bluuebunny/crossref_metadata_2025 huggingface dataset:
Modified Method for Determining Carcinoembryonic Antigen in the Presence of Human Anti-Murine Antibodies
Authors: Morrissey Nancy E , Quadri Syed Farhat , Kinders Robert , Brigham Christine , Rose Steve , Blend Michael J
Published: January 2020

Vol. 39:

p. 527. In the article by N. E. Morrissey, S. F. Quadri, R. Kinders, C. Brigham, S. Rose, and M. J. Blend entitled "Modified method for determining carcinoembryonic antigen in the presence of anti-murine antibodies," 1993;39:522-9, the graphs A and B in the left-hand column of page 527 should be exchanged with graphs A and B in the right-hand column, so that the legend for Figure 2 refers to graphs for two HAMA-negative patients and the legend for Figure 3 refers to three HAMA-positive patients.

p. 1401. In the article by J. M. Queraltó, J. C. Boyd, and E. K. Harris entitled "On the calculation of reference change values, with examples from a long-term study," 1993;39:1398-403, the last two columns of Table 4 are incorrect: in the next-to-last column, a misprint occurred in the line for sodium; in the last column, a number was omitted, causing other numbers to be misplaced. The columns should have read as follows:

See table in the PDF file

p. 1901. In the Scientific Note by R. G. Parsons, R. Kowal, D. LeBlond, V. T. Yue, L. Neargarder, L. Bond, D. Garcia, D. Slater, and P. Rogers, entitled "Multianalyte assay system developed for drugs of abuse," 1993;39:1899-903, the word "trihexylphenidyl" in line 1 of the text in column 2, page 1901, should read "trihexyphenidyl."

p. 1942. In Oak Ridge Conference paper by R. Devlin, R. M. Studholme, W. D. Dandliker, K. Blumeyer, and S. S. Ghosh, entitled "Homogeneous detection of nucleic acids by transientstate polarized fluorescence," 1993;39:1939-43, the x-axis for Figure 5 should read: "Volume of 3SR product solution (1O-4 x µL)," not (10-3 x µL).

p. 1982. In the Oak Ridge Conference Poster Session, the paper by D. Crisan, M. J. Anstett, N. Matta, and D. H. Farkas entitled "Detection of bcl-2 oncogene rearrangement in follicular lymphoma: nucleic acid hybridization and polymerase chain reaction compared," 1993;39:1980-2, the word "bone" in the first line at the top of page 1982 should have read "bone marrow."
Effect of thyroid function on concentrations of lipoprotein(a)
Authors: Engler H , Riesen W F
Published: January 2020

The effect of thyroid hormones on concentrations of lipoprotein(a) [Lp(a)] was analyzed in 60 patients with active thyroid dysfunction (hyperthyroidism 30 cases, hypothyroidism 32 cases, and 2 cases with opposite changes) and after normalization of the thyroid state. Treatment of hyperthyroidism increased the mean Lp(a) concentrations by 60% (from 73 to 102 mg/L, P < 0.002); at the same time, low-density lipoprotein cholesterol (LDL-C) increased by 53% (from 2.6 to 3.7 mmol/L, P < 0.0001) and apolipoprotein B (apo B) by 35% (from 0.91 to 1.17 g/L, P < 0.0005). In hypothyroidism, the opposite changes were observed: mean Lp(a) decreased from 136 to 114 mg/L (10%, P < 0.02), LDL-C from 4.6 to 3.9 mmol/L (13%, P < 0.01), and apo B from 1.51 to 1.20 g/L (14%, P < 0.01). Although the changes in Lp(a) concentrations did correlate with changes of LDL-C during treatment of hyperthyroidism (r = 0.43, P < 0.05), and with changes in apo B during thyroxine-substitution therapy for hypothyroidism (r = 0.46, P < 0.05), we observed no associations between Lp(a) and LDL-C or apo B in the euthyroid state. These data cannot rule out the possibility that the thyroid hormone-induced increase in LDL-C receptor activity was responsible for the decreased concentrations of Lp(a) in hyperthyroidism. Given that LDL-C is approximately 30% of the Lp(a) molecule but the changes in Lp(a) concentrations are comparable with those in LDL-C (60% vs 53%), and given that Lp(a) is metabolized by an LDL-C-receptor-independent pathway, the present data suggest a direct effect of thyroid hormones on Lp(a) synthesis.
Combined serum amylase and lipase determinations for diagnosis of suspected acute pancreatitis
Authors: Corsetti J P , Cox C , Schulz T J , Arvan D A
Published: January 2020

