bluuebunny/crossref_metadata_2025 huggingface dataset:
Concurrent liquid-chromatographic assay of retinol, alpha-tocopherol, beta-carotene, alpha-carotene, lycopene, and beta-cryptoxanthin in plasma, with tocopherol acetate as internal standard.
Authors: Thurnham D I , Smith E , Flora P S
Published: January 2020

A method is described for simultaneously determining retinol, alpha-tocopherol, beta-carotene, alpha-carotene, lycopene, and beta-cryptoxanthin in 0.25 mL of plasma. Plasma mixed with sodium dodecyl sulfate is deproteinized with ethanol containing tocopherol acetate, then extracted with heptane. The evaporated organic layer is reconstituted with mobile phase (methanol/acetonitrile/chloroform, 47/47/6 by vol) and injected onto a 100 x 4.6 mm 3-micron column of Spherisorb ODS-2 (LKB) at 1.5 mL/min. The alpha- and beta-carotenes are well resolved during the 6.5-min run. Retinol is monitored at 325 nm, the tocopherols at 292 nm, and the carotenoids at 450 nm. Extraction of concentrations as great as 135 mumol/L is complete. Intrabatch CVs were 1.7%, 2.3%, 4.1%, 10.4%, 6.4%, and 3.6% for retinol, alpha-tocopherol, beta-carotene, alpha-carotene, lycopene, and beta-cryptoxanthin, respectively. Interbatch CVs for measurements on 30 occasions over 11 weeks were about 10% for all components except alpha-tocopherol (5.3%). Results agree well with those for retinol, alpha-tocopherol, and beta-carotene in quality-control samples.
Improved assay for bismuth in biological samples by atomic absorption spectrophotometry with hydride generation.
Authors: Froomes P R , Wan A T , Harrison P M , McLean A J
Published: January 2020

This simple, rapid, sensitive, reliable, and economical assay for bismuth in plasma, erythrocytes, and urine is based on atomic absorption spectrophotometry with hydride generation. Acid digestion eliminates the problem of foaming, which hitherto has complicated such assay of bismuth in plasma and erythrocytes. The detection limit of the assay has been improved to 0.1 micrograms/L, as compared with a previously documented limit of 2.5 micrograms/L. Average recovery exceeded 95% in all biological fluids. Economy of use derives from elimination of need for electrodeless discharge lamps and atomic absorption grade borohydride. Determination of basal concentrations of bismuth in clinical samples of body fluids gave reference intervals of 0.1-3.5 micrograms/L for plasma, 0.3-4.6 micrograms/L for urine.
A dry-reagent strip for quantifying carbamazepine evaluated.
Authors: Croci D , Nespolo A , Tarenghi G
Published: January 2020

We examined a new colorimetric homogeneous immunoassay for carbamazepine based on the apoenzyme reactivation immunoassay system (ARIS) principle. The test, in dry-reagent strip format, is to be used with the Ames Seralyzer reflectance photometer. Within-run CVs (n = 20) were 3.0%, 2.7%, and 2.8% at 3.0, 6.1, and 12.1 mg/L; between-run CVs (n = 15, in 15 days) were 4.1%, 2.7%, and 1.9% at 6.0, 9.1, and 12.1 mg/L. Mean analytical recovery was 99.9 (SD 2.3)%. Results by this test (y) for clinical plasma specimens (n = 96) compared very well with those obtained by fluorescence polarization immunoassay (y = 1.01 x - 0.02, r = 0.995) and by liquid chromatography (y = 0.99 x + 0.14, r = 0.990). Bilirubin (45 mg/L), uric acid (145 mg/L), and various anticoagulants at about fourfold the usual concentrations did not interfere. High concentrations of cholesterol (4.9 g/L), triglycerides (3.8 g/L), and hemoglobin (4 g/L) caused slight positive interference. Carbamazepine-10,11-epoxide cross reacted only at greater than or equal to 5 mg/L. The two-point calibration line was validly stored for at least three weeks. Free carbamazepine also can be measured. The test is convenient and rapid (test time 80 s), and thus is particularly useful in all clinical settings where prompt testing is needed.
Intact parathyrin in postmenopausal women.
Authors: Sokoll L J , Morrow F D , Quirbach D M , Dawson-Hughes B
Published: January 2020

To establish a reference range, we measured intact parathyrin (parathyroid hormone, PTH) in 245 healthy postmenopausal women, ages 42-75 years, with use of the Allegro Intact PTH Kit from Nichols Institute Diagnostics. We also assayed serum from a subset of 120 of the women with kits specific for mid-molecule PTH. The mean intact PTH concentration for the 245 women was 32 ng/L (95% confidence interval 14-60 ng/L). Intact PTH values in these subjects were not normally distributed, although calcium concentrations in the same samples were. There was positive, but not significant (r = 0.12, P = 0.06), correlation between intact PTH and age, and a significant negative correlation between serum calcium and intact PTH that was not observed between calcium and mid-molecule PTH. The improved sensitivity of the intact PTH assay makes it useful in studies of calcium homeostasis in the normal population.
Immunoassay of low concentrations of albumin in urine by latex particle counting.
Authors: Cambiaso C L , Collet-Cassart D , Lievens M
Published: January 2020

We describe here a nonisotopic immunoassay, based on particle-counting technology, for the determination of urinary albumin. The assay takes only 35 min and has been fully automated on the IMPACT (Acade Diagnostic Systems, Brussels, Belgium) machine. The system measures albumin within a linear range between 6.25 and 50 mg/L and has a detection limit of 0.4 mg/L. Analytical recoveries at three concentrations ranged between 96% and 102%. Within-run precision ranged from 1.6% to 9.5%. The method was compared with a commercial nephelometric immunoassay system and a correlation coefficient of 0.996 was found for 216 urine samples. No antigen excess affects the shape of the curve in our system, whereas in nephelometry a 3 g/L solution of albumin starts to decrease the dose-response curve.
Stability and electrophoretic characteristics of creatine kinase BB extracted from human brain and intestine.
Authors: Chastain S L , Ketchum C H , Grizzle W E
Published: January 2020

