an eluent containing (per liter) 140 mmol of sodium, 1.10 mmol of calcium, 0.50 mmol of magnesium, and 10 mmol of 2-([2-hydroxy-1, 1-bis(hydroxymethyl)ethyl]amino)ethanesulfonate buffer (pH 7.43 at 37 degrees C) to provide the physiological conditions necessary to maintain the equilibrium of bound calcium in serum. We examined the calcium binding by each of the three protein groups. Calculating the calcium bound per gram of protein in each peak, we found that proteins in the first peak (alpha2-macroglobulin, IgM, haptoglobins) bind about the same amount of calcium per gram as does albumin. The proteins of the second peak (largely IgG and IgA) bind less calcium than does albumin, which is the protein accounting for 90% or more of the third peak. We also were able to calculate intrinsic association constants for calcium/albumin under physiological conditions.
Some aspects of sodium nitroprusside reaction with human erythrocytes.
Authors: Spiegel H E , Kucera V
Published: January 2020

Sodium nitroprusside is an excellent agent for lowering blood pressure in hypertensive emergencies, for producing controlled hypotension during anesthesia, and for treating acute myocardial infarction and chronic heart failure. Toxic effects of this drug have been reported and above-normal cyanide and thiocyanate concentrations have been observed in the blood of a small proportion of subjects receiving nitroprusside. Nitrite, syanide, and thiocyanate are major decomposition products of nitroprusside, resulting from an in vitro reaction with human blood. On the basis of the conversion mechanism, we suggest that, in the cyanide/thiocyanate cycle, only cyanide is directly responsible for any acute toxicity attributed to sodium nitroprusside. In this work, the extent of cyanide production by erythrocytes in vitro was studied. The rate of detoxification of cyanide by human liver in vitro was experimentally determined and data from a search for a possible inhibitor of the nitroprusside/hemoglobin reaction are presented. Also, the possible mechanism of the nitroprusside/hemoglobin reaction is discussed.
Calorimetric enzymic measurement of uric acid in serum.
Authors: Rehak N N , Janes G , Young D S
Published: January 2020

Uric acid in serum was determined calorimetrically with a batch type microcalorimeter, by measuring the heat evolved during a coupled uricase/catalase enzymic reaction in tris(hydroxymethyl)aminomethane HCl buffer (pH 9.0 at 30 degrees C). Heat evolution and concentration are linearly related through the physiological range of serum uric acid concentrations and the method is free of interferences of the sort encountered with spectrophotometric methods. Precision and accuracy are good (CV, 2%) and the results correlate well with those obtained by a mechanized colorimetric uricase/peroxidase system.
Delta check tables for the Technicon SMA 6 continuous-flow analyzer.
Authors: Wheeler L A , Sheiner L B
Published: January 2020

To provide a data base for the establishment of a method to detect potentially incorrect SMA 6 test results, we analyzed 2970 pairs of SMA 6 results. For each test result, as well as for the urea nitrogen/creatinine ratio and the anion gap, we estimated to probability density function of the amount of change since the most recent test of the same type. We made separate estimates for each of three ranges of the current test result and two ranges of time-spacing between the tests. Tables of nonparametric estimates of selected points from each of the six distributions for each test result and calculated quantity were developed. These tables could be used to identify unlikely test results, either manually or with a laboratory computer.
Simultaneous spectrophotometry of Fe2+ and Cu2+ in serum denatured with guanidine hydrochloride.
Authors: Williams H L , Johnson D J , Haut M J
Published: January 2020

We show how iron and copper ions may be simultaneously measured in 0.2 ml of serum after using guanidine hydrochloride to cause their release from protein. Sensitive reagents for iron (2,4,6-tripyridyl-s-triazine) and copper (disodium 2,9-dimethyl-4,7-diphenyl-1, 10-phenanthroline disulfonate) facilitate such measurement. We eliminated spectral interference between the two metal-ion complexes by selective use of pH. Results compare well with those obtained by accepted methods in which the concentrations of these ions are measured independently.
Extraction and measurement of circulating angiotensins I and II.
Authors: Barrett J D , Eggena P , Sambhi M P
Published: January 2020

