bluuebunny/crossref_metadata_2025 huggingface dataset:
Detection of BCR-ABL transcripts from the Philadelphia translocation by hybridization in microtiter wells and time-resolved immunofluorometry
Authors: Bortolin S , Christopoulos T K
Published: January 2020

Two hybridization assays have been developed to detect BCR-ABL mRNA transcripts arising from the Philadelphia translocation. Both assays use time-resolved immunofluorometric detection of polymerase chain reaction-amplified BCR-ABL mRNA sequences hybridized to specific probes. In configuration I, biotinylated amplified target is immobilized onto streptavidin-coated wells and hybridized to a probe labeled with the hapten digoxigenin. Hybrids are detected via an alkaline phosphatase-labeled antibody and fluorosalicylylphosphate as substrate. The fluorosalicylate produced forms highly fluorescent complexes with Tb(3+)-EDTA. In configuration II, biotinylated probe is immobilized onto streptavidin-coated wells. PCR, performed in the presence of hapten-labeled deoxyribonucleotide, generates labeled product, which is hybridized to immobilized probe and quantified as above. BCR-ABL transcripts from one leukemic cell amidst mRNA from 500,000 normal granulocytes are detectable with signal/background ratios as high as 36.4 and 24.6 for configurations I and II, respectively. The respective CVs for the assays were 6.6-9.0% and 5.1-12.5%.
Advances in genosensor research
Authors: Beattie K L , Beattie W G , Meng L , Turner S L , Coral-Vazquez R , Smith D D , McIntyre P M , Dao D D
Published: January 2020

Microfabricated devices containing arrays of nucleic acid hybridization sites, known as genosensors, are being developed for a variety of uses in genomic analysis. A great deal of the overall genosensor development effort involves optimization of experimental conditions in the actual use of genosensors. Here we describe a "low-tech" form of genosensor technology, involving arrays of oligonucleotides on glass microscope slides, which can be used to define optimal operating conditions and to develop applications of hybridization arrays in genome mapping and sequencing. In addition, we describe a porous silicon genosensor, which can be operated in a flowthrough mode, and discuss its advantages over current flat-surface designs. Porous silicon genosensors containing arrays of DNA fragments offer several unique capabilities in genome analysis.
Quantification of orotic acid in dried filter-paper urine samples by stable isotope dilution
Authors: McCann M T , Thompson M M , Gueron I C , Tuchman M
Published: January 2020

A rapid, sensitive, and specific method for quantification of orotic acid from dried filter-paper urine samples is described. The method involves stable isotope dilution with 1,3-[15N2]orotic acid analysis by gas chromatography-mass spectrometry. The assay is sufficiently sensitive to be used with solvent extraction techniques commonly used for urinary organic acid analysis. Extraction efficiencies of both native and isotopic orotic acid from dried filter paper and from water were 31% and 28%, respectively. The concentration of orotic acid in dried filter-paper urine specimens from 50 healthy controls was 1.1 +/- 0.67 (mean +/- SD) mmol/mol of urinary creatinine. The same 50 urine samples, analyzed directly from a 5-mL aliquot of liquid urine, gave values of 0.93 +/- 0.51. The correlation coefficient between the results obtained by the two different collection methods was 0.87. Age-related reference values in filter-paper samples are also reported. The concentrations, which are normalized to urinary creatinine, decrease with age. This method is applicable to rapid screening for urea cycle disorders and may also be used for carrier testing of ornithine transcarbamylase deficiency.
A Step Forward in Enzymatic Measurement of Creatinine
Authors: Fossati Piero , Ponti Maurmo , Passoni Gianfranco , Tarenghi Giordano , d'Eril Gian Vico Meizi , Prencipe Lorenzo
Published: January 2020

In the article by Fossati et al. entitled "A step forward in enzymatic measurement of creatinine," 1994;40:130-7, suppliers of sera were mistakenly identified. Sera were supplied by the German Society of Clinical Chemistry. Creatinine concentrations were determined by isotope dilution-gas chromatographymass spectrometry in three laboratories (D. Stöckl, INSTAND e. V., Düsseldorf; L. Siekmann, Institut für Klinische Biochemie, Bonn; A. De Leenheer/L. M. Thienpont, Universiteit Gent, Fac. Farmac. Wetenschappen, Gent), participants in the interlaboratory comparison exercise "Creatinine in Serum" (EC M+T project 376).

In the article by Taneda and Monnier entitled "ELISA of pentosidine, an advanced glycation end product, in biological specimens," 1994;40:1766-73, line 20 of the abstract should state "2630 ± 1320 nmol/L, respectively"; on page 1770, column 1, line 17, the units should have been expressed as nmol/L, not µmol/L. The units of pentosidine in Figures 8-10 contain errors. Figure 8 should state: Pentosidine by HPLC (nmol/L plasma), not (µmol/L plasma). Figure 9 should state: Pentosidine (nmol/L plasma), not (µmol/L plasma). Figure 10 should state: Pentosidine (nmol/g collagen), not µmol/g collagen).