Serum amylase and lipase measurements are often used to diagnose acute pancreatitis. This study addresses the question of whether it is advantageous to order serum amylase and lipase tests simultaneously. We evaluated performance of the two tests separately and in combination through a retrospective study of patients for whom both amylase and lipase determinations were ordered. Initial analysis of test performance was conducted with a uniformly applied criterion based on determination of optimal sensitivity-specificity pairs. Individual tests and combinations of tests, including the "AND" and "OR" rules and discriminant functions, were examined. Only the discriminant approach demonstrated better performance than the lipase test alone. This finding was subsequently confirmed by logistic regression analysis. We conclude that ordering both tests simultaneously can be advantageous in diagnosing acute pancreatitis when a bivariate approach is used; however, this must be weighed against the difficulties associated with clinical implementation of such approaches.
Densitometric determination of urinary 4-hydroxy-3-methoxymandelic acid (vanillylmandelic acid)
Authors: Agbaba D , Stojanov M , Rajacic S , Zivanov-Stakic D , Majkic-Singh N
Published: January 2020

We propose a simple and precise densitometric method for measuring vanillylmandelic acid (VMA). The method comprises direct urine application, thin-layer chromatographic (TLC) separation, postchromatographic derivatization, and in situ reading of the formed derivative at 560 nm. Quantitation was done by measuring peak area with a computer-controlled TLC Scanner and applying a five-point calibration function. The calibration function was 7.57-49.23 mumol/L. This method was used to determine urinary concentrations of excreted VMA; concentrations ranged from 4.84 to 56.06 mumol/day.
Free cortisol in serum assayed by temperature-controlled ultrafiltration before fluorescence polarization immunoassay
Authors: Lentjes E G , Romijn F , Maassen R J , de Graaf L , Gautier P , Moolenaar A J
Published: January 2020

A method is described for a temperature-controlled ultrafiltration procedure to measure free cortisol in serum. A special thermometer with a sensor was developed, measuring the temperature directly in the ultrafiltration device. The sensor is screwed on the axis of the centrifuge rotor, and the centrifuge is placed in a temperature-controlled box so that the temperature of the sample is kept at 37 degrees C +/- 0.1 degrees C. The overall CV of the free cortisol assay ranges from 2.2% to 11.4%, of which the ultrafiltration contributes only 2.2-3.6%. Increasing amounts of cortisol-binding protein, as found in women using estrogen-containing oral contraceptives, have minor but significant effects on the free cortisol concentrations in serum. Serum free cortisol concentrations in a reference population (n = 114; central 95 percentiles) were 12-43 nmol/L (4-9.5% of total cortisol); in the group of the oral-contraceptive users (n = 27), the reference interval was 11-53 nmol/L (1.5-4.5%).
Urinary Oligosaccharides In Pregnant or Lactating Women: Pitfall in Screening
Authors: Kuczynski Thaddeus W , Kendzierski Kevin S , Sewell Adrian C
Published: January 2020

Vol. 39

p. 2347: In the letter by Thaddeus W. Kurczynski, Kevin S. Kendzierski, and Adrian C. Sewell entitled "Urinary oligosaccharides in pregnant or lactating women: pitfall in screening," 1993;39:2346-7, the name of the first author was misspelled.
Modifying an enzyme immunoassay of immunoreactive trypsinogen to use time-resolved fluorescence
Authors: Ryall R G , Gjerde E M , Gerace R L , Ranieri E
Published: January 2020