Creatine kinase (CK; EC 2.7.3.2) isoenzyme BB extracted from brains of rats reportedly undergoes modification at 37 degrees C, leaving an electrophoretic variant that accounts for most of the residual CK activity. This variant, called CK-BB', migrates on electrophoresis similarly to creatine kinase isoenzyme MB. Using electrophoresis and immunoinhibition with antiserum to creatine kinase isoenzyme MM, we found CK-BB to be the only identifiable cytoplasmic isoenzyme in surgical samples from human brain and intestine. In contrast, we found that some samples of brain obtained at autopsy contain CK-BB'. We also found that CK-BB extracted from human brain was converted to CK-BB' upon incubation in serum or plasma at 37 degrees C. We found a similar development of CK-BB' in incubation mixtures of serum or plasma containing CK-BB obtained from surgical samples of human intestine. The development of CK-BB' during infarction of the gastrointestinal system may thus be a source of false-positive CK-MB in the laboratory verification of myocardial infarction when electrophoresis is used as the only method to identify CK isoenzymes.
Liquid-chromatographic assay of ibuprofen enantiomers in plasma.
Authors: Mehvar R , Jamali F , Pasutto F M
Published: January 2020

This stereospecific "high-performance" liquid-chromatographic (HPLC) assay is suitable for pharmacokinetic studies of ibuprofen (IB). Very efficient extraction of the drug and internal standard, (+/-)-2-(4-benzoylphenyl)butyric acid, from plasma with isooctane/isopropanol (95/5, by vol) is followed by sequential reaction of the enantiomers with ethyl chloroformate and (S)-(-)-1-(1-naphthyl)ethylamine. The reactions take place at ambient temperature in less than 4 min. The naphthylethylamide derivatives of IB enantiomers and internal standard are then extracted into chloroform. After the organic layer is evaporated, the reconstituted residue is chromatographed at ambient temperature on a C18 reversed-phase column with a mobile phase of acetonitrile/water/acetic acid/triethylamine (55/45/0.1/0.02 by vol) at a flow rate of 1 mL/min. The IB diastereoisomers, detected at 232 nm, are free of interfering peaks and have a resolution factor of 2.2. Within the examined enantiomer concentration range of 0.1 to 20 mg/L in plasma, the peak-area ratios varied linearly with the corresponding IB concentrations. We used the assay to study the pharmacokinetics of IB enantiomers in plasma of a subject who took a single 600-mg dose of racemic drug.
Quinidine-induced decrease of intracellular esterase activity in a hepatoma cell line.
Authors: Markovic O T , Young D S , Markovic N S
Published: January 2020

Using image-analyzing equipment, we measured the effect of quinidine on the activity of naphthol AS-D chloroacetate esterase, alpha-naphthyl butyrate esterase, alpha-naphthyl acetate esterase, and acid phosphatase in individual cells of a human Hep G2 hepatoma cell line. The impact of the drug on the morphology of the cells was also observed. Depending on the concentration of quinidine applied, various changes occurred, the most extreme being cell death. However, at some drug concentrations that did not appear to affect visible cell structures, the activity of the esterases was decreased. This lessened enzyme activity did not seem to be related to the enzymes leaking from the cells, because the activity of acid phosphatase was unaffected. Inhibition of the esterase activity was related to the interval of exposure to quinidine in the perfusing medium and to the concentration of the drug. We consider the system described here to be a potential replacement for experiments with animals in the study of hepatotoxicity.
A new enzymatic assay of chloride in serum.
Authors: Ono T , Taniguchi J , Mitsumaki H , Takahata F , Shibuya A , Kasahara Y , Koshimizu F
Published: January 2020

This new method for enzymatic assay of serum chloride is based on determination of chloride-dependent alpha-amylase (EC 3.2.1.1) activity. We evaluated the ion specificity and practicability of the method for routine use with the Hitachi 705. The analytical range of the method extends from 40 to 160 mmol of chloride ion per liter of serum. The reaction rate for samples containing 100 mmol of chloride per liter was 0.17 A/min. CVs within-run and between-run were less than 1.0%. Correlation with results of a coulometric titration method was good. The specificity for bromide is 75% of that for chloride; for the other anions, it is 0%. This enzymatic method is generally applicable to wide variety of automated chemistry analyzers.
A high-resolution method for amino acid analysis of physiological fluids containing mixed disulfides.
Authors: Lee P L , Slocum R H
Published: January 2020

In this paper we report a high-resolution method and special buffers for the analysis for 45 common amino acids and related compounds and seven less-common components occasionally found in physiological fluids. Careful optimization of the chromatographic protocol resolves cysteine-penicillamine mixed disulfide, penicillamine disulfide, and dihydroxyphenylalanine from cystine and methionine; cysteine-homocysteine mixed disulfide is eluted between tyrosine and phenylalanine. Argininosuccinic acid is eluted before ethanolamine; glycerophosphoethanolamine is separated from taurine; glucosaminic acid is eluted between urea and aspartic acid. A Beckman Instruments System 6300 Amino Acid Analyzer was used. System performance is examined in terms of coefficient of variation (CV) over the concentration range of 1.0-5.0 nmol/50 microL; average CV for all components is better than 1.5%. Method performance is assessed through the analysis of normal and abnormal urine and plasma specimens.
Factors influencing fluorescence spectra of free porphyrins.
Authors: Polo C F , Frisardi A L , Resnik E R , Schoua A E , Batlle A M
Published: January 2020