In the review by Kroll and Elin entitled "Interference with clinical laboratory analyses," 1994;40:1996-2005, the theophylline reagents used for the Hitachi 717 were obtained from GDS Diagnostics, Elkhart, IN. On page 1998, column 1, line 32, reference 32 was incorrectly cited; the correct reference is 37. On page 1998, column 2, line 17, reference 37 is listed in error.
Establishing a direct laboratory access program
Authors: Soloway H B
Published: January 2020

Direct Laboratory Access (DLA) refers to a program whereby individuals who wish to have laboratory testing performed can avail themselves of such testing independently of a physician referral. DLA benefits both physicians and consumers. Physicians benefit by not having to invest time and office resources for consumers who do not seek medical intervention but rather who visit physicians for the sole purpose of obtaining permission to have laboratory tests performed. Consumers benefit by avoiding physician encounters they do not want, by receiving state-of-the-art laboratory testing they do want, and by avoiding the added expense and inconvenience of a physician office visit. DLA appeals to an anxious, educated, and somewhat affluent niche market. The program fills a void in the provision of health services while providing a small stream of revenue for laboratories.
In vivo stability and disposition of a self-stabilized oligodeoxynucleotide phosphorothioate in rats
Authors: Zhang R , Lu Z , Zhang X , Zhao H , Diasio R B , Liu T , Jiang Z , Agrawal S
Published: January 2020

The use of antisense oligonucleotides represents a novel, genetically based therapy. The biostability and pharmacokinetics of a 33-mer self-stabilized oligodeoxynucleotide with significant anti-HIV activity was determined in rats after intravenous administration of [35S]oligodeoxynucleotide. Plasma disappearance of the labeled oligodeoxynucleotide could be described by a two-compartment model, with half-lives of 0.54 and 41.44 h. The oligodeoxynucleotide in plasma remained mainly intact. Urinary excretion represented the major elimination pathway, with approximately 27% of the administered dose excreted within 24 h and 57% over 240 h. The majority of radioactivity in urine was attached to degradative products. Fecal excretion was a minor elimination pathway. A wide tissue distribution of the oligonucleotide was observed, with the majority of radioactivity in most tissues being intact. Compared with other linear oligonucleotide phosphorothioates, the self-stabilized oligonucleotide was more stable in vivo, which may be important in development of antisense oligonucleotides as therapeutic agents.
Creatine kinase MM and MB isoforms in patients receiving thrombolytic therapy and acute angiography. TAMI Study Group
Authors: Christenson R H , Ohman E M , Topol E J , O'Hanesian M A , Sigmon K N , Duh S H , Kereiakes D , Worley S J , George B S , Pizzo C K
Published: January 2020

Creatine kinase isoforms markers, including MB2 concentration, MB2/MB1 and MM3/MM1 ratios, and MT index (based on the "tissue" M subunits), were measured in serial specimens from 207 patients receiving thrombolytic therapy followed by acute angiography. The slope of release showed a significant relation (P < 0.05) between MB2 concentrations and patency, graded as TIMI 0 through TIMI 3; with regard to the precatheterization/baseline ratio, the MB2 concentrations, the MM3/MM1 ratio, and the MT index were all significantly related to graded patency (P < 0.004). Patients having patency graded as either TIMI 2/3 (Open) or TIMI 0/1 (Closed) showed highly significant differences (P < 0.03) in the slope of release and precatheterization/baseline ratio for all markers except the MB2/MB1 ratio. Defining Open as TIMI 3 and Closed as TIMI 0/1/2 showed very similar results. Despite these significant differences between the Open and Closed groups after thrombolytic therapy, none of the C index calculations (areas under ROC curves) for any of the isoform markers--either alone or combined--exceeded 0.70, suggesting that these markers have limited diagnostic utility for assessing patency.
Carbohydrate-deficient transferrin and false-positive results for alcohol abuse in primary biliary cirrhosis: differential diagnosis by detection of mitochondrial autoantibodies
Authors: Bean P , Sutphin M S , Liu Y , Anton R , Reynolds T B , Shoenfeld Y , Peter J B
Published: January 2020

Primary biliary cirrhosis (PBC) is one of the few nonalcohol-induced liver pathologies that causes false positives in assays of carbohydrate-deficient transferrin (CDT) for diagnosing alcohol abuse. CDT was quantified by isoelectric focusing-immunoblotting-laser densitometry (IEF-IB-LD) analysis of serum from 117 women: 57 PBC patients, 20 alcohol abusers, and 40 healthy donors. Only 5% (3 of 57) of PBC patients were positive at the densitometric cutoff value chosen (> 90% specificity). Serum samples from 15 PBC patients were further evaluated by IEF-IB-LD and CDTect chromatography-RIA. Receiver-operating characteristic (ROC) analysis showed that IEF-IB-LD better discriminated between PBC and alcohol abuse than CDTect did. By ROC analysis, mitochondrial autoantibodies to pyruvate dehydrogenase antigen M2 detected by enzyme immunoassay yielded optimal test performance for diagnosing PBC. Of six patients falsely positive for CDT by CDTect, five (83%) tested M2-positive. Thus, abnormal CDT results should be further evaluated by mitochondrial antibody testing in patients with findings compatible with PBC.
Development and use of ELISA to quantify type II phospholipase A2 in normal and uremic serum
Authors: Dorsam G , Harris L , Payne M , Fry M , Franson R
Published: January 2020

