bluuebunny/crossref_metadata_2025 huggingface dataset:
Determination of fecal alpha 1-antitrypsin concentration by radial immunodiffusion: two systems compared.
Authors: Wilson C M , McGilligan K , Thomas D W
Published: January 2020

We compared Helena "QUIPlates" and Calbiochem "LC-Partigen" radial immunodiffusion systems for their ability to measure fecal concentrations of alpha 1-antitrypsin (FA1AT). Reference ranges for FA1AT concentrations in infants receiving various diets, in children, and in adults are given for each system. FA1AT values obtained with Calbiochem LC-Partigen plates averaged 30% greater than those obtained with Helena QUIPlates, but both systems distinguished between normal and high values. Studies involving variations of usual sample-handling procedures showed that storage at room temperature, repeated freezing and thawing, and long-term storage of frozen samples had no significant effect on measured FA1AT concentration. However, values obtained for lyophilized and nonlyophilized samples did not correlate well.
Heterogeneity of serum creatine kinase isoenzyme MM in myocardial infarction: standardization of patterns by use of cord-blood serum.
Authors: Williams J , Marshall T
Published: January 2020

We investigated serum creatine kinase (CK; EC 2.7.3.2) isoenzyme MM in myocardial infarction, using isoelectric focusing in polyacrylamide gels. As many as 14 sub-band species were detected, and sequential sampling revealed a progressive anodal shift in their distribution. A nomenclature for sub-band typing is proposed. It denotes the major sub-bands as 1 (pl 6.91), 2 (pl 6.65), and 3 (pl 6.35); which are usually detected in sera of normal CK range. Abnormal sub-bands (associated with increases in the concentrations of CK) are denoted as a (pl 7.55), b (pl 7.35), c (pl 7.25), d (pl 7.05), e (pl 6.85), f (pl 6.72), g (pl 6.50), h (pl 6.40), i (pl 6.28), j (pl 6.20), and k (pl 6.15). Cord-blood sera gave highly reproducible CK-MM patterns characterized by prominent detection of sub-bands 1-3 and faint detection of c, e, f, g, h, i, and j. It is recommended as a standard for CK-MM sub-band typing.
Spermidine oxidase activity in serum of normal and schizophrenic subjects.
Authors: Flayeh K A
Published: January 2020

Spermidine oxidase activity in human serum is distributed over a relatively wide range, with a highly significant difference between normal and schizophrenic subjects. The enzyme activity showed no age- or sex-related differences. It is largely inhibited by quinacrine and chloroquine.
Bioassay of parathyrin: analytical characteristics and clinical performance in patients with hypercalcemia.
Authors: Klee G G , Preissner C M , Schloegel I W , Kao P C
Published: January 2020

This new bioassay for parathyrin (PTH) in plasma (bio-PTH) combines immunoextraction on affinity columns [goat anti-hPTH (1-44) conjugated to Sepharose 4B] and a receptor assay involving an osteosarcoma cell line. The mean extraction efficacy ranges from 87% (as determined with immunopurified 125I-labeled PTH) to 62% for hPTH bioactivity. The assay is standardized with synthetic hPTH (1-84) and can detect as little as 0.9 pmol/L of PTH in 2 mL of plasma. In 100 healthy adults, the 95% reference interval for bio-PTH was less than 0.9 to 6.1 pmol/L (median, 2.0 pmol/L). In 185 patients with surgically confirmed hyperparathyroidism, bio-PTH concentrations ranged from 1.0 to greater than 120 pmol/L (median, 12.9 pmol/L); 80% of values were greater than 6.1 pmol/L. In 50 patients with both preoperative and postoperative determinations, the mean (+/--SD) concentrations of calcium in serum were 113 +/- 10 and 89 +/- 6 mg/L, respectively; the median bio-PTH concentrations were 13.6 and 2.0 pmol/L, respectively. In 22 patients with nonparathyroid-mediated hypercalcemia, the concentration of bio-PTH ranged from less than 0.9 to 5.3 pmol/L (median, 1.8 pmol/L). This bio-PTH assay is slightly less sensitive than our GP235 immunoreactive PTH (iPTH) immunoassay for detecting hyperparathyroidism (Clin Chem 1982;28:69-74); however, the bioassay is more specific and detected some cases missed by the iPTH assay. Overall, 95% of the hyperparathyroid patients had an increased test result for either the bio-PTH or the iPTH assay.
New approach to competitive lanthanide immunoassay: time-resolved fluoroimmunoassay of progesterone with labeled analyte.
Authors: Déchaud H , Bador R , Claustrat F , Desuzinges C , Mallein R
Published: January 2020

This solid-phase competitive europium immunoassay of progesterone in plasma relies on antigen labeling. With this new approach, time-resolved fluoroimmunoassay can attain sensitivity and precision similar to that of conventional radioimmunoassay. The europium-labeling involves coupling diethylenetriaminepentaacetic acid (chelating agent for Eu3+) to a progesterone-protein conjugate. The solid-phase antibody is immobilized inside polystyrene tubes in which plasma samples (50 microL) are assayed directly, without preliminary extraction. After incubation in the presence of trichloroacetic acid, the tubes are washed and the fluorescence intensity of europium is measured by time-wavelength-resolved fluorometry, with a nitrogen laser as the pulsed excitation source. Progesterone values obtained by this procedure agreed well with those obtained by radioimmunoassay (r = 0.98, n = 97). The detection limit was equivalent to that of most RIAs (0.2 micrograms/L).
Changes in serum enzymes in moderate drinkers after an alcohol challenge.
Authors: Nemesánszky E , Lott J A , Arato M
Published: January 2020