A coated microtiter-well, enzyme-linked immunometric assay for quantifying immunoreactive trypsinogen in dried blood spots was modified to use time-resolved fluorescence of europium in place of end-point enzymatic color development as the quantification step. The streptavidin-horseradish peroxidase and color development solutions supplied as packaged reagents were replaced by europium-labeled avidin, and the signal was developed with commercially available enhancement solution and read by time-resolved fluorescence. The change of label from enzyme to europium increased the dynamic range of the assay by about 5-fold, reduced the detection limit 10-fold, and halved the intra- and interassay imprecision. The improved analytical precision and stability of the modified assay resulted in a more precise description of the population distribution of immunoreactive trypsinogen values in newborns, showing less variance in the upper centiles. This effect is of paramount importance when using this assay for neonatal screening for cystic fibrosis.
Multiwavelength photometry of thermochromic indicator solutions for temperature determination in multicuvettes
Authors: Schilling K , Cumme G A , Hoffmann-Blume E , Hoppe H , Horn A
Published: January 2020

If buffer/indicator systems are used as optical thermometers in multicuvettes, the temperature resolution is limited by errors of optical measurements produced mainly by variations of pathlength and blank transmittance of the wells. Theoretical and practical approaches show that, in multicuvettes, a between-well temperature resolution of < 0.05 degrees C can be achieved by multiwavelength photometry with use of the Tris/cresol red indicator system. Using up to three absorbances (A0, A1, A2) measured in the same well at different wavelengths for calculation of differences, quotients, and quotients of differences, we found the optimum temperature signal to be In(A1/A2), with equal-ranking absorbances changing with temperature in opposite directions. We have used the method successfully to measure the temperature profiles and temporal dynamics of temperatures at all positions in two types of multicuvettes under controlled conditions, or not controlled thermostatically.
Monitoring both serum amyloid protein A and C-reactive protein as inflammatory markers in infectious diseases
Authors: Nakayama T , Sonoda S , Urano T , Yamada T , Okada M
Published: January 2020

We examined serum amyloid protein A (SAA) and C-reactive protein (CRP) as inflammatory markers of viral and bacterial infections. Both acute-phase reactants increased in the acute stage and thereafter decreased in the convalescent stage. In viral infections, the mean serum concentrations of SAA during the acute stage were 141 mg/L in infections with adenovirus, 77 mg/L with measles virus, 63 mg/L with influenza virus, 55 mg/L with parainfluenza virus, 31 mg/L with respiratory syncytial virus, and 31 mg/L in aseptic meningitis. The mean serum concentration of CRP was 19 mg/L for adenovirus infection and < 7 mg/L in all other viral infections. The SAA concentrations were 5- to 11-fold greater than the CRP concentrations. Both the SAA and the CRP concentrations were higher in bacterial infections than in viral infections. Changes in the concentrations of serum SAA paralleled those in serum CRP in bacterial infection; during the course of viral infection, however, serum SAA tended to disappear more quickly than CRP did. SAA appears to be a clinically useful marker of inflammation in acute viral infections, with or without significant changes in the CRP concentration.
Total error assessment of five methods for cholesterol screening
Authors: Miller W G , McKenney J M , Conner M R , Chinchilli V M
Published: January 2020

We report the accuracy, imprecision, total analytical errors, and patient misclassification errors for cholesterol measured from capillary whole blood, venous whole blood, and venous plasma samples by five devices used in public cholesterol screening environments: Reflotron, Vision, Ektachem DT-60, QuickRead, and Liposcan. None of the methods met the National Cholesterol Education Program (NCEP) performance recommendations of 3% CV with 3% bias. The Vision and Reflotron methods used with venous samples gave individual results with total errors consistent with a combined CV and bias in the 4-5% range; capillary blood samples had total errors > 5% (combined CV and bias criteria). The DT-60 performance was near the 5% total error criterion for capillary samples and was > 5% for venous samples. Misclassification of individuals into desirable or referral groups for venous samples was as great as 5.1% for the DT-60, 5.7% for the Vision, and 7.1% for the Reflotron. Misclassifications for capillary blood samples were as great as 6.7%, 18.3%, and 14.1% for DT-60, Vision, and Reflotron, respectively. The QuickRead and Liposcan results were substantially poorer than those obtained by the other methods.
Blood alcohol testing in the clinical laboratory: problems and suggested remedies
Authors: Frajola W J
Published: January 2020