Previously we reported that uremic plasma contained eight times more phospholipase A2 (PLA2) activity than control plasma (Costello et al., Clin Chem 1990;36:198-200). That study, however, did not distinguish between various PLA2s that could contribute to the observed increase. Therefore, we developed a sandwich ELISA to specifically quantify serum type II PLA2. By ELISA, uremic sera contained significantly more type II PLA2 than control sera (median = 1025 micrograms/L, range = 52-3320 micrograms/L vs median = 9.2 micrograms/L, range = 4.6-17.5 micrograms/L; P = 0.002). When serum samples were incubated with 1-[14C]oleate-labeled autoclaved Escherichia coli, activity was increased 14.6-fold in uremic vs normal serum, with a median of 6.5 mumol/min per liter (range 1.1-16.3) vs a control median of 0.49 mumol/min per liter (range 0.32-0.60; P = 0.002). Thus, ELISA detects about eightfold more immunoreactive type II PLA2 in uremic serum than does enzymatic analysis. Evidently, the increase in PLA2 activity previously observed in uremic plasma is primarily due to increased concentrations of type II PLA2.
Multienzyme control serum (Seraclear-HE) containing human enzymes from established cell lines and other sources. 1: Preparation and properties
Authors: Eto A , Shiki A , Chikaura Y , Oka T , Nakano N I
Published: January 2020

We have developed a new multienzyme control serum, Seraclear-HE, which was designed to function not only as an accuracy and precision control serum but also as an intermethod calibrator for unifying interlaboratory clinical enzyme data in terms of reference method values. Seraclear-HE contains as analytes the following enzymes of human origin only: aspartate aminotransferase (AST, EC 2.6.1.1) and lactate dehydrogenase (LD, EC 1.1.1.27) from erythrocytes; alanine aminotransferase (ALT, EC 2.6.1.2) from a hepatoma cell line; alkaline phosphatase (ALP, EC 3.1.3.1) from an amnion cell line; creatine kinase (CK, EC 2.7.3.2) from an embryo kidney cell line; gamma-glutamyltransferase (GGT, EC 2.3.2.2) from a macrophage cell line; and amylase (AMY, EC 3.2.1.1) from urine and saliva. The seven partly purified enzymes were lyophilized in partially delipidated human serum containing sucrose (50 g/L), pyridoxal 5'-phosphate (30 mmol/L), and other stabilizers. The material is stable for at least 2 years at temperatures < or = 10 degrees C. For two concentrations of this preparation, reference method values (mainly International Federation of Clinical Chemistry and Japan Society of Clinical Chemistry) obtained at both 30 degrees C and 37 degrees C are assigned.
Current status of US programs for training clinical laboratory scientists and anticipated impact of healthcare reform
Authors: Scott M G , Sacks D B
Published: January 2020

We surveyed directors of medical technology (MT) and postdoctoral clinical chemistry (CC) training programs and of clinical pathology (CP) and combined anatomic/clinical pathology (AP/CP) residency programs regarding the number, quality, training emphasis, and job-placement experience of trainees for 1985-95 as well as the directors' opinions on the impact of "healthcare reform." Responses were received from directors of 94 of 249 (38%) MT programs, 14 of 15 (93%) CC programs, and 63 of 138 (46%) pathology residency programs. In all four categories the numbers of trainees have increased steadily over the last 5 to 7 years but are expected to remain stable or decrease slightly. Directors of MT and CC programs expect increasing difficulty placing their graduates; directors of AP/CP and CP residency programs do not. Although > 60% of MT graduates have entered private hospitals, this is anticipated to decrease, with a concomitant increase in university hospital placements. Of the AP/CP residents, > 60% and < 5% accepted service- and research-oriented positions, respectively. In contrast, 83% of CP residents entered university hospitals, with half of these taking research-oriented positions. Among CC graduates, 41% joined university hospitals and 10-15% accepted positions in each of either private hospitals or industry or reference laboratories. The emphasis of training varies, with clinical service and pathophysiology the major focus in AP/CP programs. CP and CC programs take two distinct approaches--some accentuating management, and others emphasizing research. Finally, MT program directors appear the most optimistic regarding the opportunities that healthcare reform may present.
Clinical and analytical evaluation of an immunoradiometric assay for corticotropin
Authors: Rosano T G , Demers L M , Hillam R , Dybas M T , Leinung M
Published: January 2020

We evaluated a new IRMA developed commercially for the measurement of corticotropin (ACTH) in human plasma. The assay involves purified polyclonal goat capture antibodies specific for ACTH 26-39 and an 125I-labeled monoclonal signal antibody specific for ACTH 1-17. CVs for intraassay and total precision at ACTH concentrations between 9 and 801 ng/L ranged from 2.5% to 4.7% and from 3.3% to 9.3%, respectively, with an assay detection limit of 1.7 ng/L. The reference interval determined for adults with the new method (16-52 ng/L) differed significantly (P < 0.05) from that for an established ACTH IRMA (9-54 ng/L). Method comparison with clinical samples (n = 179) revealed a correlation coefficient of 0.970 and a best-fit equation of y (new IRMA) = (1.011 +/- 0.019)x + (4.17 +/- 3.31) with Sylx = 40.2. Both methods showed equivalent clinical sensitivity in evaluating Cushing disease, adrenal tumors, ectopic ACTH-producing tumors, hypopituitarism, steroid suppression, surgical adrenalectomy, Nelson syndrome, Addison disease, and corticotropin-releasing hormone stimulation. We conclude that the new IRMA is technically simple to perform and provides a specific and sensitive method for evaluating of adrenocortical function.
Immunoenzymometric assay of human glycogen phosphorylase isoenzyme BB in diagnosis of ischemic myocardial injury
Authors: Rabitzsch G , Mair J , Lechleitner P , Noll F , Hofmann U , Krause E G , Dienstl F , Puschendorf B
Published: January 2020