When 14 "moderate" drinkers abstained from alcohol for four weeks, the activity of gamma-glutamyltransferase (GGT; EC 2.3.2.2) in their serum showed a large decrease. Immediately after the period of abstention, an orally given ethanol challenge of 1 g/kg produced a marked increase in serum GGT at 24 h, followed by a slow decline thereafter. Aspartate amino-transferase activity in serum was significantly increased at 24 h; however, alkaline phosphate, alanine aminotransferase, and lactate dehydrogenase showed much smaller or no changes. An abnormal increase in lactate dehydrogenase isoenzyme 5 was observed in seven subjects. In some of the moderate drinkers, liver biopsies showed mild chronic hepatitis or nonspecific changes. Eight nondrinking controls showed only slight increases in serum GGT following the same alcohol challenge; results for the other enzyme tests were unchanged. We consider it probable that pre-existing liver disease affects the response to ethanol, so that greater amounts of GGT are released from hepatic tissue; alternatively, drinkers may have a higher GGT activity in this tissue as a result of enzyme induction by ethanol. The alcohol challenge test was an effective discriminator between moderate drinkers and abstainers.
Effect of creatine kinase-MM subtype composition on a CK-MB immunoinhibition assay.
Authors: Morison I M , Clayson K J , Fine J S
Published: January 2020

Immunoinhibition (INH) by use of polyclonal anti-human CK-M antibody may be used to measure CK-MB in serum. Previous studies have shown that inhibiting antibodies prepared against purified muscle extracts may inhibit CK-MM by greater than 99%. Using patients' sera and muscle homogenates incubated with human serum, we studied the effect of CK-MM subtype composition on an INH assay. We found that with increasing time from the CK-releasing event, e.g., myocardial infarction, or with longer in vitro incubation, the proportion of CK-MM1 increased and the proportion of uninhibited CK-MM increased from 0.2% to 0.7-0.8%. As a consequence, CK-MB activity may be overestimated by as much as 1.6% of total CK when uncorrected INH results are used. Inhibition was maximal in samples containing 100% CK-MM3, the tissue subtype. Because of the time-dependent change in CK-MM subtypes, published results for INH from studies in which CK-MM purified from muscle was used may not be directly applicable to clinical specimens.
Anodic stripping voltammetry procedure modified for improved accuracy of blood lead analysis.
Authors: Roda S M , Greenland R D , Bornschein R L , Hammond P B
Published: January 2020

In evaluating the accuracy and reliability of blood lead (PbB) measurements with the Environmental Science Associates Model 3010A Trace Metal Analyzer, intralaboratory comparison demonstrated that use of the operating conditions recommended by the manufacturer resulted in consistently underestimated PbB concentrations less than 400 micrograms/L and overestimated PbB values greater than 400 micrograms/L. At PbB concentrations less than 50 micrograms/L, measured concentrations were often registered as negative results. However, these negative values could be replicated to within +/- 10 micrograms/L, indicating good precision of the method, but obviously not good accuracy. In addition, lower-than-expected lead (Pb) values were measured in samples containing increased concentrations of copper (Cu), such as may occur in pregnant women. We modified the procedure to eliminate these inaccuracies by substituting manual peak-height measurements for reliance on the integrator and digital display of the instrument. We established the accuracy of the modified procedure by using calibration standards previously quantified by isotope dilution-mass spectroscopy. A quality-control program for monitoring PbB analysis is also described.
Ten highly sensitive thyrotropin assays compared by receiver-operating characteristic curves analysis: results of a prospective multicenter study.
Authors: Thonnart B , Messian O , Linhart N C , Bok B
Published: January 2020

We report a prospective multicenter study, undertaken to compare the efficacy of 10 highly sensitive thyrotropin assay kits for the diagnosis of hyperthyroidism. Performances of the kits were compared with a reference diagnosis based on clinical examination, pertinent biological tests, and determination by an independent laboratory of the concentrations in serum of free triiodothyronine and free thyroxin. No thyrotropin determination was used in establishing this reference diagnosis. Receiver-operating characteristic curves were obtained for results from 600 patients (217 hyperthyroid and 383 euthyroid) by each kit. Even though analyses were performed out of the working range of most kits, the clinical correlation was nevertheless excellent. The best results corresponded to a sensitivity of 97.5% associated with a specificity of 96.1% and were significantly better than those obtained with all other kits. Results of this comparison depended greatly on the heterogeneity of the "normal"/"abnormal" categories. When only diffuse hyperthyroidism was considered, sensitivity and specificity were improved for all kits, and there was no significant difference among the five best kits.
Improved thin-layer chromatographic determination of phospholipids in gastric aspirate from newborns, for assessment of lung maturity.
Authors: Serrano de la Cruz D , Santillana E , Mingo A , Fuenmayor G , Pantoja A , Fernández E
Published: January 2020

This one-dimensional thin-layer chromatographic method is used for assay of phospholipids in the gastric aspirate of newborns. The solvent mixture (chloroform/hexane/methanol/glacial acetic acid/water, 12/7/4/3/0.3 by vol) completely resolves lecithin, sphingomyelin, phosphatidylinositol, phosphatidylserine, phosphatidylethanolamine, and phosphatidylglycerol. The method is simple, precise, inexpensive, and rapid (chromatographic development takes less than 25 min) and gives high chromatographic resolution. We used this method to determine the lecithin/sphingomyelin densitometric ratio (L/S ratio) and the phosphatidylglycerol percentage in 200 samples of gastric aspirate and found an L/S ratio of 2.5 to be a satisfactory cutoff value for distinguishing fetal lung maturity and immaturity. We confirmed that the presence of phosphatidylglycerol excluded the possibility of respiratory distress.
Quantification of human apolipoprotein A-IV by "sandwich"-type enzyme-linked immunosorbent assay.
Authors: Rosseneu M , Michiels G , De Keersgieter W , Bury J , De Slypere J P , Dieplinger H , Utermann G
Published: January 2020