We recorded fluorescence excitation and emission spectra of uro- and coproporphyrin under different experimental conditions, to see how these conditions influence quantifications based on measurement of fluorescence intensity. We found that, for bands alpha and beta of the emission spectra and the main peak of the excitation spectra, fluorescence depends on pH and is minimal near pH 5 and near pH 7-7.5 for copro- and uroporphyrin, respectively. For band gamma of the emission spectra there was a constant decrease of fluorescence with increasing alkalinity of the solution. The intensity of porphyrin fluorescence also depends on ionic strength, reaching sharp maxima at 0.1 mol/L (for uroporphyrin) and 1 mol/L (for coproporphyrin). The organic mixture ethyl acetate:acetic acid (4:1 by vol), commonly used to extract porphyrins from biological samples, markedly diminishes the fluorescence of both porphyrins as compared with the same concentration of each porphyrin in aqueous acidic solvent. Furthermore, when we measured different ratios of uro:copro mixture at three distinct pHs and buffers, we found that at pH 10.5 (in carbonate buffer) the measured units of fluorescence depend only on total porphyrin concentration and not on the composition of the mixture.
Alpha 1-antichymotrypsin microheterogeneity in crossed immunoaffinoelectrophoresis with free concanavalin A: a useful diagnostic tool in inflammatory syndrome.
Authors: Hachulla E , Laine A , Hayem A
Published: January 2020

Using crossed immunoaffinoelectrophoresis with free concanavalin A (Con A) in the first dimension and alpha-methyl-D-glucoside incorporated in the second-dimension gel, we examined the microheterogeneity of alpha 1-antichymotrypsin (alpha 1Achy) in sera from healthy donors (N) and in sera from patients with various inflammatory syndromes. We studied three groups: patients with acute inflammation (myocardial infarction: MI, septic inflammation: SI), patients with chronic inflammation (metastatic breast cancer: MBC), and patients with chronic inflammation accompanied by acute attacks (connective-tissue disease: CTD). All pathological sera had a high alpha 1Achy concentration. Compared with N, MI and SI showed an increased proportion of Con A-reactive fraction and a decreased proportion of Con A-nonreactive fraction, which was more pronounced in SI. Unlike the patients with acute inflammation, patients with CTD showed an increased proportion of Con A-nonreactive fraction. Thus alpha 1Achy microheterogeneity in crossed immunoaffinoelectrophoresis may afford a means of differentiating between various inflammatory syndromes. In particular, it can provide a simple test: if the Con A-nonreactive fraction is in a proportion less than 17%, septic origin of an acute-phase reaction may be suspected.
Rapid, simplified radioimmunoassay of arginine-vasopressin and atrial natriuretic peptide in plasma.
Authors: Bodola F , Benedict C R
Published: January 2020

We have improved a radioimmunoassay for arginine-vasopressin (AVP) and atrial natriuretic peptide (ANP) by using Sep-Pak C18 cartridges to extract AVP and ANP from acidified plasma. The analytes are co-eluted by use of a mobile phase consisting of 1,2-dimethoxyethane and 40 g/L aqueous trifluoroacetic acid (95/5, by vol). After rapid evaporation of the solvents, AVP and ANP are assayed by a nonequilibrium radioimmunoassay method in which commercially available antibodies and radiolabeled antigens are used. The bound fractions are separated from the free by use of polyethylene glycol with human gamma globulin and rabbit anti-human IgG as the second antibody. This results in very low nonspecific binding: 0.44% for the ANP assay, 0.70% for AVP. The minimum detectable amount of ANP is 0.39 pg per tube; for AVP, it is 0.13 pg per tube. Compared with other published methods, this method is substantially more reliable, economical, and easily established in a clinical chemistry laboratory.
Secretion of atrial natriuretic peptide and digoxin-like immunoreactive substance during pregnancy.
Authors: Shrivastav P , Gill D S , D'Souza V , O'Brien P M , Dandona P
Published: January 2020

We investigated the secretion of atrial natriuretic peptide (ANP) and digoxin-like immunoreactive substance (DLIS) during pregnancy, labor, and the puerperium, as measured in maternal and umbilical cord plasma. There were no significant changes in maternal concentrations of ANP during all three periods, and the concentrations were similar to those found in nonpregnant controls and in umbilical cord plasma. Maternal concentrations of DLIS increased significantly in the second half of pregnancy, peaked during labor, then decreased abruptly within 24 h of expulsion of the infant and placenta to values approaching the nonpregnant range. DLIS concentrations in umbilical cord plasma, however, were significantly higher than in maternal plasma during labor. The abrupt fall in DLIS in maternal plasma and the absence of a significant difference in DLIS concentrations between arterial and venous cord plasma suggest that, during pregnancy, the fetus, not the placenta, is the source of DLIS in maternal plasma.
Measurement and characterization of angiotensin peptides in plasma.
Authors: Hermann K , Ganten D , Unger T , Bayer C , Lang R E
Published: January 2020