With a new immunoenzymometric assay we measured human glycogen phosphorylase isoenzyme BB (GPBB) in 116 healthy individuals, 14 patients with stable angina, 107 nontraumatic chest pain patients on admission to the emergency department [45 acute myocardial infarction (AMI), 49 unstable angina, 13 other diseases], and in serial samples from 41 AMI patients. GPBB was compared with creatine kinase (CK), CKMB mass, myoglobin, and cardiac troponin T. Receiver-operating characteristic plots demonstrated the significantly greater (P < or = 0.012) discriminatory power of GPBB to detect acute ischemic coronary syndromes compared with all other tested markers. GPBB was the most sensitive marker for detection of AMI during the first 4 h after onset of chest pain, and only GPBB was increased above the upper reference limit (7 micrograms/L) on admission in patients who had unstable angina at rest and reversible ST-T alterations. This and the high early sensitivity of GPBB are most likely explained by its function as a key enzyme of glycogenolysis.
New noncompetitive immunoassays of small analytes
Authors: Piran U , Riordan W J , Livshin L A
Published: January 2020

We developed a novel noncompetitive immunoassay format for monoepitopic analytes and describe here a model assay for triiodothyronine (T3), performed on Ciba Corning's ACS:180 analyzer. Acridinium ester (AE)-labeled bivalent anti-T3 was incubated with the sample, producing AE-anti-T3/T3 complexes and unreacted AE-anti-T3. Controlled-pore glass particles (CPG) with immobilized diiodothyronine (T2) were then added in excess, to bind AE-anti-T3 possessing two unoccupied binding sites but not AE-anti-T3 bound to one or two T3 molecules. Paramagnetic particles (PMP) with immobilized anti-AE were then added to the same cuvette to capture AE-anti-T3/T3 complexes; AE-anti-T3 bound to the surface of CPG, however, was not captured, because of steric hindrance. After the incubation, the PMP was magnetically separated to remove the liquid phase and the suspended CPG from the cuvette. The chemiluminescence associate with the PMP remaining in the cuvette was then measured. This noncompetitive T3 assay exhibited a 10-fold lower detection limit than the equivalent competitive T3 assay, i.e., 0.3 vs pg/test. Imprecision (CV) in the clinically significant range was 6% or less. The assay also displayed two- to sevenfold lower cross-reactivities and a wider dynamic range.
Dual-label one-step immunoassay for simultaneous measurement of free and total prostate-specific antigen concentrations and ratios in serum
Authors: Mitrunen K , Pettersson K , Piironen T , Björk T , Lilja H , Lövgren T
Published: January 2020

We developed a simple one-step dual-label immunoassay for simultaneous measurement of the free, noncomplexed form of prostate-specific antigen (PSA) and total PSA. The assay is based on time-resolved fluorescence and includes a stable fluorescent chelate of Eu to label a monoclonal antibody (mAb) that detects only free PSA, whereas a second mAb labeled with a fluorescent chelate of Tb provides equimolar detection of both free PSA and PSA complexed to alpha 1-antichymotrypsin. A third mAb on a solid phase captures the free and complexed forms of PSA in an equimolar fashion. The simultaneous measurement of the free-to-total PSA ratio (F/T) with the one-step dual assay is not sensitive to variations in the sample volume. The discrimination between benign prostatic hyperplasia and prostate cancer patients, i.e., the area under the receiver-operating characteristic curve, increased from 0.64 (total PSA assay) to 0.78 and 0.81 when the F/T ratio was measured with single and dual assays, respectively.
Immunometric assays of luteinizing hormone (LH): differences in recognition of plasma LH by anti-intact and beta-subunit-specific antibodies in various physiological and pathophysiological situations
Authors: Mitchell R , Hollis S , Crowley V , McLoughlin J , Peers N , Robertson W R
Published: January 2020

Restricted immunoreactivity of plasma luteinizing hormone (LH) has been described in some subjects when assayed with certain methods involving antibodies against intact LH. We have compared the performance of the Amerlite LH-30 (A) and Delfia LHSpec (D) assays (which include anti-intact and beta-specific antibodies, respectively) in normal and pathological conditions. As shown previously, results of the two systems were highly correlated with each other and, as we show here, with those of a bioassay. We found eight outliers (results outside the 95% confidence interval of the regression) among 427 samples studied from 121 subjects. Of the outliers, five had Delfia results in a range (< 1 IU/L) that was associated with poor assay precision for that assay, and the ratios of their values by both methods (A:D ratios) were very low. This ratio was affected by endocrine status, e.g., was lower in postmenopausal women than in premenopausal controls, and varied intraindividually within the same menstrual cycle. The restricted immunoreactivity described previously for assays involving anti-intact LH antibodies may, in part, reflect these differences, which, in turn, may reflect the presence of isoforms (e.g., glycoforms) that are differentially recognized by assays that have different antibody configurations.
Serial lecithin/sphingomyelin ratios and surfactant/albumin ratios in tracheal aspirates from term infants with respiratory failure receiving extracorporeal membrane oxygenation
Authors: Lotze A , Stroud C Y , Soldin S J
Published: January 2020

Serial tracheal aspirate samples were obtained for determination of lecithin/sphingomyelin (L/S) ratios from 47 term infants in respiratory failure. Phospholipids were extracted with Folch solution (chloroform:methanol, 2:1 by vol) and analyzed by HPLC with use of silica column and a mobile phase of acetonitrile:methanol:water (48:31:21, by vol). Surfactant/albumin (S/A) ratios were determined with the TDx Fetal Lung Maturity Assay (Abbott Labs). L/S ratios increased significantly over time in all patients (F = 19.42, P < 0.0001). The S/A ratio correlated with the L/S ratio (r = 0.554, P < 0.001). This study suggests that postnatal surfactant deficiency in term newborns with respiratory failure is a component of the newborn's lung injury.
Clinical Utility of Biochemical Analysis of Cerebrospinal Fluid
Authors: Watson Mark A , Scott Mitchell G
Published: January 2020

In the Review by Watson and Scott entitled "Clinical utility of biochemical analysis of cerebrospinal fluid," 1995; 41:343-60, the total protein concentration in Table 1 should have been expressed in grams per liter (g/L), not milligrams per liter.