A specific and sensitive "sandwich"-type enzyme-linked immunosorbent assay (ELISA) has been developed for quantifying human apo A-IV. Using apo A-IV immunosorbent columns, we isolated monospecific anti-apo A-IV antibodies for coating the ELISA plates and for preparing peroxidase-antibody conjugate. The assay can detect as little as 0.20 ng of apo A-IV, with mean intra- and interassay CVs of 3.6% and 8.2%, respectively. The apoA-IV concentrations in normolipemic and hyperlipemic plasma were unaffected by either delipidation or treatment with detergents or urea. To validate the ELISA assay we compared it with an immunoelectrophoretic technique. ApoA-IV concentrations in plasma from normo- and dyslipemic subjects compared well by the two assays (r = 0.89). The mean apo A-IV concentration, measured by ELISA in plasma from 50 normolipemic subjects, was 143 (SD 52) mg/L; values for dyslipemic subjects were not significantly different. We also used this new assay to monitor apo A-IV profiles of normolipemic and hypertriglyceridemic plasma after chromatographic fractionation.
Reliability of IL Monarch ion-selective electrode module for sodium, potassium, and chloride measurements.
Authors: Guagnellini E , Spagliardi G , Bernardi G , Stella P
Published: January 2020

We evaluated the IL Monarch random-access centrifugal analyzer for measurement of Na+, K+, and Cl- by an indirect potentiometric method. For different concentrations of control material, the total precision (CV) ranged between 0.82% and 1.14% for the three electrolytes; linearity was acceptable within a range of 103 to 215 mmol/L for Na+, 1.6-15.25 mmol/L for K+, and 80-173 mmol/L for Cl-. Data correlated well with those by flame photometry for Na+ and K+ and with those by coulometry for Cl-, both for various biological materials--sera, urines, dialysis fluids--and commercial control materials from various producers. Stability of the potentiometric signal was acceptable: daily variations were 0.2 mV for Na+, 0.05 mV for K+, and 0.03 mV for Cl-. Accordingly, we conclude that the system supplies reproducible and accurate results while being easy to use and requiring little maintenance. The use of indirect potentiometry offers results consistent with those obtained with traditional methods, and easily interpretable by clinical staff. However, better information about the actual ion activity in the tested sample for certain pathologies such as hyperlipemia and dysproteinemia could be obtained by methods involving direct potentiometry.
Antidepressants in the medically ill: diagnosis and treatment in primary care.
Authors: Katon W , Roy-Byrne P P
Published: January 2020

Major depression is one of the most common clinical problems seen by primary-care physicians. The prevalence is 5% to 10% in primary-care outpatients and 15% in medically ill inpatients. Despite this high prevalence rate, depression is frequently not accurately diagnosed. Evidence suggests that misdiagnosis often occurs because patients with depression frequently focus preferentially on somatic complaints or amplify complaints of chronic medical illness. The tricyclic antidepressants are more effective than placebo, both in treating medically ill patients with depression and primary-care patients with major depression. Common side effects of the tricyclic antidepressants such as the anticholinergic, cardiac, and blood-pressure problems are reviewed, with emphasis on the use of specific tricyclics in medically ill populations.
Prenatal diagnosis of cystic fibrosis: microvillar enzymes and DNA analysis compared.
Authors: Buffone G J , Spence J E , Fernbach S D , Curry M R , O'Brien W E , Beaudet A L
Published: January 2020

We have established reference ranges for three microvillar intestinal enzymes--alkaline phosphatase (EC 3.1.3.1), gamma-glutamyltransferase (EC 2.3.2.1), and leucine aminopeptidase (EC 3.4.1.1)--measured in amniotic fluid in a reference population of 1875 women presenting for routine amniocentesis. These data were derived for use in prenatal diagnostic studies in a population at risk (1:4) for cystic fibrosis. False-positive or indeterminate results were noted for fewer than 3.5% of all low-risk cases for each enzyme evaluated. Total alkaline phosphatase and its isoenzymes and leucine amino-peptidase and gamma-glutamyltransferase were measured in amniotic fluid sampled between the 15th and 19th weeks of gestation. Restriction fragment length polymorphism analysis of DNA was also performed when possible. In 52 cases examined for cystic fibrosis thus far, 46 were diagnosed on the basis of DNA analysis and (or) by sweat testing; for the other six cases, only abnormal enzyme results were obtained before termination of pregnancy. Predictions based on microvillar enzyme results were falsely negative in three cases. In only one case was there a discrepancy between enzyme results and DNA analysis. Diagnostic accuracy was highest during the 17th and 18th week of gestation. Preliminary results suggest the false-negative rate of this diagnostic strategy may be greater than or equal to 10%.
Six direct radioimmunoassays of estradiol evaluated.
Authors: Schiøler V , Thode J
Published: January 2020