We report a method for the extraction of angiotensin peptides from plasma with a mixture of acetone, 1 mol/L HCl, and water (40/1/5 by vol). The method is highly reproducible for the measurement of angiotensin I and angiotensin II in small sample volumes, with analytical recoveries of about 80% for both peptides. We investigated the influence of sample handling and found a standard procedure for blood collection, plasma preparation, and extraction was essential. The method was used to measure angiotensin I and II in rat and human plasma. In rat plasma, the mean (+/- SEM) concentrations of angiotensin I and angiotensin II were determined to be 67 (+/- 8) and 14 (+/- 1) pmol/L (n = 10), respectively. Neither angiotensin I nor angiotensin II was detectable 24 h after bilateral nephrectomy. Acute oral administration of the converting-enzyme inhibitor ramipril caused a significant increase of angiotensin I from 85 (+/- 6) to 257 (+/- 33) pmol/L (n = 10; P less than 0.001) and a significant decrease of angiotensin II from 12 (+/- 1) to 7 (+/- 0.4) pmol/L in rat plasma (n = 9; P less than 0.001). In human plasma, angiotensin I and angiotensin II values of 21 (+/- 1) and 6.6 (+/- 0.5) pmol/L (n = 10) were found. A single oral dose of the diuretic furosemide increased angiotensin I significantly from 21 (+/- 1) to 32 (+/- 1.7) pmol/L (n = 5); P less than 0.001), whereas angiotensin II remained unchanged, 6.6 (+/- 0.5) vs 6.4 (+/- 0.4) pmol/L (n = 5). Extracted peptides could be identified as [IIe5]-angiotensin I and [IIe5]-angiotensin II by HPLC in combination with specific radioimmunoassays for angiotensin I and angiotensin II.
Nucleotides, nucleosides, and oxypurines in human kidneys measured by use of reversed-phase high-performance liquid chromatography.
Authors: Maessen J G , van der Vusse G J , Vork M , Kootstra G
Published: January 2020

An HPLC technique is presented for determining adenine nucleotides and related substances in renal cortical tissue. Nineteen metabolic substances can be resolved in a single 25-min run, with use of a gradient-elution system. The mean intra-assay CV is 2.4%, the interassay CV 5%. The lower detection limit for substances commonly present in kidney tissue--such as ATP, ADP, AMP, GTP, GDP, GMP, IMP, inosine, adenosine, hypoxanthine, and xanthine--ranges from 0.6 to 3.6 mumol/L, corresponding to 18 and 107 pmol applied to the column. For reliable analysis, a specimen of renal cortex weighing at least 5 mg (wet weight), taken during donor nephrectomy, during cold storage of the kidney, and 1 h after the onset of reperfusion, can be used. The method presented provides a rapid, reproducible diagnostic tool for assessing the chemical energy status of human kidneys in renal surgery and transplantation.
Monoclonal antibodies for radioimmunoassay of cyclosporine: a multicenter comparison of their performance with the Sandoz polyclonal radioimmunoassay kit.
Authors: Holt D W , Johnston A , Marsden J T , Vernillet L , Keown P A , Rosano T G , Shaw L M , Rosenthaler J
Published: January 2020

The performance of a radioimmunoassay kit containing monoclonal specific and nonspecific antibodies to cyclosporine (Sandimmun-Kit; Sandoz Ltd., Basle, Switzerland) was compared with that of the original Sandoz polyclonal radioimmunoassay kit (Ciclosporin RIA-Kit). A total of 1320 blood and plasma samples from patients receiving cyclosporine after kidney, heart, liver, and bone-marrow transplantation were analyzed at six centers. For blood samples the median result on using the specific assay was about 50% of the polyclonal assay result after kidney and bone-marrow transplantation, about 33% after heart and liver transplantation; comparable figures for plasma samples were 70 and 40%. The monoclonal nonspecific-antibody assay produced results 10% to 140% higher than polyclonal-assay results, depending on sample matrix and transplant indication; the largest difference was seen in samples from heart- and liver-transplant recipients. Evidently the specific-antibody assay provides a convenient alternative to high-performance liquid chromatography for specific measurement of the drug, but the role of the new nonspecific antibody, possessing an even broader spectrum of cross-reactivity with cyclosporine metabolites than the original polyclonal antiserum, has yet to be defined.
Atypical patterns of lactate dehydrogenase isoenzymes in acute myocardial infarction.
Authors: Rotenberg Z , Weinberger I , Davidson E , Fuchs J , Sperling O , Agmon J
Published: January 2020

Total lactate dehydrogenase (LD; EC 1.1.1.27) activity in serum and LD isoenzymes were quantified in 190 patients with acute myocardial infarction (AMI) 24, 48, and 72 h after admission. In 90% of the 570 blood specimens an LD isoenzyme pattern typical of AMI (LD-1/LD-2 greater than 0.76) was found. The other 56 blood specimens showed an LD isoenzyme pattern atypical of AMI (LD-1/LD-2 less than 0.76). They were divided into three groups: 28 specimens with isomorphic pattern (relative increase in all five LD isoenzymes); 18 with relatively increased LD-3 proportion (greater than 35%); and 10 specimens with increased LD-5 proportion (greater than 10%). No difference was found in mean total LD activity in serum between the typical isoenzyme group and the three atypical groups. The LD isomorphic pattern was found in 60% of AMI patients complicated by cardiogenic shock. Fifty percent of AMI patients admitted with pulmonary edema showed increased LD-3 proportion and half of the patients with AMI and congestive heart failure, predominant right, demonstrated increased LD-5 proportion. We conclude that although most patients with AMI present at diagnosis with a typical LD isoenzyme pattern, it is important to recognize that some may present with atypical LD isoenzyme patterns, which may be associated with specific AMI complications.
Early detection of skeletal muscle injury by assay of creatine kinase MM isoforms in serum after acute exercise.
Authors: Apple F S , Hellsten Y , Clarkson P M
Published: January 2020