Laboratory directors or others responsible for blood alcohol testing are advised to reevaluate their procedures for alcohol tests. The demand for better traffic safety has resulted in a large increase in the number of arrests for drinking and driving violations and in a concomitant increase in the litigation of alcohol-related cases. My experience in the last 20 years indicates that many difficulties in the forensic aspects of these cases and many miscarriages of justice could have been reduced or even avoided if laboratories doing alcohol analyses had followed established guidelines. The most frequently encountered problems involve the physician-patient privilege ("the test was done for medical purposes only") and the fact that alcohol results were frequently reported as blood alcohol concentrations when they were, in fact, serum alcohol concentrations. Here I discuss these problems and suggest some solutions.
Liposome-based flow-injection immunoassay for determining theophylline in serum
Authors: Locascio-Brown L , Plant A L , Chesler R , Kroll M , Ruddel M , Durst R A
Published: January 2020

We developed a method for quantitatively determining theophylline in serum, using a heterogeneous immunoassay called flow-injection immunoanalysis. The reaction involves competition between serum theophylline and theophylline-labeled liposomes. Separation occurs on a solid-phase reactor column containing immobilized antibody to theophylline incorporated in a flow-injection system. Subsequent lysis of the bound liposomes provides sensitive detection of the analyte. Effective regeneration of the immobilized antibody activity allows the reactor to be reused for hundreds of sequential samples. Comparison of the results of the flow-injection immunoassay method with results obtained with a commercially available fluorescence polarization method showed an excellent correlation.
Clinical and laboratory studies of a new immunoradiometric assay of parathyroid hormone-related protein
Authors: Fraser W D , Robinson J , Lawton R , Durham B , Gallacher S J , Boyle I T , Beastall G H , Logue F C
Published: January 2020

Parathyroid hormone-related protein (PTHrP) was measured in plasma by a new immunoradiometric assay (IRMA) from Nichols Institute. The assay is specific for PTHrP and shows excellent parallelism when measuring keratinocyte fluid, samples with high PTHrP content, and PTHrP-supplemented plasma. A precision profile established the assay detection limit at 0.7 pmol/L. PTHrP was unstable in plasma, but the degradation rate was patient-specific. Because delay in separation resulted in loss of PTHrP immunoreactivity, samples were collected into tubes containing protease inhibitors (aprotinin, leupeptin, pepstatin, and EDTA) and separated within 30 min. Among normal subjects, 78% had PTHrP values greater than the detection limit; the reference range established was < 0.7-2.6 pmol/L. Of patients with hypercalcemia associated with malignancy, 46% had PTHrP > 2.6 pmol/L. PTHrP was increased in patients with breast (73%), genitourinary (64%), or lung (46%) malignancy but was rarely above normal in patients with hematological (29%) or gastrointestinal (33%) malignancy. PTHrP and nephrogenous cyclic adenosine monophosphate (NcAMP) were strongly correlated (r = 0.63, P < 0.01) in 37 patients with PTHrP values greater than the detection limit, but 8 patients had PTHrP and parathyroid hormone [PTH(1-84)] values below the limit of detection with inappropriate or increased NcAMP. Five of these eight patients had small cell carcinoma of lung. These patients may have secreted a factor that is not detected by the IRMAs of PTHrP or PTH used in this study but that produces hypercalcemia by means of cAMP-mediated mechanisms.
Rapid enzymatic analysis for human immunodeficiency virus type 1 DNA in clinical specimens
Authors: Sevall J S , Prince H , Garratty G , O'Brien W A , Zack J A
Published: January 2020

A clinical procedure for rapid detection of human immunodeficiency virus type 1 (HIV-1) by DNA amplification is demonstrated. The rapid procedure reduces handling requirements and amplification time and eliminates use of radioactivity for the detection of the amplification product. Total leukocyte lysates are the amplification substrates. Two conserved regions in the HIV-1 genome are amplified by 45 cycles of a two-temperature thermal cycle and the amplification products are detected by ultraviolet light after electrophoresis on agarose gels. Twenty-four specimens clinically diagnosed by detection of antibody (IgG) to HIV-1 were confirmed by the rapid DNA amplification procedure. In a blind study, 56 samples positive for HIV-1 DNA were detected in 503 individuals by the current classical polymerase chain reaction method; the same 56 positive samples were also detected by the rapid amplification protocol. No false-positive or false-negative results were obtained. The turnaround time for analysis has been reduced to < 24 h without compromising test results.
Automated enzymatic determination of sodium in serum
Authors: Quiles R , Fernández-Romero J M , Fernández E , Luque de Castro M D , Valcárcel M
Published: January 2020