We evaluated the analytical performance of six commercial direct radioimmunoassays of 17 beta-estradiol, those from Radioassay Systems Laboratories, IRE Medgenix, Biotecx Laboratories, Farmos Spectria, International CIS, and Diagnostic Products Corp. The mean value for estradiol (E2) and the within- and between-run CVs were determined for serum pools and control materials, measured in seven to 10 runs with each method. Mean values for E2 in pooled sera deviated by 75% to 350% from the means measured with our routinely used extraction method. Between-run CVs ranged from 4% to 14% for the direct assays as compared with 10% for the extraction assay. We also investigated, for two of the direct assays, the effect of extraction with diethyl ether before radioimmunoassay, with respect to improvement in the agreement with our extraction method. All of the assays were easy to perform and results were obtained within 4-5 h, but we conclude that matrix effects may be expected in direct assays of estradiol.
Lack of effect of warfarin on uric acid concentration.
Authors: Walker F B , Becker D M , Kowal-Neeley B , Krongaard L S
Published: January 2020

Uric acid concentrations reportedly are increased in patients being treated with warfarin. We measured uric acid in 40 patients before and during warfarin administration. The mean pre- and post-warfarin uric acid concentrations for our patients were 0.39 mmol/L and 0.40 mmol/L, respectively, not a significant difference. Further observations of hyperuricemic patients started on warfarin are needed to prove that their risk of gout is not increased.
Identification of HIV-specific oligoclonal immunoglobulins in serum of carriers of HIV antibody.
Authors: Papadopoulos N M , Costello R , Ceroni M , Moutsopoulos H M
Published: January 2020

Zone electrophoresis on agarose gel was performed on serum samples from HIV-antibody carriers and negative controls. Nitrocellulose strips precoated with an HIV preparation were then placed on top of the gels and developed by an immunoblotting procedure. A positive reaction was demonstrated between the HIV antigens and the HIV-antibody-positive serum samples with hypergammaglobulinemia and oligoclonal IgG bands. A negative reaction was found between the HIV antigens and HIV-antibody-negative serum samples from a normal person and a patient with monoclonal gammopathy. The presence of oligoclonal IgG bands in the serum of HIV-antibody carriers, and their positive identification with HIV antigens, indicates a specific immune response of the host to the HIV infection and supports the use of oligoclonal IgG bands as markers to follow the course of HIV infection.
One-step, high-yield purification of human prostatic acid phosphatase from seminal fluid by gel-filtration HPLC under nondenaturing conditions.
Authors: Rusciano D , Agostini E , Ceccarini C , Terrana B
Published: January 2020

This is a fast, efficient method for purification to homogeneity of human prostatic acid phosphatase [orthophosphoric-monoester phosphohydrolase (acid optimum), EC 3.1.3.2], from seminal fluid. Use of a "high-pressure" liquid-chromatographic gel-filtration column permits high-yield recovery of the purified enzyme with most of its enzymatic and immunological activity retained.
Are the classical markers sufficient to describe uremic solute accumulation in dialyzed patients? Hippurates reconsidered.
Authors: Schoots A C , Dijkstra J B , Ringoir S M , Vanholder R , Cramers C A
Published: January 2020

Interdependencies of accumulated solutes, analyzed by liquid chromatography in dialyzed and non-dialyzed patients, were studied by multivariate statistical analysis. In principal component analysis, three principal components (PC1-PC3) were retained from the data on 22 accumulated compounds in dialyzed patients, whereas only one principal component was retained from analogous data of a non-dialyzed patient group. PC1 in the dialyzed patient group comprises concentrations of hippuric acid, p-hydroxyhippuric acid, tryptophan, and five unidentified fluorescent solutes in serum. Concentrations of the classical markers urea, uric acid, creatinine, and phosphate were closely related to PC2 in these patients. Indoleacetic acid and two unidentified fluorescent compounds constitute PC3. The compounds associated with the groups found by principal component analysis may be characterized by chemical structure and by the mechanism of their excretion via the remaining nephrons of dialyzed patients. In the non-dialyzed group, most of the solutes could be described by a single PC. This PC and PC1 from the dialyzed group correlated significantly with residual renal function, and with total ultraviolet absorbance and total fluorescence emission. The data suggest that it is of value to introduce a marker of uremic solute retention in addition to urea, to account for renal-function-related "organic-acid-like" compounds that are excreted by renal tubular secretion in dialyzed patients. The hippurates may serve this purpose.
An enzymatic method for erythrocyte acetylcholinesterase.
Authors: Abernethy M H , Fitzgerald H P , Ahern K M
Published: January 2020

The acetylcholinesterase (EC 3.1.1.7) in 50 microL of a 61-fold dilution of erythrocytes in water hydrolyzes acetylcholine during a timed 20-min reaction at 37 degrees C. The resulting choline is measured by use of choline oxidase coupled to peroxidase, with phenol and aminoantipyrene to give a pink product that absorbs maximally at 500 nm. For calibration, a choline iodide standard is included in each batch of up to 19 samples. Accuracy was assessed by using specific inhibitors and measuring choline in the presence of excess erythrocyte solution. The standard curve for the assay is linear to threefold the normal enzyme activity. Between-batch precision was 0.40 kU/L at a mean of 11.5 kU/L (CV 3.5%), and comparison with an acetylthiocholine procedure (x) gave a good correlation: y = 1.02x - 0.27 kU/L (r = 0.991). Long-term precision (10 months), assessed from three sets of assays of samples from 17 individuals, was 0.71 kU/L at a mean of 11.7 kU/L (CV 6.1%).
Development of standard materials for use with the hematofluorometer.
Authors: Parsons P J , Meola J R , Mitchell D G
Published: January 2020

