bluuebunny/crossref_metadata_2025_split huggingface dataset:
Role of antibody valency in hapten-heterologous immunoassays
Authors: Piran U , Silbert-Shostek D , Barlow E H 
Published: January 2020

Abstract
We studied the effects of hapten heterology on immunoassays of triiodothyronine (T3), digoxin, and cortisol, in a format involving labeled monoclonal antibodies and immobilized, protein-conjugated ligands. Replacing the homologous conjugated ligands T3, digoxin, and cortisol with their respective analogs diiodothyronine, digitoxin, and corticosterone led in each case to a decrease in the midpoint of displacement (ED50) for the same zero-dose signal. The mechanism of this phenomenon was studied by converting the bivalent anti-T3 to a monovalent whole antibody (bispecific monoclonal anti-T3 x anti-glucose-6-phosphate dehydrogenase) by cell fusion. The monovalent antibody was effective as a tracer in the homologous T3 assay, but generated a very low zero-dose signal with the heterologous solid phase, thus precluding sensitivity enhancement. On the basis of these results and additional kinetic and double-labeling experiments, we propose that the use of hapten heterology relies on bivalent binding of the antibody to the solid phase to compensate for a lower intrinsic affinity. This binding mechanism leads to lower assay concentrations of the ternary complex analyte-labeled antibody-immobilized hapten, thereby providing enhanced sensitivity.
Medically significant concentrations of prostate-specific antigen in serum assessed
Authors: Bernstein L H , Rudolph R A , Pinto M M , Viner N , Zuckerman H 
Published: January 2020

Abstract
Vol. 36
p. 517. In the article by L.H. Bernstein, R.A. Rudolph, M.M. Pinto, N. Viner, and H. Zuckerman entitled "Medically significant concentrations of prostate-specific antigen in serum assessed," 1990;36:515-8, the text describing Table 4 reports reversed values for sensitivity and positive predictive value and likewise for specificity and negative predictive value. The text should read, "Table 4, a reorganized Table 3, defines the occurrence of binary class patterns with respect to carcinoma of the prostate by stage compared with nondisease. The table allows the estimation of 44.4% sensitivity, 97.1% specificity, a negative predictive value of 79%, and a positive predictive value of 87.8% for PSA &gt;22.8 ng/mL."
Vol. 38
p. 2557. In the latter to the editor by J.D. Mitchell, B.J. Perrigo, and V.A. Mason-Daniel entitled "Falsely negative urine drug assay results due to filtration," 1992;38:2556-7, the units in the first paragraph at the top of p. 2557 should have been micrograms per liter (µg/L), not pg/L.
Biological variability in concentrations of serum lipids: sources of variation among results from published studies and composite predicted values
Authors: Smith S J , Cooper G R , Myers G L , Sampson E J 
Published: January 2020

Abstract
To obtain the best estimates of the average intraindividual biological variability (CVb) in the concentrations of total cholesterol (TC), low-density lipoprotein cholesterol (LDLC), high-density lipoprotein cholesterol (HDLC), and triglyceride serum lipids in a person's blood, we evaluated results from 30 studies published from 1970 to 1992. The usually more applicable random-effects model estimated an average CVb of 6.1% for TC, 7.4% for HDLC, 9.5% for LDLC, and 22.6% for triglyceride. Composite estimates of the average CVb from all evaluated published studies by different models of estimation ranged from 6.0% to 6.4% for TC, from 6.2% to 7.5% for HDLC, from 7.0% to 9.6% for LDLC, and from 22.4% to 22.9% for triglyceride. Two important factors influenced the reported biological variation of the study subjects: (a) the magnitude of the variability of the analytical method used and (b) the design characteristics of the study--primarily the number of subjects, the sampling interval, and the number of measurements per subject. For TC, we found a statistically significant positive correlation between the reported mean CVb and both the number of study subjects and the analytical variation. For TC and LDLC we estimate CVb as a function of the study design features. The number of patient specimens required to obtain reliable estimates for serum lipid concentrations are determined from the CVb and the current analytical variation.
Evaluation of colorectal cancer-associated mucin CA M43 assay in serum
Authors: van Kamp G J , von Mensdorff-Pouilly S , Kenemans P , Verstraeten R , Yedema C A , Wobbes T , Bon G G , Kok A , Brown J P , Hilgers J 
Published: January 2020