In the Case Report by S. Coyle, M.D. Penney, P.W. Masters, and B.E. Walker entitled "Early diagnosis of ectopic arginine vasopressin secretion." 1933;39:152-4, in column 2, paragraph 3, page 152, and the title of The 1, the concentration of hypertonic saline infusion should have been expressed as 50 grams per liter (g/L), not milligrams per liter.
Clinical laboratory consultation
Authors: Burke M D
Published: January 2020

The future of healthcare delivery systems will depend on their ability to provide the best care at the least cost. In the clinical laboratory, this translates to ensuring that decreased test use means appropriate use. The latter is unlikely to occur unless clinical laboratories provide practicing physicians with advice on test strategy and interpretation of results. Effective clinical laboratory consultation requires an identification with clinical problems. Important components of the identification process are an appreciation of the probabilistic rather than deterministic nature of the relation of clinical and laboratory data to disease and the inevitable tradeoff between false negatives and false positives that clinical circumstances demand. Institution of consultation services requires that the clinical laboratory adopt a proactive approach to advocating test strategy and interpreting results. Consultation on test strategy is likely to be facilitated by further developments in practice guidelines and by advances in informatics and telemedicine.
Low plasma concentrations of retinol-binding protein in individuals with mutations affecting position 84 of the transthyretin molecule
Authors: Waits R P , Yamada T , Uemichi T , Benson M D
Published: January 2020

Retinol-binding protein (RBP), the principal carrier for vitamin A, is known to form a complex with transthyretin (TTR) for transport in plasma. Individuals from a kindred with the amino acid substitution of serine for isoleucine at position 84 (Ser84) of the TTR molecule show substantial reduction in plasma concentrations of RBP. In the present study, we measured plasma RBP in individuals from several kindreds, demonstrating 17 different point mutations within the TTR gene. In each case, these mutations caused single amino acid substitutions at various positions throughout the TTR molecule. Of all the individuals examined, only those with mutations causing amino acid substitutions at position 84 of the TTR molecule (Ser84 and Asn84) demonstrated substantial decreases in plasma concentrations of RBP. These results suggest that the isoleucine at position 84 on the TTR molecule may be critically involved in mediating RBP binding. Further, these findings demonstrate the importance of considering TTR gene mutations when clinically evaluating patients with low RBP.
HPLC-microparticle enzyme immunoassay specific for tacrolimus in whole blood of hepatic and renal transplant patients
Authors: Firdaous I , Hassoun A , Otte J B , Reding R , Squifflet J P , Besse T , Wallemacq P E
Published: January 2020

Tacrolimus is a relatively new immunosuppressant used in organ transplantation to prevent graft rejection. However, its use is not devoid of side effects, making it important to maintain blood concentrations within therapeutic ranges. Several analytical methods are currently available for routine drug monitoring. However, these methods are based on use of the same monoclonal antibody, which also cross-reacts with some metabolites, resulting in overestimation of some blood concentrations. Even though this antibody appears appropriate for therapeutic drug monitoring, no reference method measures only the parent drug, mainly because of the poor absorptivity of tacrolimus in ultraviolet light. We have developed a method displaying an increased specificity towards the unchanged drug, using conventional equipment available in most clinical laboratories. After chromatographic separation of the blood extract, the tacrolimus fraction is analyzed by an automated microparticle enzyme immunoassay (MEIA) performed on the IMx analyzer (Abbott Labs.). This method is linear from 0 to 40 micrograms/L, yields CVs from 8.5% to 18.2%, and has a detection limit of 5 micrograms/L. Tacrolimus concentrations obtained by HPLC-MEIA in hepatic and renal transplant patients are from 47.5% to 18.8% lower than those obtained by MEIA, according to liver function tests and metabolite accumulation, even though no significant differences were observed between the methods for drug-free blood samples supplemented with known amounts of tacrolimus.
Measurement of urinary oxalate and citrate by capillary electrophoresis and indirect ultraviolet absorbance
Authors: Holmes R P
Published: January 2020

We describe a method for measuring urinary oxalate and citrate with capillary electrophoresis (CE) and indirect ultraviolet absorbance detection. Sample preparation is minimal, requiring an acidification, brief centrifugation, and dilution. The method is rapid, with oxalate and citrate having mean migration times of 4.02 and 4.50 min, respectively. The minimal detectable concentration (signal-to-noise ratio of 7) of both oxalate and citrate in urine was 7 mg/L. Total imprecisions (CV) were 1.2-5.6% for three urine samples with oxalate and citrate concentrations of 8-60 mg/L and 80-860 mg/L, respectively. The recovery of added oxalate ranged from 94% to 101%. Results of CE analyses agreed well with enzymatic determinations of oxalate and citrate. Rapid analysis time, accuracy, and reproducibility make this procedure well suited for routine urinary oxalate and citrate determinations.
History of NIST's contributions to development of standard reference materials and reference and definitive methods for clinical chemistry
Authors: Schaffer R , Bowers G N , Melville R S
Published: January 2020