Two methods for preparing erythrocytes were evaluated for the purpose of developing stable, blood-based standard materials for the hematofluorometer. Erythrocytes washed with citrate-phosphate-dextrose solution and reconstituted with platelet-free plasma were stable for 12 weeks. Plasma-free erythrocytes washed with isotonic saline and resuspended in citrate-glycerol solution were stable for 18 weeks when stored at 4 degrees C. Consequently, plasma-free erythrocytes were successfully used as proficiency test specimens in the New York State Department of Health's Clinical Laboratory Evaluation Program for hematofluorometer users.
Potassium permanganate used to eliminate interference with the Porter-Silber reaction in the butanol extraction method for determination of urinary 17-hydroxycorticosteroids.
Authors: Maeda T , Matsuzaki H , Sekine T
Published: January 2020

We describe a simple, rapid, and less interference-susceptible method for using the Porter-Silber reaction for determination of urinary 17-hydroxycorticosteroids. The procedure is based on butanol extraction of the steroids after the urine sample is treated with potassium permanganate and sodium bisulfite. These treatments, together with an additional acid- and alkali-washing of the extract, could eliminate most of the substances in urine that interfere with the Porter-Silber reactions. Values so obtained correlated well (r = 0.95) with those by Furuya's method in which beta-glucuronidase is used. Our method may be useful and suitable for a screening test of adrenocortical and pituitary functions.
Inaccuracy in measuring glycated albumin concentration by thiobarbituric acid colorimetry and by boronate chromatography.
Authors: Johnson R N , Baker J R
Published: January 2020

We compared thiobarbituric acid colorimetry and boronate chromatography for measuring glycation of serum proteins. With 14C-glycated human albumin as a test material, both methods were acceptably linear and precise. However, comparable estimates (mmol/L) of albumin glycation ranged from 0.22 for thiobarbituric acid to 0.05 for boronate, representing yields relative to [14C]glycosylprotein of 42% and 10%, respectively. The low yield with thiobarbituric acid was corroborated independently on the basis of kinetic differences between the reactions of fructose standards and of glycosylprotein, leading to underestimation of glycosylprotein concentration. The lower estimate of glycosylprotein by boronate chromatography was related to an apparent requirement for two [14C]glyco groups per albumin molecule to effect binding.
Determination of plasma bicarbonate of neonates in intensive care.
Authors: Masters P , Blackburn M E , Henderson M J , Barrett J F , Dear P R
Published: January 2020

Most modern blood-gas analyzers are programmed to use the Henderson-Hasselbalch equation to calculate a value for plasma bicarbonate. It has been suggested, however, that among acutely ill patients, including newborns, these calculated values may be at variance with measured total CO2. To assess the clinical significance of such errors, we compared calculated bicarbonate with measured total CO2 in 79 blood samples from 40 babies in intensive care. The calculated bicarbonate values consistently exceeded the measured values by about 1.5 mmol/L. Of the errors, 94% were within the range -10% to +20%. When the systematic bias was removed, calculated and measured bicarbonate values agreed within +/- 3.30 mmol/L in 95% of cases. Because calculated values can be obtained much more quickly and frequently than laboratory measurements, we believe that these limits are clinically acceptable.
Confirmation and certainty in toxicology screening.
Authors: Spiehler V R , O'Donnell C M , Gokhale D V
Published: January 2020

Confirmation of presumptive positive urine drug screens, necessary to minimize the reporting of false-positive results, can be costly and time-consuming. The predictive value model can be used to select the confirming tests and to calculate the confidence of the result. The predictive value of a test result is the probability, based on the sensitivity and specificity of the test, that the result is a true positive or a true negative. The predictive value model applied to toxicology screening tests for drugs of abuse showed that prevalence, in addition to sensitivity and specificity, was the factor controlling the confidence level of a result. For example, the predictive value of a positive result for a screening test that has a sensitivity of 99% and a specificity of 99%, applied to screening in a population with a prevalence of 1% is 0.50; for a prevalence of 10%, it is 0.92. Confirmation with a second, chemically independent, test of equal sensitivity and specificity increases the predictive value to 0.99.
Radioimmunoassay of salivary cyclosporine with use of 125I-labeled cyclosporine.
Authors: Coates J E , Lam S F , McGaw W T
Published: January 2020

We prepared 125I-labeled cyclosporine (125I-CS) by modifying the procedure of Mahoney and Orf (Clin Chem 1985;31:459) and characterized it with regards to maximal immunoreactivity (greater than 90%), trichloroacetic acid precipitability (greater than 90%), and stability (90% immunoreactive after five half-lives of 125I). For a particular preparation of 125I-CS, we estimated its immunoreaction concentration (50 pmol/L) and the equilibrium constant for its reaction with Sandoz polyclonal antiserum (K = 3.9 X 10(9) L/mol). By substituting 125I-CS as tracer in the Sandoz radioimmunoassay and by modifying other aspects of the assay, we developed a procedure that is sufficiently sensitive (0.34 micrograms/L) to allow measurement of "trough" (lowest inter-dose) cyclosporine concentrations in parotid saliva. Of 38 kidney-transplant patients, 35 had measurable concentrations in saliva (mean 8.3, SD 5.2 micrograms/L), and these correlated moderately with paired serum concentrations (r = 0.68, P less than 0.001). We believe that measurement of salivary cyclosporine may offer a simple way of estimating the free fraction of the drug in serum or plasma.
Re-evaluation of the fructosamine reaction.
Authors: Phillipou G , Seaborn C J , Phillips P J
Published: January 2020