Abstract
The technical performance of a newly developed assay for CA M43, a serum marker for colorectal cancer, was evaluated and its preliminary clinical potential assessed. The heterologous double-determinant enzyme immunoassay for the detection of the tumor-associated mucin CA M43 utilizes two monoclonal antibodies (CT 43 and CT 66) selected for their binding capacity to two distinct epitopes present on mucins in the sera of patients with colorectal cancer. CT 66 recognizes both Lewis(a) and sialylated Lewis(a) antigen; CT 43 is directed toward a mucin epitope of an as-yet uncharacterized structure. Precision experiments revealed interassay CVs of 11.8%, 5.9%, and 4.9% at 9.3, 11.9, and 78.9 units/mL, respectively; intraassay precision was 2.0% at 95.1 units/mL. The upper normal value was set at 7.5 units/mL, which included 99% of the values found in healthy controls. In colorectal cancer patients, CA M43 showed a positivity rate equivalent to that of carcinoembryonic antigen (CEA) and superior to that of CA 19.9, with only one CA 19.9-positive serum being negative for CA M43. Interestingly, CA M43 appeared to be complementary to CEA, with CA M43 and CEA together reaching 87% positivity in metastatic disease.
Analytical evaluation of i-STAT Portable Clinical Analyzer and use by nonlaboratory health-care professionals
Authors: Jacobs E , Vadasdi E , Sarkozi L , Colman N 
Published: January 2020

Abstract
We evaluated the performance of the i-STAT Portable Clinical Analyzer, a hand-held instrument that, with its current cartridge, analyzes for electrolytes, urea nitrogen, glucose, and hematocrit in approximately 60 microL of whole blood in approximately 90 s. Accuracy, imprecision, and linearity studies were performed with aqueous controls and standards and by split-sample analysis. Intrarun imprecision (CV) ranged from 0.34% to 3.97%. Total imprecision over a 2-month period ranged from 0.42% to 4.83%, with urea nitrogen and glucose analyses generating the higher values. Patients' results from the Portable Clinical Analyzer correlated well with those obtained for whole blood or plasma by the Nova Stat Profile 5, the Beckman Synchron CX3, or the Technicon H1 Hematology Analyzer, with Sylx values &lt; 0.2 mmol/L for potassium; &lt; 1.5 mmol/L for sodium, glucose, and urea nitrogen; &lt; 2.4 mmol/L for chloride, and &lt; 2.4% for hematocrit. We also ascertained imprecision and accuracy of the system placed in a cardiothoracic intensive-care unit and operated by nurses. There were no significant differences in either the imprecision or accuracy of the system in this setting. We conclude that operator technique is not a factor in the analytical performance of the system and that it can be used by nonlaboratorians with a high degree of confidence that reliable results will be obtained.
Reported effects of long-term freezer storage on concentrations of retinol, beta-carotene, and alpha-tocopherol in serum or plasma summarized
Authors: Comstock G W , Alberg A J , Helzlsouer K J 
Published: January 2020

Abstract
A literature review of the effects of long-term freezer storage on concentrations of antioxidant micronutrients in serum or plasma showed that a high proportion of the studies that compared the concentrations of retinol, beta-carotene, or alpha-tocopherol in fresh and stored serum were deficient because of small numbers of observations, imprecise descriptions of procedures, and short periods of storage. Data from nested case-control studies of the associations of these micronutrients with cancer are confounded by differences in the study populations. Nevertheless, the overall impression is strong that retinol is stable at storage temperatures as warm as -20 degrees C for at least 15 years, that moderate losses of alpha-tocopherol occur at temperatures above -40 degrees C during that period, and that only a small proportion of beta-carotene persists at storage temperatures above -40 degrees C. At -70 degrees C or colder, all three micronutrients appear to be stable for at least 15 years.
Commercial kits for 1,25-dihydroxyvitamin D compared with a liquid-chromatographic assay
Authors: Bertelloni S , Baroncelli G I , Benedetti U , Franchi G , Saggese G 
Published: January 2020

Abstract
Three commercially available kits for 1,25-dihydroxyvitamin D [1,25(OH)2D] determination were compared with a liquid-chromatographic method. Serum samples were analyzed for 1,25(OH)2D from 70 healthy subjects (age 2.2-17.5 years; 35 males, 35 females), some (n = 5) given orally high vitamin D3 doses. In addition, 1,25(OH)2D was measured in 28 patients with untreated diseases associated with low (n = 16) or high (n = 12) serum concentrations of the hormone. The results showed that the commercial kits were sufficiently accurate with respect to the comparison method and suitable for routine clinical work.
Sequential ultracentrifugation micromethod for separation of serum lipoproteins and assays of lipids, apolipoproteins, and lipoprotein particles
Authors: Brousseau T , Clavey V , Bard J M , Fruchart J C 
Published: January 2020