We could detect skeletal muscle injury early after an acute exercise bout by measuring creatine kinase (CK, EC 2.7.3.2) MM isoforms in serum. Eleven men performed 120 alternating-arm, eccentric (muscle lengthening) biceps contractions with the intensity of each contraction being 110% of maximal concentric strength--a form of exercise previously shown to cause significant increases of CK in serum at 24 h and muscle soreness 48 h after exercise. Total CK and CK-MM isoform activities in serum were determined before and at 0.5, 0.75, 1, 1.5, 2, and 6 h after exercise. Using thin-film agarose gels and a rapid isoelectric focusing technique, we separated the MM isoforms into MM3 (skeletal muscle form), MM2, and MM1 (in vivo conversion forms). The isoforms reflected the MM form released into the serum from tissue as well as the conversion of one form to another. There were no significant increases in total CK from before to 6 h after exercise: 75 (SD 36) vs 91 (SD 33) U/L. However, CK MM3 in serum increased significantly (P less than 0.01) within 2 h after exercise from 22 (SD 6)% to 28 (SD 6)%. The MM3 to MM1 ratio also increased significantly (P less than 0.05) during this time, from 0.6 (SD 0.3) to 0.9 (SD 0.4). Thus, quantification of CK MM isoforms permitted very early detection of skeletal muscle enzyme release.
Urinary lipoperoxides quantified by liquid chromatography, and determination of reference values for adults.
Authors: Knight J A , Smith S E , Kinder V E , Pieper R K
Published: January 2020

Urinary lipoperoxides, measured as the malondialdehyde-thiobarbituric acid adduct, were quantified by adapting to urine the liquid-chromatographic method of Wong et al. (Clin Chem 1987;33:214-20) to plasma. Reference intervals for untimed urine specimens from 121 men, ages 16 to 67 years, and 107 women, ages 15 to 55, were determined. Their concentrations differed significantly (P = 0.015), males having a mean (and SD) of 0.89 (0.35) nmol of malondialdehyde per milligram of creatinine, females 0.78 (0.30). In both groups, the values were slightly skewed to the higher values. Our early studies suggest that measuring urinary lipoperoxide may have advantages over plasma in studying certain disorders. The presence of other urinary chromophores or TBA-reactive substances stresses the need for chromatographic techniques when lipoperoxides are measured in biological samples.
An enzyme immunoassay compared with a ligand-binding assay for measuring progesterone receptors in cytosols from breast cancers.
Authors: Smyth C M , Benn D E , Reeve T S
Published: January 2020

To assay progesterone receptor (PR), we compared Abbott's enzyme immunoassay (PR-EIA) with a ligand-binding assay involving dextran-coated charcoal (PR-DCC), using cytosols prepared from 109 breast-cancer biopsies. Results by the two PR methods agreed well. Least-squares analysis produced a line of best fit having a slope of 0.88, an intercept on the PR-EIA axis of 16 fmol per milligram of protein, and a correlation coefficient (r2) of 0.87. To evaluate whether accurate PR-EIA measurements could be obtained on stored cytosols, we compared PR-EIA values for fresh cytosols with values for cytosols stored for various lengths of time up to 13 weeks. Agreement was excellent, especially when the samples showing very high binding (greater than 600 fmol per milligram of protein) were excluded. The lines of best fit after least-squares analyses of the remaining values had slopes between 1.0 and 1.1, intercepts less than 3 fmol/mg, and r2 all greater than 0.91.
Multi-wavelength analyses of second-derivative spectra for rapid determination of acetaminophen in serum.
Authors: Dingeon B , Charvin M A , Quenard M T , Thome H
Published: January 2020

Measurement of acetaminophen by analysis of the second derivative of its spectrum is specific and sensitive. The method of extraction and the use of just one phosphate buffer as reagent makes this method very convenient. Readings are reliable from 10 to 1500 mg/L. A turnaround time of 20 min makes this method well suited for emergency cases. Precision and accuracy of the method are presented. Results are not biased by interferences, not even from N-acetylcysteine.
Alanine aminopeptidase in serum: biological variations and reference limits.
Authors: Sanderink G J , Artur Y , Schiele F , Gueguen R , Siest G
Published: January 2020

We studied the effect of various factors on the biological variation of alanine aminopeptidase (AAP, EC 3.4.11.2) in serum in a population of 2178 apparently healthy subjects and in subjects taking specific drugs. To measure AAP, we used an automated assay, with alanyl-4-nitroanilide as substrate. AAP activity concentrations were significantly higher in males than in females in all age groups between 10 and 55 years. The highest mean AAP values were found for children between 10 and 14 years. In male heavy smokers, AAP values were 8% higher than in moderate or nonsmokers (P less than 0.001); in females, this effect was less pronounced. AAP concentrations were higher in subjects consuming over 44 g of ethanol a day than in those consuming less (P less than 0.01 for men, P less than 0.05 for women). The use of oral contraceptives increased AAP values by 12% (P less than 0.001). Hypolipidemic drugs caused lower values for AAP in men (P less than 0.001). Subjects showing induction of gamma-glutamyltransferase (EC 2.3.2.2) by anticonvulsant drugs had also higher AAP activities (by 23%). Taking these data into account, we established reference limits for AAP in serum.

The aim of this study was to describe the major factors producing variation in levels of serum alanine aminopeptidase. The study population consisted of 1065 men and 1113 women screened at the Center for Preventive Medicine in Vandoeuvre-les-Nancy in 1987. Mean values were significantly higher for men than for women between age 10 and age 55. Alanine aminopeptidase activity was higher in prepubertal children than in adults and decreased with age, but increased again in postmenopausal women. Overweight status had no effect, nor did systolic blood pressure below 160 mmHg. Serum levels fell by 10% for men over this limit, but increased by 15% for women above this limit. Enzyme levels rose with alcohol consumption for both men and women. Men who were heavy smokers (over 20 cigarettes/day) had elevated levels of serum alanine aminopeptidase. In women oral contraceptives and antihypertensive drugs increased enzyme levels. Mean levels increased by 23% in both men and women taking anticonvulsant drugs who also had increased gamma-glutamyltransferase activity. Factors affecting serum alanine aminopeptidase levels, in decreasing order of importance, were sex, age, alcohol consumption, and cigarette smoking.
Effect of ethanol on the ratio between testosterone and epitestosterone in urine.
Authors: Falk O , Palonek E , Björkhem I
Published: January 2020