The issuance of cholesterol as a Standard Reference Material (SRM) in 1967 started the National Institute of Standards and Technology (NIST; then named the National Bureau of Standards) on a major effort to help clinical laboratories establish and improve the quality of measurements they make. NIST now issues three kinds of SRMs for that purpose: analyte samples of certified purity as primary standards, serum samples having certified analyte concentrations as accuracy controls, and materials certified for calibrating instruments. In working with clinical laboratory scientists to establish Reference Methods (RMs) for measuring the analytes, NIST developed Definitive Methods (DMs) to use for evaluating RM accuracy and then used the DMs for assigning analyte values to its SRMs. The development of SRMs and DMs is discussed.
Development and implementation of an expert information system (BRITE) used in technical support of medical diagnostics customers
Authors: Hopkins T , Advani R , Gudmunson G
Published: January 2020

We developed BRITE (Bringing Resources and Information to Employees), an expert information retrieval system, for problem solving and retrieving technical product information. Specialists in our Technical Assistance Center (TAC) use the BRITE system on a Pentium workstation to access information in < 3 s. Manuals, technical bulletins, parts lists, and administrative tools such as word processing and network fax are available electronically. The system allows consistent troubleshooting and definition of customers' problems by TAC specialists. BRITE seamlessly integrates expert system, document retrieval, and relational database technologies; the system runs on a token ring local area network, which is part of a Ciba Corning Diagnostics wide area network. Information received over the wide area network is used to develop and update BRITE daily. BRITE is kept current by input or troubleshooting and new applications from TAC specialists.
Laboratory computing--process and information management supporting high-quality, cost-effective healthcare
Authors: Buffone G J , Moreau D R
Published: January 2020

One currently observes many healthcare institutions rushing to reengineer and install information systems with the expectation of achieving enhanced efficiency, competitiveness, and, it is hoped, higher patient satisfaction resulting from timely, high-quality care. Unfortunately, information system concepts, design, and implementation have not yet addressed the complexity of representing and managing clinical processes. As a result, much of the synergy one might expect to derive from understanding and designing clinical processes to gain efficiency and quality while maintaining humanness is not readily achievable by implementing traditional information systems. In this presentation, with laboratory services as an example, we describe a conceptually different information systems model, which we believe would aid care-givers in their efforts to deliver compassionate, quality care while addressing the highly competitive nature of market-driven healthcare.
BibleCard: network-based virtual database for laboratory information
Authors: Fine J S , Ching A K , Schneider J B , Pollum D , Astion M L
Published: January 2020

The clinical laboratory's use of computers has evolved beyond the single minicomputer stand-alone system. Our laboratory information system is now part of an institutional network. The laboratory also uses smaller systems and workstations for a wide variety of functions, often with much data duplication among systems. We have been developing a network-based virtual database for laboratory test information. This system uses World Wide Web standards for hypertext and multimedia displays, which allows for the display of information retrieved from various department computer sources without the necessity of data duplication, modification of existing systems, or centralization of data. The medical technologists can continue to write testing procedures on their word processors. Maintenance of reference values, specimen requirements, etc., can continue as a laboratory information system function. Yet information from all of these disparate sources can be viewed in a consolidated format that has platform independence.
Analyte detection with DNA-labeled antibodies and polymerase chain reaction
Authors: Joerger R D , Truby T M , Hendrickson E R , Young R M , Ebersole R C
Published: January 2020

We demonstrate immuno-polymerase chain reaction (PCR) assays for two clinical analytes--human thyroid-stimulating hormone and chorionic gonadotropin (hTSH, hCG)--using DNA-labeled antibodies and PCR for amplification of assay response. DNA-antibody conjugates were synthesized by using heterobifunctional cross-linker chemistries to covalently attach single- or double-stranded DNA labels through amine or sulfhydryl groups on the analyte antibodies. These approaches yielded molecular chimeras possessing both analyte-specific antibody binding and nucleic acid amplification functionalities. Dose-response relationships were demonstrated for immuno-PCR assays of both analytes in a microtiter plate-based, two-antibody sandwich assay format. Detection limits for hTSH (1 x 10(-19) mol, < 1.4 mIU/L) and hCG (5 x 10(-18) mol, 0.025 IU/L) exceeded those of conventional enzyme immunoassays by 2-3 orders of magnitude. We also evaluated various DNA design factors influencing label amplification and assay performance, such as primer sequence, strand number, and DNA length. Our findings, in concert with previous reports, suggest that this hybrid technology could provide the basis for a new generation of ultra-sensitive immunoassays offering multianalyte capabilities.
Solution-phase immunoassay for determination of cortisol in serum by capillary electrophoresis
Authors: Schmalzing D , Nashabeh W , Fuchs M
Published: January 2020

We describe a competitive solution-phase immunoassay for serum cortisol determination that involves capillary electrophoresis (CE) combined with laser-induced fluorescence for separation and quantification. A polyclonal antibody preparation and fluorescein-labeled cortisol are used as assay reagents. 8-Anilino-1-naphthalenesulfonic acid was added to the serum sample during the assay to promote the release of cortisol from endogenous binding proteins. Conditions for the rapid separation of free and bound labeled antigen by CE were developed. Aspects of assay performance are evaluated in this report. The resulting assay protocol allows for the analysis of serum samples without extraction or other sample preparation steps.
Physiological variations in thyroid hormones: physiological and pathophysiological considerations
Authors: Fisher D A
Published: January 2020