Abstract
We describe a fast sequential separation of very-low-density, low-density, and high-density lipoproteins from 400 microL of serum, using the Beckman TL100 ultracentrifuge. The cumulative centrifugation time is 9.5 h. The purity of lipoprotein fractions was verified by a gel-filtration procedure. The major contaminant is the serum albumin, which can be eliminated by a second centrifugation at the same density. Enzymatic measurement of lipids shows good recovery (&gt; 91%) and weak within-sample variation (&lt; 7%). In comparison with a density-gradient procedure, the deleterious effects on the lipoprotein structure appear to be limited, as shown by the low concentrations of apolipoprotein (apo) E and apo A-I in the fraction &gt; 1.21 kg/L. Furthermore, the micro-ultracentrifugation also gives a better recovery rate. Finally, we have studied the distribution of lipids, apolipoproteins, and lipoprotein particles (LpA-I:A-II, LpB:C-III, LpB:E) in each fraction separated from 10 serum samples from healthy subjects.
Redox status and protein binding of plasma aminothiols during the transient hyperhomocysteinemia that follows homocysteine administration
Authors: Mansoor M A , Guttormsen A B , Fiskerstrand T , Refsum H , Ueland P M , Svardal A M 
Published: January 2020

Abstract
We administered reduced L-homocysteine perorally (67 mumol/kg of body wt) to 12 healthy subjects and injected the same dose into one person, and determined the kinetics of the alterations in reduced, oxidized, and protein-bound concentrations of homocysteine, cysteine, and cysteinylglycine. After oral intake, reduced homocysteine increased rapidly (tmax &lt; or = 15 min), reaching concentrations [3.97 (SD 2.99) mumol/L] 20-fold above fasting values, and then declined towards the normal concentration within 2 h. There was a similar increase in reduced cysteine and a moderate increase in reduced cysteinylglycine. During this response, we observed a positive correlation between the reduced/total ratio for homocysteine and cysteine. When homocysteine was injected, the increase in reduced homocysteine preceded the increase in reduced cysteine by about 3 min. After oral loading, oxidized homocysteine showed a transient increase (tmax = 30 min) that lagged behind the increase of reduced homocysteine. Oxidized cysteine and cysteinylglycine were stable or decreased slightly. Protein-bound homocysteine increased the least rapidly after homocysteine administration (tmax = 1-2 h), and returned to normal values slowly. Changes in protein-bound homocysteine essentially mirrored a concurrent decrease in protein-bound cysteine, suggesting displacement of bound cysteine. These data show that plasma homocysteine has a pronounced, direct effect on the redox status and protein binding of other plasma thiol components. Such effects should be recognized when studying the mechanisms behind the atherogenic effect of increased plasma homocysteine.
Quantification of lipoprotein(a) particles containing various apolipoprotein(a) isoforms by a monoclonal anti-apo(a) capture antibody and a polyclonal anti-apolipoprotein B detection antibody sandwich enzyme immunoassay
Authors: Taddei-Peters W C , Butman B T , Jones G R , Venetta T M , Macomber P F , Ransom J H 
Published: January 2020

Abstract
A quantitative sandwich ELISA for lipoprotein(a) [Lp(a)], utilizing a monoclonal capture antibody that recognizes human and rhesus monkey apolipoprotein(a) [apo(a)] isoforms in combination with a polyclonal anti-apolipoprotein B-peroxidase conjugate was developed. This assay generates a linear calibration curve from 31.2 to 1000 mg/L, is highly reproducible (intra- and interassay CV of &lt; 5% and &lt; or = 12%, respectively), and shows no interference from plasminogen (1 g/L), low-density lipoprotein (6.00 g/L), triglycerides (27.00 g/L from chylomicrons and 10.00 g/L from very-low-density lipoprotein), hemoglobin (5 g/L), or bilirubin (30 mg/L). This assay format quantifies the concentration of Lp(a) on an equal molar basis regardless of apo(a) isoform. In contrast, a commercially available ELISA [Macra Lp(a)] method with a monoclonal anti-apo(a) capture antibody and a polyclonal anti-apo(a) conjugate was found to underestimate the Lp(a) concentrations of individuals with lower-M(r) apo(a) isoforms--whether quantifying the Lp(a) in plasma or the purified lipoprotein. This demonstrates the importance of assay format selection in quantifying Lp(a).
Simple method for extracting RNA from cultured cells and tissue with guanidine salts
Authors: Zolfaghari R , Chen X , Fisher E A 
Published: January 2020