The testosterone/epitestosterone weight ratio in urine is used to detect cases of doping when an athlete has treated himself with exogenous testosterone. When this ratio exceeds 6, it is considered evidence of testosterone doping. We show here that intake of ethanol can affect this ratio. Ingestion of 110-160 g of ethanol, about 2 g per kilogram body weight, increased the ratio between testosterone and epitestosterone in urine from 1.14 +/- 0.07 to 1.52 +/- 0.09 in four healthy male volunteers. The increase ranged from 30% to 90% in the different subjects studied (mean 41%). In cases where doping with testosterone is suspected, the possibility should be considered that at least part of an observed increased testosterone/epitestosterone ratio in urine is ascribable to previous ingestion of ethanol.
The role of precision in determining the performance of a thyrotropin assay in diagnosing hyperthyroidism.
Authors: Wilke T J , Utley D J
Published: January 2020

We examined the relationship between analytical sensitivity, precision at the lower limit of the reference interval, and diagnostic performance in hyperthyroidism for one radioimmunoassay and five immunometric assay kits for thyrotropin. The analytical sensitivity of these kits extended from 0.05 to 1.56 milli-int. units/L. Diagnostic efficiencies of the immunometric assays, in discriminating between euthyroidism and hyperthyroidism, ranged between 93% and 98%. There was a highly significant correlation (r = 0.99, P less than 0.001) between analytical sensitivity and diagnostic efficiency. The between-assay coefficients of variations, at the lower limit of the reference interval, ranged from 26% to 87%. There was no correlation (r = 0.36) between precision, at this concentration, and diagnostic efficiency. We conclude that analytical sensitivity and not precision is the major determinant in controlling the diagnostic performance of a thyrotropin assay in hyperthyroidism.
Screening for neutral and basic drugs in blood by dual fused-silica column chromatography with nitrogen-phosphorus detection.
Authors: Turcant A , Premel-Cabic A , Cailleux A , Allain P
Published: January 2020

We describe a capillary gas-chromatographic method for detection and quantification of basic and neutral drugs in the plasma of patients thought to be poisoned after dangerous overdose. Without further derivatization, the drugs are extracted from 1 mL of plasma, at basic pH, into diethyl ether. The extracts are injected onto two fused-silica capillary columns of different polarity (Ultra 1 and CP Sil 19 CB) coupled to nitrogen-phosphorus detectors. Under these conditions, drug-free plasmas give blank chromatograms, with a peak only for the internal standard (RN 927, an antihistamine not being marketed). Plasma samples from patients who have taken drugs show additional peaks, the relative retention times (RRTs) of which are used to identify the drugs. Here we list the RRTs of about 200 drugs on the two columns. Analyses are routinely performed with an automatic injector; overall analysis time is about 1 h per sample. During the last six years, more than 1000 plasma samples per year have been analyzed. We find this method a powerful tool for toxicological analysis, especially in cases of multi-drug intoxications.
Lithium determined in serum with an ion-selective electrode.
Authors: Bertholf R L , Savory M G , Winborne K H , Hundley J C , Plummer G M , Savory J
Published: January 2020

We evaluated the performance of the lithium ion-selective electrode (ISE) in the Du Pont Na/K/Li analyzer. Lithium concentrations in 106 serum samples from patients being treated with lithium were measured in duplicate with the ISE and by flame photometry. The slope of the regression line for the two methods was 1.004 with a standard error of the estimate of 0.049 mmol/L (x = flame photometry, y = ISE). Lithium measurements by the ISE method in serum or aqueous standards were linear to greater than 2.0 mmol/L. Within-run CVs for low (0.31 mmol/L) and high (1.15 mmol/L) lithium controls were 5.9% and 1.7%, respectively (n = 20). Day-to-day CVs for the same controls were 9.8% and 3.3%, respectively (n = 20). There was no significant interference when the concentrations of sodium, potassium, calcium, or magnesium were varied, nor did intervening urinary lithium analyses affect the measurement of serum lithium. Results for lithium measurement in four serum-based survey materials compared well with results by isotope dilution/mass spectrometry.
Significance of isolated increases in total lactate dehydrogenase and its isoenzymes in serum of patients with bacterial pneumonia.
Authors: Rotenberg Z , Weinberger I , Davidson E , Fuchs J , Sperling O , Agmon J
Published: January 2020

Total lactate dehydrogenase (LD, EC 1.1.1.27) activity in serum and LD isoenzymes were quantified at the time of diagnosis in 320 patients with bacterial pneumonia. In eighty, LD activity was increased, but this was accompanied by either other pathological results for liver-function tests or associated diseases that could explain it. The remaining 240 patients were divided into four groups, based on their total serum LD values: group A, less than 225 U/L (normal limit); group B, 226-350 U/L; group C, 351-499 U/L; and group D, greater than 500 U/L. Total LD was above normal at diagnosis in 40% of the patients. Recovery time was twice as long in group D as in groups A, B, and C. In five patients from group D, the pneumonia reflected underlying lung cancer. In groups B and C, the LD-3 ratio was increased in comparison with group A; in group D, LD-4 and LD-5 were increased up to twice the normal limit. Evidently nearly half of patients with bacterial pneumonia may show isolated increases in total LD activity (mostly LD-3) in serum. In cases with high activity, prolonged recovery time is expected. Intensive follow-up and extensive investigation are warranted in these patients, because some may have underlying lung cancer.
Immunofluorometric demonstration and quantification of placental protein 5 in the absence of pregnancy.
Authors: Bützow R , Alfthan H , Stenman U H , Suikkari A M , Bohn H , Seppälä M
Published: January 2020