An automated method based on the principles of flow-injection analysis is proposed for the enzymatic determination of sodium ion in serum. The method relies on the activation of beta-galactosidase by the analyte. The features of the proposed method include linear range between 1 and 1700 mumol/L, a sampling rate of 50 samples/h, a sample volume of 50 microL, and the absence of interferences from species usually present in serum. The results obtained were consistent with those provided by widely used methods such as those based on flame spectrometry and direct potentiometry with ion-selective electrodes.
High-molecular-mass alkaline phosphatase as a tumor marker for colorectal cancer: comparison of two test methods
Authors: Wei J S , Chung N C , Wei L L , Tzeng W F , Liu T Z , Wang J Y
Published: January 2020

We measured high-molecular-mass alkaline phosphatase (HiMwALP) in serum samples from patients with colorectal cancer by polyacrylamide gel electrophoresis (PAGE) and by column chromatography on diethylaminoethyl (DEAE)-cellulose. Determination of carcinoembryonic antigen (CEA) by an enzyme immunoassay, a frequently used cancer assessment method, was used for comparison. We studied patients with primary colorectal cancer (n = 72), using others with hemorrhoids (n = 38) for a comparison group. HiMwALP activities twice those of pooled normal sera were adopted as cutoff values. The diagnostic sensitivity of the PAGE method for 72 colorectal cancer patients was 63% vs 36% for the DEAE method and 50% for the CEA method. The diagnostic specificities of the PAGE, DEAE, and CEA methods were 89%, 79%, and 95%, respectively. Using both HiMwALP (PAGE method) and CEA for the detection of primary colorectal cancer increased the sensitivity to 72% but decreased specificity to 87%.
Equivalent discrimination among states of thyroid function by immunochemiluminimetric and immunoradiometric determination of thyrotropin
Authors: Roden M , Nowotny P , Hollenstein U , Schneider B , Vierhapper H , Waldhäusl W
Published: January 2020

New immunochemiluminometric assays (ICMA) of thyrotropin (TSH) have been reported to facilitate the diagnosis of hyperthyroidism. To compare the accuracy of ICMA-TSH with that of a conventional immunoradiometric assay (IRMA-TSH), we examined 115 consecutive patients of a thyroid outpatient clinic. On the basis of complete thyroid-function testing, including thyroliberin tests, the untreated patients (n = 89) were subclinically hyperthyroid (n = 21), subclinically hypothyroid (n = 13), or euthyroid (n = 55). The receiver-operating characteristic (ROC) curve, used for comparing the TSH values obtained by these two methods, was shifted to the left for the IRMA-TSH. The area under the curve was greater for IRMA-TSH than for ICMA-TSH (0.984 vs 0.869, respectively), which suggests equal or better clinical performance of IRMA-TSH vs ICMA-TSH in discriminating between hyperthyroidism and euthyroidism. Both assays displayed similar clinical sensitivity/specificity for evaluating thyroid function in hypothyroid and treated patients. We conclude that the ICMA-TSH tested is not more accurate than an IRMA in distinguishing between hyperthyroidism and euthyroidism.
Monoclonal antibodies specific for prothrombin fragment 1.2 and their use in a quantitative enzyme-linked immunosorbent assay
Authors: Hursting M J , Butman B T , Steiner J P , Moore B M , Plank M C , Szewczyk K M , Bell M L , Dombrose F A
Published: January 2020