Abstract
We have developed a simple protocol for isolating RNA from both cell culture and tissue from human and animal sources, using guanidine thiocyanate and guanidine hydrochloride, but no organic solvents. The protocol reproducibly yielded 15 to 25 micrograms of high-quality RNA per 10(6) cells of human and animal origin and 1 to 1.1 mg of RNA per gram of human placental tissue. The RNA so obtained was ribonuclease-free and not contaminated by DNA. It was suitable for reverse transcription-polymerase chain reaction, Northern blot analysis, and in vitro expression of proteins. Thus, the molecular assessment of both research and clinical samples can be readily and reliably initiated by the application of this protocol.
Enzyme immunoassay of beta-hexosaminidase A and B in serum: carrier detection of GM2-gangliosidoses, and equivalence of enzyme activity and enzyme protein reactivity
Authors: Isaksson A , Hultberg B , Masson P , Landels E , Fensom A 
Published: January 2020

Abstract
beta-Hexosaminidase (Hex; EC 3.2.1.52) isoenzymes A and B were analyzed in sera from a control group of 22 apparently healthy subjects, 13 obligate carriers of Tay-Sachs disease (TSD), 10 obligate carriers of Sandhoff disease (SHD), and 4 affected TSD patients by enzyme immunoassay methods based on enzyme activity. No Hex A activity was detected in the sera of patients with TSD. The activities of Hex A in the obligate carriers of TSD and SHD tended to be lower (nonsignificantly) than in the control group. Hex B activities tended to be higher in TSD patients as well as in carriers of TSD, although the mean activities did not significantly differ from the corresponding mean for the control group. However, Hex B activities were decreased in the carriers of SHD in comparison with the other groups. Sera from 900 postmenopausal women, all of age 55 years, were also analyzed for Hex isoenzymes; the results indicated a carrier frequency of about 1 in 200 for both TSD and SHD. We also compared the enzyme immunoassay method based on enzyme activity with one based on the antigenic (enzyme protein) reactivity alone. Because both methods yielded similar information, we conclude that no significant amounts of inactive enzyme protein are present in the circulation.
Governmental perspectives on evaluating laboratory performance
Authors: Boone D J 
Published: January 2020

Abstract
The quality of laboratory analytical performance required to support medical decision-making has been defined in four major ways: (a) by the analytical variance of the state of the practice; (b) by the total variance, including analytical and biological variability; (c) by the loss of diagnostic efficiency attributable to analytical error; and (d) by medical-usefulness criteria. From the federal government's perspective, the answer to the question "How good must a laboratory test result be to be medically relevant?" must take into account the clinical context of the test, with accompanying concerns about access, timeliness, and cost, as well as limits for precision and accuracy in the analytical process and the frequency and potential patient-care impact of error in the pre- and postanalytical steps of the total testing process. Therefore, medically relevant goals should encompass not only analytical precision and accuracy but also goals to provide access to clinically effective tests and to reduce errors in the total testing process that can lead to medically misleading information. Development of more appropriate regulatory requirements for laboratories, as well as any needed improvements in instrumentation and methodology, should focus on ensuring that goals for medically relevant results are met by appropriate design and management of the entire process of laboratory testing.
Application of gene transfer technologies to the production of enzyme reference materials: example of gamma-glutamyltransferase
Authors: Siest G , Oster T , Visvikis A , Thioudellet C , Angèle C , Schiele F , Wellman M 
Published: January 2020

Abstract
Protein reference materials are traditionally prepared by purification from mammalian or human tissues. The supply of these tissues is limited; consequently, there is a growing need for applied molecular and cellular biology technologies for the production of human recombinant proteins. This is especially true when only small amounts of the proteins are available in the tissues. We review the current knowledge necessary for high-level production of such proteins in different heterologous expression systems, using our data on gamma-glutamyltransferase (EC 2.3.2.2) as an example. We describe the steps required to achieve the expression of enzymes and other proteins in Escherichia coli, yeast, or mammalian cells. We list many of the problems investigators may face in preparing recombinant proteins, and provide information on selecting the most appropriate system as well as the most favorable experimental conditions. Depending on the expression system, recombinant proteins can potentially be obtained for most, if not all, enzymes of interest in clinical chemistry, and such proteins should possess characteristics very similar to those of the corresponding human native proteins. Studies suggest that these products can be used as reference materials in clinical chemistry laboratories.
Homocysteine and other thiols determined in plasma by HPLC and thiol-specific postcolumn derivatization
Authors: Andersson A , Isaksson A , Brattström L , Hultberg B 
Published: January 2020