This time-resolved immunofluorometric assay (IFMA) developed for measurement of placental protein 5 (PP5) involves two antibodies: a monoclonal anti-PP5 antibody attached to a solid phase and an europium(III) chelate-labeled polyclonal anti-PP5 antibody as a tracer. The measuring range is 0.05-100 micrograms/L and the detection limit is 20 times lower than that of a PP5 radioimmunoassay (RIA) performed with the same polyclonal antiserum. By IFMA, PP5 could be detected and quantified in all plasma and serum samples of nonpregnant and pregnant individuals, whereas PP5 was undetectable by RIA in serum of healthy men and nonpregnant women. The mean concentration of PP5 in sera from men was 0.43 micrograms/L (SD 0.13, range 0.19-0.75, n = 47) and in sera from nonpregnant women 0.49 micrograms/L (SD 0.19, range 0.20-0.90, n = 41). PP5 concentrations in serum showed no systematic variation during the menstrual cycle. In serum samples from 60 pregnant women the results obtained by IFMA and RIA correlated well (r = 0.97).
Indications for measurement of total carbon dioxide in arterial blood.
Authors: Kost G J , Trent J K , Saeed D
Published: January 2020

There is increasing evidence of variability in pK1', the practical dissociation coefficient used in the Henderson-Hasselbalch equation to calculate arterial bicarbonate from measurements of arterial pH and pco2. The case presented here illustrates not only potential technical artifacts in arterial blood sampling, which can confuse, but also irreconcilable differences in the values of calculated arterial bicarbonate vs measured arterial and venous total carbon dioxide (carbon dioxide content). Measurements of total carbon dioxide in arterial blood will resolve such conflicts, particularly for acutely ill patients, and will reflect the correct bicarbonate measurements for use in therapeutic decisions.
Chromogenic ionophore-based methods for spectrophotometric assay of sodium and potassium in serum and plasma.
Authors: Kumar A , Chapoteau E , Czech B P , Gebauer C R , Chimenti M Z , Raimondo O
Published: January 2020

We report spectrophotometric methods for quantifying sodium and potassium in serum and plasma without sample pretreatment or solvent-extraction steps. The methods are based on novel chromogenic ionophores characterized by molecular structures highly preorganized for binding, which result in exceptionally high sensitivity and selectivity (negligible responses to sodium at 80 to 200 mmol/L in the potassium assay and to potassium at 0 to 10 mmol/L in the sodium assay). The within-run imprecisions are comparable with those for ion-selective electrodes (for K, 1.0% at 4.5 mmol/L; for Na, 1.1% at 136 mmol/L); the analytical range is 2-10 mmol/L for potassium, 80-170 mmol/L for sodium. The results for greater than 100 patients' samples correlated well with results obtained with ion-selective electrodes (r = 0.99 for K, 0.97 for Na). The ready-to-use aqueous reagents will be available under the trade name of ChromoLyte for use with Technicon's RA family of clinical analyzers.
Enzyme electrode for determining glucose in whole blood.
Authors: Genshaw M A
Published: January 2020

In this sensor for glucose in whole blood, glucose oxidase (EC 1.1.3.4) is directly coupled to a platinum electrode by a silane coupling agent and glutaraldehyde. This very thin catalytic layer is then coated with a thin rubber membrane. The response to glucose in whole blood varies linearly with glucose concentration to at least 6000 mg/L with a small positive intercept. The response time is a few minutes. The key factor in the success of this sensor is the unitized construction, with both membrane and enzyme chemibonded to the electrode. The compatibility with whole blood and the high oxygen permeability of the membrane also contribute to the good performance. The coefficient of variation in replication (glucose concentration 1 g/L, n = 13) is about 3%. The sensor is insensitive to hematocrit and has a lifetime of several days after initial use.
Direct determination of free thyroxin in undiluted serum by equilibrium dialysis/radioimmunoassay.
Authors: Nelson J C , Tomei R T
Published: January 2020

We have designed a re-usable dialysis cell and a complex dialysis buffer, with which undiluted serum samples can be dialyzed with minimal changes in their serum matrix. Dialysate thyroxin (free T4) is then measured by a sensitive RIA for T4. The range of reportability was 2-128 ng/L, the normal range was 8-27 ng/L, and the interassay CV was 7%. Free T4 concentrations in various disorders were as follows: hyperthyroidism, 32-478 ng/L; in both excess thyroxin-binding globulin (TBG) and familial dysalbuminemic hyperthyroxinemia, 9-27 ng/L; primary hypothyroidism, less than 2-7 ng/L; central hypothyroidism, 4-6 ng/L; severe TBG deficiency, 9-25 ng/L; hypothyroxinemias of nonthyroidal illness, 8-35 ng/L. With this free-T4 assay, which is adaptable to clinical laboratory use, one can differentiate hyperthyroidism from the major euthyroid hyperthyroxinemias and hypothyroidism from the major euthyroid hypothyroxinemias.
Quick method of determining lipoproteins, including those of intermediate density, in serum.
Authors: Fontanals-Ferrer N , Serrat-Serrat J , Sorribas-Vivas A , Gonzalez-Garcia C , Gonzalez-Sastre F , Gomez-Gerique J
Published: January 2020

We describe an ultracentrifugation method for isolating the different lipoprotein classes relatively quickly. In this method the very-low-density lipoproteins are first separated by non-density-adjusted ultracentrifugation. The resulting infranatant material is then stained with Coomassie Brilliant Blue R-250 and ultracentrifuged in a density gradient. The intermediate-density lipoproteins (IDL), low-density lipoproteins, and high-density lipoproteins fractions are separated by aspiration from the top of the tube. This method can be used to separate, analyze, and quantify lipoproteins, including anomalous lipoproteins such as the IDL. The CVs for the present method never exceeded 15%.
High-density lipoprotein apolipoproteins in urine: I. Characterization in normal subjects and in patients with proteinuria.
Authors: Gomo Z A , Henderson L O , Myrick J E
Published: January 2020