Thyroid hormone production is regulated via pituitary thyrotropin (TSH) modulation of thyroxine (T4) prohormone secretion by the thyroid gland and regulation of active triiodothyronine (T3) production in peripheral tissues via metabolic events influencing activities of the iodothyronine monodeiodinase enzyme systems. Control at both levels is developmentally regulated and modified in serious nonthyroidal illness (trauma, infection, cancer, metabolic diseases). Racial and gender differences are of little significance except for the effects of placental estrogens and chorionic gonadotropin during pregnancy. There is a circadian rhythm of TSH secretion, with peak values at the onset of sleep and nadir concentrations during the afternoon hours. Peak and nadir concentrations differ by approximately +/- 50%. The effect on circulating T4 and T3 concentrations is not significant because of the large size of the extrathyroidal T4 pool. In healthy subjects there is no significant impact of body weight, physical training, body habitus, posture, immobilization, exercise, or ambulatory status on thyroid function, and no significant geographic environmental variation. Nutrition also has a minimal impact except for variation in iodine intake. Subthreshold concentrations of iodine intake are associated with increased TSH secretion, goiter, increased thyroid iodine uptake, decreased T4 production, an increased T3/T4 secretion ratio, and an increased ratio of circulating T3/T4 concentrations. Excessive iodine intake can block thyroid hormone biosynthesis by inhibiting the enzymes involved in the biosynthetic process, resulting in reduced T4 secretion, increased TSH concentrations, goiter, and hypothyroidism if the iodine excess is chronic.
Analytical performance of free and total thyroxine assays
Authors: Nelson J C , Wilcox R B
Published: January 2020

Excessive bias and imprecision are major analytical problems associated with some assays for free and total thyroxine (T4). Bias in free T4 methods is largely proportional to variations in serum T4 binding. In direct methods, this is attributable to requirements for substantial quantities of protein-bound T4 to replace analytical losses of free T4. In some total T4 methods, bias is inversely proportional to the amount of serum T4 binding and is attributable to the incomplete release of serum protein-bound T4. In others, bias is fixed and attributable to inaccurate calibration. Manufacturers should report the bias in their methods. Calibrations should be standardized. Imprecision varies widely among methods, but is generally less for total T4 methods than for free T4 methods. A consensus on quantitative analytical performance goals for free and total T4 methods would be helpful. Here, performance goals are proposed, based in part on the best achievements of current methods.
Clinical usage recommendations and analytic performance goals for total and free triiodothyronine measurements
Authors: Klee G G
Published: January 2020

The major clinical role for total triiodothyronine (TT3) and (or) free T3 (FT3) is the assessment of hyperthyroidism in patients with suppressed sensitive thyrotropin (sTSH) concentrations. The assays are particularly important in hyperthyroid patients with normal free thyroxine (FT4) concentrations to assess potential T3 thyrotoxicosis. Other specialized uses for T3 and FT3 measurements are monitoring thyroid hormone replacement therapy, the evaluation of amiodarone-induced thyrotoxicosis, and predicting outcome of antithyroid drug therapy in patients with Graves hyperthyroidism. The roles of these tests in assessing heart function in cardiopulmonary bypass surgery, evaluation of patients with neuropsychiatric disorders, and monitoring of patients on anticonvulsant therapy are not well defined. These assays are not recommended for diagnosis of hypothyroidism. Analytic recommendations include CV < 5.2% for T3 and < 3.8% for FT3; < 0.2% cross-reactivity with L-T4; and < 1.0% cross-reactivity with D-T4, D-T3, and reverse T3.
Current status and performance goals for serum thyroglobulin assays
Authors: Spencer C A , Takeuchi M , Kazarosyan M
Published: January 2020

Serum thyroglobulin (Tg) measurements are used as a tumor marker for monitoring patients with differentiated thyroid carcinoma. The clinical utility of six different Tg methods [RIA or immunometric assay (IMA)] currently used in Europe and the US was evaluated, with focus on methodologic standardization, sensitivity, interassay precision across the typical clinical monitoring interval (6 to 12 months), "hook" effects (IMA methods), and Tg autoantibody interference. The methods evaluated were: DYNOtest Tg (Henning), OptiQuant Tg (Kronus), SELco Tg (Medipan), Thyroglobulin IRMA (Pasteur), Nichols Chemiluminescent ICMA (Corning Nichols), and an RIA developed by us (USC Endocrine Services Laboratory). The clinical impact of the current methodologic problems on the use of serum Tg measurements is reviewed. Optimal performance goals are recommended for manufacturers developing and laboratories and physicians selecting a serum Tg method to use for serial long-term monitoring of thyroid cancer patients.
Modern concepts of primary thyroid gland failure
Authors: Ridgway E C
Published: January 2020

Primary thyroid gland failure is a common medical disorder occurring in mild or severe forms in 10% to 15% of our population. Symptoms may be classical and easy to recognize or very subtle, escaping clinical detection. This disorder is more common in females and increases with advancing age. The most important diagnostic test is measurement of the serum thyrotropin (TSH) concentration, which will increase above the normal range in both mild and severe cases. Most clinical effects of thyroid hormone deficiency can be explained by the "nuclear thyroid hormone hypothesis," which states that thyroid hormones act predominantly by effecting the transcription of key genes in affected tissues. Therapy of hypothyroidism is easy, inexpensive, and precise, involving pure L-thyroxine and measuring dose requirements and efficacy by monitoring serum TSH concentrations.
Blood glutathione concentrations in a large-scale human study
Authors: Richie J P , Skowronski L , Abraham P , Leutzinger Y
Published: January 2020