The difference in spectral characteristics between 1-deoxy-1-morpholinofructose (DMF) and protein/plasma samples in the fructosamine reaction has been related to the solubility of the diformazan formed by reduction of nitro blue tetrazolium chloride. Addition of the surfactant Triton X-100 (20 g/L) to the reagent buffer not only corrects this anomaly but also enhances the absolute response. Detailed investigation of DMF and dihydroxyacetone as calibration standards for the reaction established a clear preference for the latter. Fundamental differences in reaction kinetics were also noted between the Amadori rearrangement products of glucose formed from morpholine (DMF) or the amino lysine groups of protein (glycated albumin). From the reactivity of dihydroxyacetone, as well as glyceraldehyde, observed in the fructosamine reaction, and the presence of this class of compounds (trioses) in human plasma, we infer that they may also contribute to the differentiation of diabetic and non-diabetic samples.
Liquid-chromatographic determination of total hydroxyproline in urine.
Authors: Dawson C D , Jewell S , Driskell W J
Published: January 2020

In this method for quantifying 4-hydroxyproline in human urine, 50 microL of urine is hydrolyzed, derivatized with phenylisothiocyanate (PITC), and then quantified by reversed-phase HPLC with ultraviolet detection. The detection limit in urine is 373 pg of hydroxyproline per 50-microL injection. The total CVs for high- and low-concentration pools are 5.3% and 3.9%, respectively (10 runs in 10 days). The standard curve of the assay is linear over a range of 0 to 22 nmol per injection. We estimated the normal range for hydroxyproline excretion in men on an unrestricted diet to be 123-308 mumol/24 h. We also report hydroxyproline concentrations in patients with metastatic bone disease and cirrhosis of the liver.
Sensitive, rapid procedure for time-resolved immunofluorometry of lutropin.
Authors: Khosravi M J , Morton R C , Diamandis E P
Published: January 2020

In this new immunofluorometric method for quantification of lutropin in serum, the "sandwich" principle is combined with time-resolved fluorescence measurements, with the europium chelate 4,7-bis(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA) used as label. A monoclonal antibody to the alpha-subunit of lutropin is adsorbed onto the walls of white-opaque microtiter wells to form the solid-phase capture antibody, and a biotin-labeled soluble monoclonal antibody is used for antigen quantification. The detection system is completed with streptavidin, which has been linked to a protein bulking agent labeled with multiple BCPDA residues. In the presence of excess europium, the fluorescence of the final complex attached to captured lutropin molecules is measured on the dried solid phasse with an automated time-resolved fluorometer. The assay can be performed as a rapid (less than 60 min incubation) or regular (150 min incubation) procedure. The rapid assay is well-suited for routine daily monitoring of increasing or ovulatory lutropin concentrations; the regular assay, with its greater sensitivity (0.5 int. unit/L), is a practical procedure for lutropin measurements in hyposecretory states. The assay measures up to 240 int. units/L, and results compare well with those by a commercially available radioimmunoassay, an immunoradiometric assay, and another time-resolved immunofluorometric procedure.
An evaluation of the trends in analytical performance of international apolipoprotein A-1 and B assays.
Authors: Smith S J , Henderson L O , Cooper G R , Hannon W H
Published: January 2020

Two international apolipoprotein A-1 and B (apoA-1 and apoB) surveys were conducted in 1983 and 1986 with use of a lyophilized serum measured by 55 and 91 participants, respectively. The chief source of variability in both surveys was among laboratories, but among-method variation was significant for apoB. Comparing the 1986 with the 1983 findings, we saw a 60% decrease in apoA-1 variability among laboratories, and a similar decrease (53%) was found for apoB. Evaluation of individual measurements suggests that some collaborators may have adjusted their calibrators toward the consensus values published in 1983.
Development and performance of a fully automated method for assay of C-reactive protein in the aca discrete clinical analyzer.
Authors: Schwartz M W , Schifreen R S , Gorman E , Tuhy P M , Bienvenu J , Warkentin D L
Published: January 2020

A quantitative immunoassay for C-reactive protein (CRP) has been developed for use in the Du Pont aca discrete clinical analyzer. Particle-enhanced turbidimetric immunoassay (PE-TIA) technology is used. The method has a CV of less than 10% in the range 2 to 120 mg/L. Neither hemolyzed samples (Hb less than 5 g/L), icteric samples (total bilirubin less than 300 mg/L), lipemic samples (triglyceride less than 15 g/L), nor some commonly used drugs interfere. Dithioerythritol is used to eliminate interference from rheumatoid factor. Good correlation was seen when the Du Pont CRP method was compared with the Beckman ICS, Syva EMIT, TDx, and Behring methods for CRP. The normal reference interval is 0 to 9 mg/L. The method, which is fully automated, is fast, requires only a few microliters of serum, and is well suited to emergency-room requirements.
Factitious proteinuria: diagnosis and protein identification by use of isoelectric focusing.
Authors: Sutcliffe H , Rawlinson P S , Thakker B , Neary R , Mallick N
Published: January 2020

Factitious proteinuria is an unusual finding. We present a case in which clinical suspicion was aroused by the disparity between the clinical history and findings and the 24-h excretion of protein in urine. Electrophoresis of the patient's serum and urine confirmed the presence of an unusual protein. By isoelectric focusing we identified it as egg-white, a finding confirmed by immunofixation with antiserum to egg-albumen. In the past, confirmation of the identity of such a protein has required specific antiserum for immunofixation or immunodiffusion. Such antiserum may not always be available. However, isoelectric focusing gives sufficient resolution for positive identification of exogenous proteins, even in the presence of true proteinuria.
Human and primate monoclonal antibodies for in vivo therapy.
Authors: Ehrlich P H , Moustafa Z A , Justice J C , Harfeldt K E , Gadi I K , Sciorra L J , Uhl F P , Isaacson C , Ostberg L
Published: January 2020