Abstract
We describe a versatile high-performance liquid-chromatographic method for determining homocysteine and other plasma sulfhydryls. Using three different procedures for preparation of plasma, we determined total, free (non-protein-bound), and reduced forms of homocysteine, cysteine, glutathione, cysteinylglycine, and gamma-glutamylcysteine in human plasma. Sample preparation involves disulfide reduction with dithiothreitol and protein precipitation with sulfosalicylic acid. The assay utilizes isocratic reversed-phase ion-pair liquid chromatography at pH 2.4, postcolumn derivatization with 4,4'-dithiodipyridine, and colorimetric detection at 324 nm. The intra-assay precision (CV) of the method for total homocysteine is 1.5%; the interassay precision over 2.5 months is 2.5%. The detection limit for homocysteine is &lt; 50 nmol/L plasma.
Determination of clozapine, norclozapine, and clozapine-N-oxide in serum by liquid chromatography
Authors: Volpicelli S A , Centorrino F , Puopolo P R , Kando J , Frankenburg F R , Baldessarini R J , Flood J G 
Published: January 2020

Abstract
We report a new assay to measure the serum concentrations of the atypical antipsychotic drug clozapine and two major metabolites, norclozapine and clozapine-N-oxide. The analytes and an internal standard (triprolidine) were extracted from alkalinized samples into ethyl acetate and back-extracted into 0.1 mol/L HCl. The acid extracts were chromatographed on a reversed-phase liquid chromatographic column with photodiode array detection (210-340 nm). With the 254-nm signal, between-run imprecision (CV) was &lt; 2% for clozapine and norclozapine at 400 micrograms/L, and 4.1% for clozapine-N-oxide at 100 micrograms/L. Absolute recovery exceeded 65%, and the detection limit was approximately 3-4 micrograms/L. In 25 patients at steady state at a mean daily clozapine dosage of 269 mg (3.09 mg/kg), clozapine averaged 231 +/- 144 micrograms/L (mean +/- SD); norclozapine and clozapine-N-oxide concentrations averaged 84% and 23% that of clozapine. Analyte concentrations were significantly correlated with daily dose. The method's ability to quantify clozapine and two major metabolites simultaneously with precision and sensitivity makes it useful in pharmacokinetic studies and therapeutic monitoring.
Free thyroxine assessed with three assays in sera of patients with nonthyroidal illness and of subjects with abnormal concentrations of thyroxine-binding proteins
Authors: Docter R , van Toor H , Krenning E P , de Jong M , Hennemann G 
Published: January 2020

Abstract
Three methods for estimating free thyroxine (FT4) in serum were studied: equilibrium dialysis, the SPAC-ET FT4 radioimmunoassay kit, and the Amerlite MAB FT4 luminometric assay. Serum samples from 10 subjects with above-normal thyroxine-binding globulin (TBG), 6 with low TBG, 30 with familial dysalbuminemic hyperthyroxinemia (FDH), 13 with nonesterified fatty acids (NEFA) concentrations in serum &gt; 1.0 mmol/L, and 178 patients with various degrees of nonthyroidal illness (NTI) were measured and compared with samples from 42 euthyroid blood donors. The Amerlite MAB FT4 assay compared well with equilibrium dialysis, whereas the SPAC-ET assay averaged 40% lower. All three assays were not influenced by changes in TBG and showed no or only little changes in the presence of NEFA. Mean FT4 values in the FDH samples were somewhat higher than in controls when measured with the SPAC-ET assay, about equal with equilibrium dialysis, and somewhat below the mean control value with the Amerlite MAB FT4 assay, although individual results were within the control reference range. In NTI patients, no FT4 values were below the control reference range by the Amerlite MAB FT4 assay, 4 of 178 were below this range by equilibrium dialysis, and 1 of 178 was below this range by the SPAC-ET assay. In all assays a large proportion of the NTI samples showed FT4 values above the control reference range, a result that will interfere with the efficacy of these assays for assessing thyroid function in NTI patients.