We describe a method for simultaneous extraction of angiotensins I and II from human blood. Blood was diluted with 10 volumes of isotonic saline containing ammonium ethylenediaminetetraacetate, and the angiotensin peptides were extracted by cation-exchange chromatography. The dilution effectively inhibits in vitro production of angiotensin I, conversion of angiotensin I to angiotensin II, and degradation of the peptides. Analytical recovery of exogenous angiotensin I or II added to whole blood in physiological concentrations was essentially complete. The extraction procedure removes blood and plasma components that interfere with radioimmunoassay of angiotensin, thus eliminating the need to correct for nonspecific interference. Circulating angiotensins I and II, measured in patients with essential hypertension before and after a week of diuretic administration, increased in parallel with plasma renin activity.
Biological variability in aspartate aminotransferase activity in serum of healthy persons, and effect of in vitro supplemantation with pyridoxal 5-phosphate.
Authors: Horder M , Bowers G N
Published: January 2020

Aspartate aminotransferase (EC 2.6.1.1) activities in sera from nine healthy individuals were monitored during two weeks, both with and without first supplementing the serum with pyridoxal phosphate. Pyridoxal phosphate supplementation caused a mean increase of 39% (range, 33-55%) in measured activity. The biological variability during the two-week period was independent of pyridoxal phosphate supplemantation. The intra-individual variability (CV) was 5.3% and 5.1% with and without pyridoxal phosphate supplementation, respectively; the corresponding inter-individual variability was 13.2% and 13.6%. We conclude that the reference interval will be insensitive to intra-individual fluctuations in aspartate aminotransferase activity in serum, whether or not the serum is supplemented with pyridoxal phosphate.
Differential serum amylase determination by use of an inhibitor, and design of a routine procedure.
Authors: O'Donnell M D , FitzGerald O , McGeeney K F
Published: January 2020

We describe a new method for measuring pancreatic and salivary-type amylases in serum that requires no electrophoresis or chromatography. An inhibitor protein (from wheat) with a 100-fold greater specificity for human salivary than for human pancreatic amylase was used to analyze mixtures of the two enzymes. The concentration of pancreatic and salivary amyalase was determined in 141 normal sera (72 men and 69 women). Statistically significant differences were found for serum pancreatic amylase between mean and women, higher values being shown in women. No sex-related difference was found for the salivary component of serum amylase. With this method, the increase in serum amylase activity in pancreatitis was shown to be attributable to the pancreatic component. In mumps, the increase is attributable to the salivary component. In pancreatic insufficiency, serum pancreatic amylase activities were significantly lower than in the controls. Our method is simple and rapid; our results agree well with those of other authors who used chromatographic or electrophoretic methods.
Micro-scale method for theophylline in body fluids by reversed-phase, high-pressure liquid chromatography.
Authors: Orcutt J J , Kozak P P , Gillman S A , Cummins L H
Published: January 2020

We describe a micro-scale method for determining serum theophylline. The chromatography system includes a muBondapack C18 column and acetonitrile, 70 ml/liter of sodium acetate buffer (10 mmol/liter, ph 4.0) as the mobile phase. Test serum or plasma, 30 mul, is mixed with an equal quantity of a solution containing the internal standard, beta-hydroxyethyltheophylline in acetonitrile/sodium acetate buffer (20 mmol/liter, pH 4.0), 7/43 by vol. After the precipitate is removed by centrifugation, the mixture is chromatographed and the amount of theophylline calculated from the ratio between peak heights for theophylline and the internal standard. Advantages include easy sample preparation, involving only addition of internal standard and centrifugation before injection, long column life, and the suitability of the internal standard, which is adjusted to a peak height equivalent to 20 mg of theophylline per liter for easy computation of results.
The Gallium Melting-Point Standard: A Determination of the Liquid—Solid Equilibrium Temperature of Pure Gallium on the International Practical Temperature Scale of 1968
Authors: Thornton Donald D
Published: January 2020

The sharpness and reproducibility of the gallium melting point were studied, and the melting temperature of gallium in terms of IPTS-68 was determined. Small melting-point cells designed for use with thermistors are described. Nine gallium cells including three levels of purity were used in 68 separate determinations of the melting point. The melting point of 99.99999% pure gallium in terms of IPTS-68 is found to be 29.7714 ± 0.0014 °C; the melting range is less than 0.0005 °C and is reproducible to ±0.0004 °C.
Acetaminophen in Serum and Plasma Estimated by High-Pressure Liquid Chromatography: A Micro-Scale Method
Authors: Blair Doyce , Rumack Barry H
Published: January 2020