Human monoclonal antibodies, owing to their decreased immunogenicity, are expected to be an improvement over mouse monoclonal antibodies for in vivo therapy. Human and primate monoclonal antibodies are best produced with a human x mouse heteromyeloma. Several human chromosomes are stable in the human x (human x mouse) hybrids. Chimpanzee anti-digoxin monoclonal antibodies were prepared and characterized. Because they are structurally very similar to human antibodies, they should be well tolerated in humans. The anti-digoxin antibodies can be used for therapy of extreme overdoses or as an in vivo diagnostic tool for slight overdoses. Because the advantage of using human monoclonal antibodies is their lack of immunogenicity, preparation of the antibody must be scrupulous so as not to introduce extraneous immunogens. Analysis to ensure the purity of the preparation can be complicated by the presence of high concentrations of the antibody and the low levels of contamination that must be detected. We describe a Western blot assay for Protein A that is sensitive even in the presence of human IgG.
A non-flow cytometric system for detecting antibodies and cellular antigens in blood.
Authors: Ghazarossian V , Laney M , Vorpahl J , Pease J , Skold C , Watts R , Jeong H , Dafforn A , Cook R , Ullman E F
Published: January 2020

The system described here can distinguish between single and agglutinated erythrocytes by use of a non-flow fiber optic fluorometer. The method is capable of detecting cell-surface antigens and antibodies to cell-surface antigens present in blood. Significant features include high-efficiency fluorescent dyes that intercalate into cell membranes, a stretching membrane for transport and mixing of samples, charged colloidal magnetite for magnetic separation of erythrocytes, and an immersible fiber optic probe for measuring fluorescence associated with cells in a 1-nL volume of a bulk solution. We describe the application of the system to automation of ABO/Rh grouping and antibody screening.
Enzyme-linked immunosorbent assay for serum amyloid A apolipoprotein with use of specific antibodies against synthetic peptides.
Authors: Saïle R , Delpierre C , Puchois P , Hocke G , Cachera C , Gesquiere J C , Steinmetz A , Tartar A , Fruchart J C
Published: January 2020

We describe a noncompetitive enzyme-linked immunosorbent assay for amyloid A apolipoprotein in human serum (apo SAA) in which specific antibodies against synthetic peptides are used. Microtiter plates were used as solid phase and coated with affinity-purified antibodies raised against SAA1-(95-104) peptide. After incubation with delipidated plasmas, the bound apo SAA was revealed by labeled antibodies raised against SAA1-(58-69) peptide. The assay offers several advantages over existing techniques: sensitivity, specificity, simplicity, and non-use of radioisotopes. Results correlate well with those by a nephelometric method in which polyclonal antibodies are used.
Liver-function studies in heart-transplant recipients treated with cyclosporin A.
Authors: Gulbis B , Adler M , Ooms H A , Desmet J M , Leclerc J L , Primo G
Published: January 2020

Cyclosporine (CsA) hepatotoxicity has been reported but has not been studied systematically. This study includes 17 patients undergoing heart transplantation (HTx) and being treated with CsA, azathioprine, and corticosteroids. We assessed liver function in these patients before HTx and during the following month by five biological tests: total bile acids (BA), alkaline phosphatase (AP), gamma-glutamyltransferase (GGT), and total bilirubin in serum, and the aminopyrine breath test (ABT). With these tests we could classify the patients into three groups before HTx: normal (group I), mildly altered (group II), and severely altered (group III) liver function. During CsA therapy we did not observe any changes in any test results except for BA and GGT, and these only in group III. The ABT improved significantly in this group. During kinetic studies in patients without liver dysfunction, we confirmed a direct interference of CsA on BA secretion mechanism, but not the GGT increase, which remains to be explained.
Kinetic measurement of the combined concentrations of acetoacetate and beta-hydroxybutyrate in serum.
Authors: Nuwayhid N F , Johnson G F , Feld R D
Published: January 2020

This is an automated method for the kinetic measurement of the combined concentrations of acetoacetate and beta-hydroxybutyrate in a single channel of the "Multistat III" centrifugal analyzer. Acetoacetate is first reduced with high concentrations of NADH by catalysis with 3-hydroxybutyrate dehydrogenase (EC 1.1.1.30). This reaction mixture is diluted with excess NAD+. The endogenous beta-hydroxybutyrate and that resulting from acetoacetate are then measured kinetically. Comparing the combined concentration of acetoacetate and beta-hydroxybutyrate (y) with the sum of acetoacetate and beta-hydroxybutyrate measured as described by Hansen and Freier (Clin Chem 1978;24:475) (x) yielded the relationship: y = 0.99x - 0.57 (r = 0.93, n = 25). The run-to-run CVs for low (5 mmol/L) and high (15 mmol/L) acetoacetate controls were 12% and 6%, respectively. The method is useful for determining the concentration of ketone bodies in 2-microL samples of serum of patients with diabetic ketoacidosis. The sensitivity can be increased to determine ketone body concentration in nonketotic individuals by increasing sample volume to 10 microL.
Enzymic determination of the free cholesterol fraction of high-density lipoprotein in plasma with use of 2,4,6-tribromo-3-hydroxybenzoic acid.
Authors: Moshides J S
Published: January 2020