Prothrombin fragment 1.2 (F1.2) is an activation peptide generated during a critical event of blood coagulation, the conversion of prothrombin to thrombin. As a marker of thrombin generation, F1.2 has clinical potential in assessing thrombotic risk and monitoring anticoagulant therapy. In developing a highly specific, monoclonal antibody-based immunoassay of human plasma F1.2, we generated six murine anti-F1.2 monoclonal antibodies, using as immunogen a synthetic peptide (sequence: CGSD-RAIEGR) similar to the unique carboxyl terminus of F1.2. Each antibody bound F1.2 but not prothrombin. Epitope mapping studies with one antibody (5-3B) showed that optimum binding required six to eight amino acids plus a terminal arginine to emulate the F1.2 carboxyl terminus. A quantitative sandwich ELISA for human plasma F1.2 was configured with monoclonal antibody 5-3B as the capture antibody and peroxidase-labeled polyclonal antibodies to the F1.2 amino-terminal region as detector antibodies. Calibrators were prepared by adding purified F1.2, 0-10 nmol/L, to F1.2-depleted plasma. Assay characteristics included the following: mean (+/- SD) analytical recovery of 98% +/- 13%; no interference from lipemia, hemolysis, icterus, or thrombolytic agents; 0.08 nmol/L sensitivity; and mean intra- and interassay imprecision (three lots) < 12% at both low and high concentrations of F1.2.
Multicenter evaluation of Iso-ALP test kit for measurement of bone alkaline phosphatase activity in serum and plasma
Authors: Rosalki S B , Foo A Y , Burlina A , Prellwitz W , Stieber P , Neumeier D , Klein G , Poppe W A , Bodenmüller H
Published: January 2020

A test kit (Iso-ALP, Boehringer Mannheim) for measuring human bone alkaline phosphatase activity in serum or plasma was evaluated in five laboratories in three countries. The assay is based on the principle described by Rosalki and Foo (Clin Chem 1984;30:1182-6) and uses wheat germ lectin to precipitate bone alkaline phosphatase. Residual ALP in the supernate in comparison with total ALP is used to quantify the bone fraction. The imprecision of residual ALP measurement was low (median between-run CV 4.9%) and comparable with that of total ALP. Linearity of precipitation was demonstrable up to a bone ALP activity (diethanolamine buffer 37 degrees C) of 2000 U/L, though a matrix effect was observed for dilutions of high-activity sera in saline or bovine serum albumin. For assaying patients' samples, different batches of lectin demonstrated excellent comparability. Taking electrophoresis as a basis for standardization, we determined that the lectin precipitated approximately 90% of bone ALP, but < 5% of nonbone ALP. From this we derived serum/plasma upper reference limits for bone ALP activity in adults and children.
Clinical and laboratory studies of time-resolved fluorescence immunoassays of thyrotropin and free triiodothyronine
Authors: Taimela E , Aalto M , Koskinen P , Irjala K
Published: January 2020

We evaluated a new time-resolved immunofluorometric assay (TR-IFMA) for thyrotropin (TSH) and a time-resolved fluoroimmunoassay (TR-FIA) for free triiodothyronine (FT3) in serum. Delfia hTSH Ultra is a solid-phase two-site TR-IFMA involving three monoclonal antibodies directed against different antigenic sites on the human TSH molecule. It offers improved analytical (+3 SD of zero) and functional (10% between-assay CV) detection limits of 0.001 and 0.016 mlU/L, respectively. Results of the assay agreed closely with those of a second-generation immunoradiometric assay (TSH-IRMA) for samples with TSH > 0.2 mlU/L (n = 114). The nonparametric reference interval for TSH in euthyroid men and women was 0.5-4.7 mlU/L (n = 200). The TSH concentrations of hyperthyroid subjects (n = 29) were < 0.016 mlU/L (median; range < 0.016-0.063 mlU/L). The Delfia FT3 assay, a solid-phase TR-FIA, is based on the back-titration principle and second-antibody separation. The parametric reference interval for FT3 in euthyroid men and women was 4.2-7.6 pmol/L (n = 320). The FT3 concentration in hyperthyroidism (n = 29) was 23 +/- 22 pmol/L (mean +/- SD; range 7.9-85 pmol/L). These methods offered rapid results (2-4 h for the whole procedure) and were simple to run.
Effects of age, race, sex, and smoking on prothrombin fragment 1.2 in a healthy population
Authors: Hursting M J , Stead A G , Crout F V , Horvath B Z , Moore B M
Published: January 2020