We describe a capillary-sampling method for serum or plasma acetaminophen by cation-exchange chromatog-raphy. As little as 1.5 ill of plasma or serum and an equal volume of the internal standard (N-butyryl-p-aminophenol) were run, with a precision of ±5 % between duplicates. Acetaminophen and the internal standard chromato-graphed in 32 and 50 min, respectively, distinct from in-trinsic plasma peaks and peaks caused by other medica-tions.
Some problems associated with assay of 25-hydroxycalciferol in human serum.
Authors: Mason R S , Posen S
Published: January 2020

Methods available for the assay of 25-hydroxycalciferol in human serum are evaluated and compared to one another. Ethanol was chosen for use in the initial extraction procedure and rat-kidney cytosol as the binding protein, although good alternative methods are also available. We used silicic acid for chromatography and found this an essential step. Reproducibility was increased when, after "bound" and "free" material were separated, an aliquot of the supernate was pipetted into the counting vial instead of the entire supernatant fluid being decanted. Beta-lipo-protein added to the assay system was of no advantage; added bovine serum albumin interfered with the assay by giving rise to high blank values. With ethanol extraction, silicic acid chromatography, rat kidney cytosol and separation on dextran-coated charcoal, sera from normal subjects showed a mean 25-hydroxycalciferol concentration of 28.5 microng/liter (range, 13.1 to 43.9) during the fall season. Coefficients of variation for a control serum were 4.9% (intra-assay) and 10.9% (interassay).
Improved staining characteristics of serum lipids after halogenation and esterification of thin-layer chromatograms.
Authors: Nelson W R , Natelson S
Published: January 2020

We describe a procedure for treatment of thin-layer chromatographic serum lipid patterns so that they may be stained by dyes for evaluation by densitometry. After development of the chromatogram [Clin. Chem. 18, 384 (1972)] the plates are dried and sprayed with butyryl chloride. This esterifies the free cholesterol. After drying, the plate is treated with iodine monobromide, to add iodline to the double bonds. The triacylglycerols (triglycerides), cholesterol esters, free cholesterol, and phospholipids are now all in the form of esters with no unsaturated double bonds. The free fatty acids are also now saturated. The chromatogram is now stained with basic fuchsine in acetate buffer (0.1 mol/liter, pH 5.0). Excess stain is removed with a buffered solution of guanidine hydrochloride. Erythrosine B may also be used. With basic fuchsine the background will be a uniform pink. With erythrosine B the background is white, but the stain tends to be washed out of the free fatty acids. The chromatograms are evaluated by densitometry, with use of a 540-nm filter for basic fuchsine and a 520-nm filter for erythrosine B. The stained chromatograms and densitometric scans accurately represent the relative concentration of the various lipid fractions as compared to that of an internal standard, and correlate with the nature of the disease being explored.
Sequential-saturation-type assay for serum 25-hydroxyvitamin D.
Authors: Ellis G , Dixon K
Published: January 2020

We describe a method for measuring 25-hydroxyvitamin D in serum. The serum is denatured with ethanol, and the extract purified by chromatography on silicic acid to prevent potential interference from vitamin D and by other substances. 25-Hydoxyvitamin D is assayed by sequential saturation analysis in Tris-HCI buffer, pH 8.6, containing the surfactant Triton X-405 (2 ml/liter) to solubilize the steroid. Diluter, unpurifed normal human serum is used as the binder. The method is sensitive and precise. The dose-response curve is linear. The assay normally gives high count rates (12 000 to 20 000 cpm for the zero standard) and consequently 25-hydroxyvitamin D of low specific activity can be used if necessary. Serum 25-hydroxyvitamin D was determined in normal and osteomalacic subjects.
Computer-assisted quality control in clinical chemistry.
Authors: Sher P P
Published: January 2020

A MUMPS comuter program, which stores and retrieves quality-control data from all automated and manual work-station in the laboratory, has been developed as part of a laboratory information system. Tabular displays, Levey-Jennings charts, and summary statistics are available on a real-time basis. Significant economy over previous manual methods has been observed, and the total quality-control program in the laboratory has become a more active and timely process.
Triiodothyronine uptake assay with immobilized triiodothyronine antibody as the bound-free separating agent.
Authors: Coleman L H , LeMat B , Diel V , Barbieri A , Barrett J
Published: January 2020