A highly sensitive enzymic colorimetric reagent is described for determination of the free cholesterol fraction of high-density lipoprotein (HDL), which represents about 20% of the total cholesterol content of this lipoprotein. For greater sensitivity with respect to cholesterol, I used 2,4,6-tribromo-3-hydroxybenzoic acid instead of phenol in the cholesterol oxidase/peroxidase/4-aminoantipyrine reagent system. This allows determination of the free cholesterol fraction of HDL isolates prepared with polyethylene glycol 6000, a method for precipitating beta-lipoprotein that involves a twofold dilution of plasma. The reagent, adapted for use with a Cobas-Bio centrifugal analyzer, results in between-run and within-run CVs of less than 3% and a linearity to at least 400 mg of HDL free cholesterol per liter. Comparison with results by Trinder's cholesterol method, which measures cholest-4-en-3-one at 232 nm, showed good correlation (r = 0.9829, slope 1.0001, and y-intercept +2.4797 mg/L). With the manual procedure for HDL free cholesterol, between-batch and within-batch CVs were less than 5%, and results correlated well with those by the automated method (r = 0.9975, slope 0.9839, and y-intercept +2.4327 mg/L). The mean (and SD) HDL free cholesterol for 123 men was 96.8 (30.6) mg/L and for 122 women 136.4 (36.8) mg/L, indicating a distinct sex-related difference, similar to that found for HDL total cholesterol. HDL free cholesterol in plasma may therefore be a potential new predictor of coronary heart disease.
Performance of two new algorithms for estimating within- and between-method carryover evaluated statistically.
Authors: Stephens T W
Published: January 2020

Accurate and precise algorithms for estimating within-method carryover, based on the minimization of a unique "carryover sum of squares," and between-method carryover, based on a weighted Deming regression of first sample recovery vs carryover-corrected "true" recovery, are described and compared with traditional methods by use of a Monte Carlo study. In addition, I have studied the experimental parameters that influence the accuracy and precision of carryover estimation. The new algorithm for estimating within-method carryover is unbiased under most conditions, whereas the traditional algorithm is biased low under most conditions. The new algorithm is also more precise, owing to more-efficient utilization of information contained in an analytical run performed for carryover estimation. Between-method carryover in a random-access analyzer is estimated quantitatively by the second proposed algorithm and is found to be readily and precisely determinable. Use of these methods in combination to evaluate analytical interaction should allow the prediction of carryover error under most current analytical situations.
Totally automated liquid-chromatographic assay for cortisol and associated glucocorticoids in serum, with use of ASTED sample preparation.
Authors: Turnell D C , Cooper J D , Green B , Hughes G , Wright D J
Published: January 2020

We describe estimation of total cortisol in serum by HPLC. Incorporation of a new technique to totally automate sample preparation decreases costs. The benefits of HPLC ensure that interferences from other steroids are avoided. The method can also be used to estimate concentrations of other glucocorticoids in serum.
Improved agarose electrophoretic method for separating alkaline phosphatase isoenzymes in serum.
Authors: Van Hoof V O , Lepoutre L G , Hoylaerts M F , Chevigné R , De Broe M E
Published: January 2020

A modified agarose electrophoretic system for the separation of alkaline phosphatase (ALP, EC 3.1.3.1) isoenzymes is described. Bone, liver, high-molecular-mass, and intestinal ALP are separated with high reproducibility. The sensitivity of the agarose system is superior to cellulose acetate in detecting high-Mr ALP. Correlation is good between bone ALP fractions scanned before and after treatment with neuraminidase. Immunoglobulin-bound ALPs, the ALP-lipoprotein-X complex, and the additional ALP fraction observed in transient hyperphosphatasemia in children are detected by their peculiar electrophoretic mobility in the proposed system. Approximately 25% of the samples contained an additional fraction of intestinal-type ALP, as evidenced by neuraminidase treatment and use of polyclonal and monoclonal antibodies. Because the electrophoretic mobilities of this "intestinal variant" and of some immunoglobulin-bound ALP fractions are identical to those of bone and intestinal ALP, respectively, treatment of the samples with a polyclonal antibody that reacts with intestinal ALP is advised.
Simultaneous determination of carbamazepine and its epoxide and transdiol metabolites in plasma by microbore liquid chromatography.
Authors: Riad L E , Sawchuk R J
Published: January 2020

Simultaneous monitoring of carbamazepine and its metabolities is helpful in providing information on the induction process via the epoxide-diol route. The present method involves the liquid-chromatographic analysis of 1.0 mL of plasma for carbamazepine at concentrations of 0.025 to 4.0 mg/L and for both the 10,11-epoxide and 10,11-transdiol metabolites at concentrations of 0.01 to 1.0 mg/L. Weighted regression equations, expressing peak area ratios as a function of concentrations of carbamazepine and its epoxide and transdiol metabolites in the standards, were used to determine concentrations in plasma samples.
A commercial enzyme immunoassay method (EMIT) compared with liquid chromatography and bioassay methods for measurement of chloramphenicol.
Authors: Schwartz J G , Casto D T , Ayo S , Carnahan J J , Jorgensen J H
Published: January 2020

A new enzyme immunoassay method (EMIT; Syva Co.) was compared with conventional high-performance liquid chromatography (HPLC) and agar-diffusion bioassay methods for measurement of chloramphenicol in human serum. Forty-nine serum samples were assayed by each of the three methods. Excellent correlation was observed between values by EMIT and by the two conventional methods (r = 0.986 and 0.961). Precision was acceptable (CV less than 5%) with EMIT. Assay of samples containing chloramphenicol glucuronide and chloramphenicol succinate demonstrated that EMIT recognizes only the biologically active (base) form of the drug. The capability to test serum samples as small as 0.2 mL, adaptation to widely available instrumentation, and provision of rapid results are principal advantages of the EMIT method for routine chloramphenicol measurements.