Prothrombin fragment 1.2 (F1.2) is a biomarker of thrombin generation during blood coagulation and has diagnostic potential for assessing thrombotic risk and monitoring anticoagulation therapy. We used a monoclonal antibody-based immunoassay for plasma F1.2 to establish a well-defined reference interval and to evaluate the effects of age, race, sex, and smoking status on F1.2 concentrations in a healthy population. Plasma samples and demographic information were obtained from 357 healthy individuals. F1.2 concentrations more closely followed a lognormal than a gaussian frequency distribution. In a multiple linear-regression model in which the logarithms of F1.2 concentrations were regressed on age, race, sex, and smoking status, the significant explanatory variables were age and, to a lesser extent, sex and smoking. A segmented "hockey stick" regression model indicated that F1.2 concentrations and age were unrelated for individuals < 44 years old but were positively correlated above that age threshold. The estimated 95% tolerance interval (P = 0.95) for F1.2 in healthy individuals < 44 years old (n = 268) was 0.21-2.78 nmol/L. We conclude that age-matched F1.2 reference intervals may be important for studies evaluating the diagnostic utility of F1.2 measurements, and that the clinical relevance of increased thrombin generation during aging warrants further investigation.
Quantitative analysis of serum proteins separated by capillary electrophoresis
Authors: Kim J W , Park J H , Park J W , Doh H J , Heo G S , Lee K J
Published: January 2020

The possibility of open tubular capillary electrophoresis for clinical diagnostic use is examined. Capillary electrophoresis was performed in an untreated 50 microns (i.d.) x 100 cm (65 cm to detector) capillary with detection of absorbance at 200 nm. Conditions for the separation of serum proteins without adsorption to the capillary surface were established. Quantitative analyses of serum samples from 38 patients with liver cirrhosis, nephrotic syndrome, or polyclonal gammopathy by capillary electrophoresis were done and the results were compared with those by conventional agarose gel electrophoresis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. All samples were analyzed in duplicate. We evaluated linearity of response, within-run CV, and the correlation between capillary electrophoresis and agarose gel electrophoresis.
Rapid cycle allele-specific amplification: studies with the cystic fibrosis delta F508 locus
Authors: Wittwer C T , Marshall B C , Reed G H , Cherry J L
Published: January 2020

Rapid cycle DNA amplification is a polymerase chain reaction technique with improved product specificity and cycle times of 20-60 s, allowing complete 30-cycle reactions in 10-30 min. The presence or absence of the delta F508 deletion and wild-type allele was determined in 104 cystic fibrosis patients by rapid cycle DNA amplification. In separate allele-specific assays, sequences on both sides of the delta F508 locus were amplified with the 3' end of a discriminating primer at the delta F508 locus, with either a 3-bp or a 1-bp mismatch. With rapid cycling (35-s cycles), single-base discrimination was achieved over a broad range of annealing temperatures (50 degrees C or lower); with conventional cycling and "hot starts" (160-s cycles), only annealing temperatures of 61-62 degrees C sufficiently discriminated between alleles. With rapid cycling, genotype could still be assessed with annealing temperatures as low as 25 degrees C. We conclude that faster temperature cycling can improve the results of allele-specific amplification.
Pyrophosphatase-based enzyme-linked immunosorbent assay of total IgE in serum
Authors: Peuravuori H , Korpela T
Published: January 2020

Human IgE was purified to near homogeneity by a two-step procedure consisting of immunoaffinity chromatography and high-performance liquid chromatography. Mouse hybridoma cell lines secreting antibodies against IgE were generated. Monoclonal and polyclonal antibodies were used to develop a "sandwich"-type ELISA for determining total IgE in human serum. Inorganic pyrophosphatase (EC 3.6.1.1), an enzyme having a high turnover number, was used as the label. The mean analytical recovery of our ELISA was 95.2% and the results showed good linear correlation with an established RIA of IgE. We found pyrophosphatase to be a good alternative label for use in immunoassays.
Assay of complexed alpha 1-antichymotrypsin in plasma
Authors: Heidtmann H H , Havemann K
Published: January 2020