Available systems for evaluation of degree of unsaturation of thyroxine-binding globulin are hampered at the bound-free separation step. In current assays, inorganic sorbants or ion-exchange resins are used to separate free 125I-labeled triiodothyronine from that bound to thyroxine-binding globulin. The techniques are laborious, time consuming, and not readily adaptable to total automation. In the assay we describe, triiodothyronine antibody, immobilized on the walls of polypropylene test tubes, is used as the bound-free separating agent in the evaluation of degree of unsaturation of thyroxine-binding globulin. The assay is simplified to four steps; washing, centrifugation, and use of columns are eliminated; and the procedure is readily automatable with existing pipetting equipment. Correlation with existing methods is excellent.
Furildioxime method for nickel analysis in serum and urine by electrothermal atomic absorption spectrometry.
Authors: Mikac-Dević D , Sunderman F W , Nomoto S
Published: January 2020

We describe a method of analysis for nickel in serum and urine. After the sample (1 ml) has been subjected to acid digestion, Ni(II) is extracted into methylisobutylketone as a furildioximate complex, and is measured by electrothermal atomic absorption spectrometry. The method is more sensitive and more convenient than previous methods, and is less subject to nickel contamination because the digestion and extraction steps are performed in a single tube. Analytical recovery of Ni(II) (10 or 20 microng/liter) added to 20 specimens of serum or urine averaged 101% (range, 94 to 107). The following values (mean +/- SD) were obtained for nickel concentrations in specimens of serum and urine from 19 healthy adults living in central Connecticut: serum = 3.1 +/- 1.6 microng/liter; urine = 2.7 +/- 1.6 microng/liter. The method provides a practical means for monitoring occupational exposures of workers to nickel compounds.
Binding of dye to albumin, as studied by ultrafiltration.
Authors: Yap W , Schaffer R
Published: January 2020

We describe a procedure for measuring the binding of small molecules to proteins by means of a dynamic ultrafiltration system. A continuous binding curve is obtained by continuously monitoring the absorbance of ultrafiltrate as a function of ultrafiltrate volume, and corrections are made for the retention of the small molecule by the membrane and the leakage of the protein through the membrane. The method is applied to the binding of bromcresol green to human albumin. Some major sources of uncertainties in the method are discussed.
Increased concentrations of serum free fatty acids falsely increase serum thyroxine as determined by competitive protein-binding.
Authors: Pannall P R , Swanepoel E , Bennett J M , Pauw F H
Published: January 2020

Markedly increased concentrations of free fatty acids after a fatty meal and heparin injection already have been shown to falsely increase thyroxine values measured by competitive protein-binding techniques, where ethanol extraction in used. We report here the effect of lesser increases. In 10 patients receiving heparin during hemodialysis we found significant increases in serum thyroxine by competitive protein-binding (mean, 20 nmol/l) and in free fatty acid concentrations (164 micronmol/l). Thyroxine measured as iodine did not change significantly. In seven subjects who had fasted for 27 h, we also noted significant increases in thyroxine by competitive protein-binding (mean 63 nmol/l) and in free fatty acid concentrations (624 micronmol/l). In a single subjects, serial measurements showed a close association of the two variables and the overall correlation between the increases was good. This artefact limits the value of many currently used thyroxine methods because it imposes restrictions on transport of samples and selection of patients.
Evaluation of a discrete samples/stopped-flow mixer system for equilibrium and kinetic analyses.
Authors: Pardue H L , Gaw H T , Mieling G E , Mathews V L , Fast D M , Rubesch M A
Published: January 2020