Little is known about the variability of blood glutathione (GSH) in human subjects. Thus, we wanted to develop and validate a rapid method for measuring GSH concentrations in whole blood and apply this method to ascertain the variation of GSH in a large-scale study of free-living adults. The assay was highly sensitive (detection limit < 5 pmol) and precise, with an interassay variation of 2.3% and a sampling variation of 3.6%. Applying this method to screen 715 adults, we observed a threefold range of GSH concentrations, with a mean of 1.02 mmol/L and CV of 17%. GSH concentrations were 8-10% greater in smokers than in nonsmokers (P < 0.001). Although we observed no sex differences for GSH, GSH/hemoglobin ratios were 8-18% greater in women < 55 years old than in older subjects (P < 0.05). The results presented here validate the use of this method for large-scale human studies and provide information on the variation and normal values of blood GSH in adults.
Logical observation identifier names and codes (LOINC) database: a public use set of codes and names for electronic reporting of clinical laboratory test results
Authors: Forrey A W , McDonald C J , DeMoor G , Huff S M , Leavelle D , Leland D , Fiers T , Charles L , Griffin B , Stalling F , Tullis A , Hutchins K , Baenziger J
Published: January 2020

Many laboratories use electronic message standards to transmit results to their clients. If all laboratories used the same "universal" set of test identifiers, electronic transmission of results would be greatly simplified. The Logical Observation Identifier Names and Codes (LOINC) database aims to be such a code system, covering at least 98% of the average laboratory's tests. The LOINC database should be of interest to hospitals, clinical laboratories, doctors' offices, state health departments, governmental healthcare providers, third-party payors, organizations involved in clinical trials, and quality assurance and utilization reviewers. The fifth release of the LOINC database, containing codes, names, and synonyms for approximately 6300 test observations, is now available on the Internet for public use. Here we describe the LOINC database, the methods used to produce it, and how it may be obtained.
Detection of Mycobacterium tuberculosis DNA with thermophilic strand displacement amplification and fluorescence polarization
Authors: Walker G T , Linn C P
Published: January 2020

Strand displacement amplification (SDA) is an isothermal, in vitro method for diagnostics that amplifies a target DNA sequence by using a restriction enzyme and DNA polymerase. We have combined a new thermophilic form of SDA that involves restriction enzyme BsoBI and polymerase exo-Bca with fluorescence polarization for detection of Mycobacterium tuberculosis DNA by using the IS6110 insertion element as the target sequence. A 5'-fluorescein-labeled oligodeoxynucleotide detector probe hybridizes to the amplified product as it rises in concentration during SDA, and the single- to double-stranded conversion is monitored through an increase in fluorescence polarization. The associated change in polarization upon amplification of the target sequence is enhanced by specific polymerase binding to the double-stranded detector probe. Fewer than 10 M. tuberculosis genomes can be amplified and detected with an extremely simple protocol that takes only 20 min and uses relatively simple instrumentation and reagents, all of which can be purchased off-the-shelf.
Standardization of pyridinium crosslinks, pyridinoline and deoxypyridinoline, for use as biochemical markers of collagen degradation
Authors: Robins S P , Duncan A , Wilson N , Evans B J
Published: January 2020

The collagen crosslinks, pyridinoline and deoxypyridinoline, have been developed as urinary markers of bone resorption but, despite wide clinical application of the technique, comparatively little attention has been paid to the standardization of these compounds. In this study, pyridinoline and deoxypyridinoline have been purified from bone and converted completely to monochloride trihydrochloride salts. In addition to mass spectrometry and NMR spectroscopy, the purity of the isolated materials was assessed by microelemental analysis including the chloride counterions. These purified compounds were used to establish individual molar absorptivity values as primary standardization criteria for the two crosslinks. For pyridinoline in 0.1 mol/L HCl, epsilon at 295 nm was 5490 L mol(-1) cm(-1); in 50 mmol/L sodium phosphate, pH 7.5, epsilon at 325 nm was 5785. The corresponding values for deoxypyridinoline at acid and neutral pH were 5160 and 5290 L mol(-1) cm(-1). The availability of standardization criteria for the crosslinks will allow more meaningful comparisons of clinical data between different laboratories.
Simple spectrofluorometric determination of p-aminobenzoic and p-aminosalicylic acids in biological fluids by use of terbium-sensitized luminescence
Authors: Lianidou E S , Ioannou P C
Published: January 2020

A novel, sensitive, and selective method has been developed for determination of p-aminobenzoic (PABA) and p-aminosalicylic (PAS) acids in the N-benzoyl-L-tyrosyl-PABA/ PAS test. PAS is measured as a ternary complex with terbium and EDTA (lambda(ex) = 324 nm, lambda(em) = 546 nm) in alkaline aqueous solution (pH approximately 12.6), whereas both compounds (PABA and PAS) are measured as ternary complexes with terbium and tri-n-octylphosphine oxide (lambda(ex) = 292 nm, lambda(em) = 546 nm) in weakly acidic aqueous solution (pH approximately 5.5). We inve stigated and implemented optimum conditions for formation of these complexes, yielding respective detection limits for PABA and PAS of 0.07 and 0.02 micromol/L and ranges of application of 0-10 and 0-40 micromol/L (final concentration). The method has been successfully applied to determinations of PABA and PAS in urine and, after alkaline hydrolysis, to determinations of PABA in serum that has been deproteinized with acetonitrile. Within-run imprecision of the PABA determination ranges from 0.8% to 4.2 % for urine samples and from 3.9% to 8.2% for serum samples; day-to-day imprecision varies from 3.2% to 10% for serum samples.