A high-resolution two-dimensional electrophoretic method for protein, with silver staining, has been used to characterize and identify urinary high-density-lipoprotein apolipoproteins (HDL-Apos) and their isoforms in healthy subjects and in patients with kidney disease. Analytical techniques based on both molecular mass and ultracentrifugal flotation properties were used to isolate urinary lipoprotein particles with characteristics identical to those of HDL in plasma. HDL-Apos identified in urine of normal subjects and patients with glomerular proteinuria were Apos A-I, A-II, and C. Five isoforms of Apo A-I were present. Immunostaining of electroblotted proteins further confirmed the presence of HDL-Apos in urine. Creatinine clearance rate was decreased in the patients with proteinuria, and ranged from 32.5 to 40 mL/min. Concentrations of cholesterol and triglycerides in serum were greater in the patients' group, whereas mean HDL-cholesterol (0.68, SD 0.10 mmol/L) and Apo A-I (0.953, SD 0.095 g/L) were significantly (each P less than 0.01) lower. Results of this study suggest that measurement of urinary Apo A-I will reflect excretion of HDL in urine.
Cholesteryl ester transfer activity in plasma of patients with familial high-density lipoprotein deficiency.
Authors: Sparks D L , Frohlich J , Pritchard P H
Published: January 2020

We determined cholesteryl ester transfer activity in whole plasma and in lipoprotein-depleted plasma of normolipidemic subjects and of patients with severe high-density lipoprotein (HDL) deficiency: Tangier disease, lecithin:cholesterol acyltransferase (LCAT) deficiency, and "fish-eye" disease. Transfer rates in plasma were positively correlated (r = 0.950) with rates measured in the absence of the endogenous lipoproteins. This suggests that lipoprotein composition and content may not affect total cholesteryl ester transfer activity in normolipidemic and the HDL-deficient subjects. Cholesteryl ester transfer from solid-phase-bound HDL to plasma lipoproteins was decreased by 39% in fish-eye disease and 33% in LCAT deficiency but increased by 57% in Tangier disease, as compared with normal values. Changes were similar for lipoprotein-depleted plasma from the same individuals. Transfer to plasma HDL was significantly decreased in all HDL-deficient patients, whereas transfer to very-low- and low-density lipoproteins was increased only in Tangier disease. Differences in transfer rates between the patients studied appeared to reflect the LCAT activity and the need to transport cholesteryl ester rather than the HDL cholesterol concentration. Thus, the concentration of HDL in plasma does not directly affect total cholesteryl ester transfer activity in HDL deficiency.
Immunoturbidimetric method for routine determinations of apolipoproteins A-I, A-II, and B in normo- and hyperlipemic sera compared with immunonephelometry.
Authors: Siedel J , Schiefer S , Rosseneu M , Bergeaud R , De Keersgieter W , Pautz B , Vinaimont N , Ziegenhorn J
Published: January 2020

We describe a method for routine immunoturbidimetry of apolipoproteins (apo) A-I, A-II, and B in both normo- and hyperlipemic sera. A special antiserum reagent, consisting of a highly concentrated mixture of nonionic and anionic detergents (final concentration in the assay, 36 g/L), rapidly removes intrinsic turbidities of even strongly lipemic sera without interfering with the antigen-antibody precipitation reaction. The method has good precision, and obviates the need for special sample pretreatment, extended incubation periods, and measurment of sample blanks. A comparison with established immunoephelometric assays generally showed close agreement for analytical recoveries of the three apolipoproteins. However, in samples containing greater than or equal to 18 g of triglycerides per liter, the nephelometric assays yielded about two- to threefold higher values for apo A-II and B than did the turbidimetric procedure. To elucidate this discrepancy, we used the turbidimetric methods to assay sera with and without enzymatic lipolytic pretreatment. Even for samples with triglyceride concentrations up to 60 g/L, complete enzymatic lipolysis (as evidenced by thin-layer chromatography) did not significantly alter the recoveries of apo A-II and B from those obtained with the untreated specimens. Thus the immunoturbidimetric methods yield reliable results for apo A-I, A-II, and B, not only in normo- but also in hyperlipemic sera.
Determination of testosterone in serum not bound by sex-hormone-binding globulin: diagnostic value in hirsute women.
Authors: Loric S , Guéchot J , Duron F , Aubert P , Giboudeau J
Published: January 2020

We compared the diagnostic value of information given by total testosterone (I), free testosterone (II), the free androgen index (III), and testosterone not bound by sex-hormone-binding globulin (SHBG) (IV) as measured by a new differential ammonium sulfate precipitation technique, each step of which is conducted at 37 degrees C. SHBG and albuminemia were also measured. To examine the clinical value of IV, we analyzed single blood samples from 15 hirsute women and 15 age-matched healthy control volunteers. Values for I, II, III, and IV testosterone were all significantly higher in the hirsute group (P less than 0.01), whereas SHBG was decreased (P less than 0.01) and albumin concentrations were similar for the two groups. Overlap between values for normal and for hirsute women was 33.3% for I, 13.3% for II, and 0% for III and IV. The presented data suggest that IV measured by ammonium sulfate precipitation is the preferred discriminator for detecting hyperandrogenism, because this assay is technically simpler and less expensive than the II assay for routine investigation. It closely reflects the pool of bioavailable testosterone; thus, its main use might be as a screening test for androgen excess in women.