This paper describes the evaluation of a system for computer-controlled discrete sampling and stopped-flow mixing for equilibrium and kinetic determinations of several sorts of analytes in human serum. The instrumental system features a wash-out sampling system that permits rapid change-over from one sample and (or) reagent type to another, and a mixing-measurement system that can provide reliable data as soon as 10 ms after reagent and sample are mixed. Examples discussed include equilibrium procedures for glucose and cholesterol, slow kinetic procedures for glucose and lactate dehydrogenase, and a fast kinetic method for thiocyanate. The regression equation for all stopped-flow results (n = 114) vs. results by conventional methods is y = (103 +/- 0.01)x - (0.016 +/- 0.019) for numerical values of y between 0.3 and 3.0. The correlation coefficient for these data was 0.991. These results demonstrate that the stopped-flow method is a viable analytical approach for equilibrium, slow kinetic, and fast kinetic determinations that require measurement times shorter than 0.1 s.
Analysis for platinum in biological material by flameless atomic absorption spectrometry.
Authors: Pera M F , Harder H C
Published: January 2020

A method for determining platinum in biological materials by flameless atomic absorption spectrometry has been developed for use in pharmacological studies of platinum antitumor compounds. Tissue digests are analyzed in a heated graphite atomizer, with use of background correction and ramp temperature charring. We have overcome problems of apparent loss of platinum in Pt-supplemented tissue samples with time and of difficulty in analyzing tissue lipid. The validity of the assay was verified by injecting rats with radiolabeled cis-dichlorodiammineplatinum (II), then measuring platinum in tissues by gamma scintillation spectrometry and flameless atomic absorption spectrometry. Correlation between the two methods was excellent. The limit of the assay in its present form is about 0.1 migrogram/gram of wet tissue.
Continuous-flow determination of dialyzable calcium in serum.
Authors: Toffaletti J , Savory J , Gitelman H J
Published: January 2020

We describe a procedure for measuring dialyzable calcium in 60 sera per hour with use of AutoAnalyzer (Technicon) modules. The serum is buffered, then dialyzed in a regular 6-inch (15-cm) dialysis block. A constant proportion of the dialyzable calcium is fluorometrically measured. Analysis of samples containing protein, prepared by an equilibrium dialysis technique, shows that protein-bound calcium is not detected. The method requires less than 0.4 ml of serum, and is accurate, precise, and durable. No unusual care or special techniques are required during sample collection or analysis.
Determination of urinary normetanephrine, metanephrine, and 3-methoxytyramine by liquid chromatography, with amperometric detection.
Authors: Shoup R E , Kissinger P T
Published: January 2020

We describe a sensitive procedure for measuring the basic catecholamine metabolites--normetanephrine, metanephrine, and 3-methoxytyramine--in urine by use of liquid chromatography, with electrochemical detection. These substances are isolated from hydrolyzed urine by passage through small ion-exchange columns and then pre-concentrated by a rapid set of solvent extractions before being injected onto the liquid-chromatographic column. Concentration and instrumental response are linearly related to 2.0 mg/liter for all three compounds, and practical lower detection limits are about 20 micrograms/liter for actual urine samples. The high selectivity that accrues from the combination of the extraction procedure, high-performance liquid chromatography, and electrochemical detection makes this procedure suitable for quantitative studies of catecholamine metabolism. A set of six samples can be taken through the extraction procedure in parallel in less than 2 h; once prepared, the extracts can be analyzed at a rate of three per hour.
Continuous-flow determination of serum inorganic phosphate with a single reagent--the vanadomolybdate method re-evaluated.
Authors: Baadenhuijsen H , Seuren-Jacobs H E , Jansen A P
Published: January 2020

The merits of the vanadomolybdate method in the determination of inorganic phosphate are highly underestimated with regard to the simplicity of the method and the stability of both the reagents and the color that is formed. The absorption curve of the phosphovanadomolybdate complex, with its maximum at 335 nm, extends into the visible range of the spectrum. This permits measurements with inexpensive tungsten-lamp colorimeters. On-stream dialysis is best done in a nitric acid medium, 0.15 mol/liter. Paralleled by the change in the H2PO4-/H3PO4 ratio, appreciable protein binding and poor dialysis efficiency are seen at lower acid concentrations (pH greater than 1.0). Optimum reaction-mixture concentrations of vanadium and molybdenum appeared to be respectively 0.2 and 5 mmol/liter up to 3 mmol of phosphate per liter, in a final acid medium of 0.2 mol/liter, concentrations considerably lower than those used in some studies published earlier. Color development with the stable combined reagent is complete after only 20 s at room temperature and the color is stable for at least 2 h. Figures on precision and accuracy demonstrate the reliability of the method.