We introduce an assay to measure complexed alpha 1-antichymotrypsin in human plasma. The assay works on the principle that the target proteinase cathepsin G has a very high affinity to the surface of microtiter plates, even if these surfaces are blocked with albumin and Tween 20. alpha 1-Antichymotrypsin, when complexed to cathepsin G, is thus immobilized and can be detected with specific antibodies. The mean (SD) concentration of alpha 1-antichymotrypsin in 50 healthy individuals was 1.73 (0.58) nmol/L. The detection limit was 0.84 nmol/L. The results of the assay are linear to at least 14 nmol/L. We propose to use this assay for diseases in which increased turnover of alpha 1-antichymotrypsin or cathepsin G can be expected to clarify further the function of this enzyme-inhibitor system.
Measurement of urinary sugars by HPLC in the estimation of intestinal permeability: evaluation in pediatric clinical practice
Authors: Willems D , Cadranel S , Jacobs W
Published: January 2020

Determinations of the permeability of the intestine to various sugars have been used to assess intestinal mucosal abnormalities, but the widespread application of such tests has been prohibited by the complexity of most classically used techniques. In this study, we evaluated the sensitivity and practicability of using HPLC to estimate intestinal permeability, with mannitol and lactulose as mono- and disaccharides, respectively. The results of the permeability tests were compared with those of intestinal biopsy in 20 children. All patients with an abnormal intestinal biopsy showed a low mannitol recovery, and the patients with allergic symptoms showed a high lactulose recovery. Our preliminary data suggest that HPLC is reliable, gives results comparable with those of other methods, and is a practical way to determine intestinal permeability noninvasively.
Enzymatic determination of anti-NAD+ glycohydrolase in serum
Authors: Gella F J , Viver J , Canalias F , Beleta J , Gonzalez-Sastre F
Published: January 2020

A colorimetric method for measuring anti-NAD+ glycohydrolase in human sera has been developed. The assay involves the inhibition of NAD+ glycohydrolase (EC 3.2.2.5) by the antibody and determination of the noninhibited enzyme activity by using an enzymatic amplifying system for NAD+. The assay is easily carried out and has the additional advantage of a direct relationship between signal and antibody concentration. The results obtained for 100 human sera compare favorably with other tests commonly used to obtain evidence of streptococcal infections or their complications, such as the anti-streptolysin O and the anti-DNase B tests.
Effect of double-blind crossover selenium supplementation on biological indices of selenium status in cystic fibrosis patients
Authors: Portal B , Richard M J , Ducros V , Aguilaniu B , Brunel F , Faure H , Gout J P , Bost M , Favier A
Published: January 2020

Twenty-seven cystic fibrosis patients received selenium supplementation (2.8 micrograms of sodium selenite per kilogram of body weight per day) or a placebo. This 5-month trial was conducted as a double-blind, placebo-controlled study. After an interval of 2 months, treatments of the two groups were interchanged (crossed over) for another 5-month period. A group of healthy subjects, living in the same area, was investigated simultaneously. No selenium deficiency was found either in plasma or in erythrocytes before the supplementation. This result was inconsistent with a previous study performed in 1988 in our laboratory. This change in selenium status can be explained by progress in the nutritional nursing care of children and by the addition of selenium to the diet. During the study, selenium concentrations in plasma decreased when patients received placebo treatment and increased during selenium intake. In one of the two groups a similar variation was found for glutathione peroxidase activities in plasma and erythrocytes, whereas erythrocyte selenium was normal and did not change in any group. Nowadays, in the Grenoble area, the selenium status of cystic fibrosis patients is close to normal. Nevertheless, this study indicates a fragile equilibrium, given that selenium concentrations cn be lowered by placebo or mildly increased by supplementation.
Improvement and assessment of enzyme-linked immunosorbent assay to detect low FK506 concentrations in plasma or whole blood within 6 hours
Authors: Wallemacq P E , Firdaous I , Hassoun A
Published: January 2020

FK506, a promising new immunosuppressant, is currently under clinical investigation. Because dose-dependent toxicity is possible, blood concentrations of FK506 should be monitored. We improved the original ELISA of FK506 by shortening the incubation time. With some modification of materials, results are obtained within 6 h instead of 2 days, with similar or even better precision. Internal and external quality-control programs showed that our results correlated satisfactorily both with values determined by the original method and the theoretical expected values. Either plasma (detection limit 0.1 microgram/L) or whole-blood (detection limit 1 microgram/L) samples can be used. The sensitivity of the method makes it particularly useful for accurate pharmacokinetic studies or measurement of low blood concentrations. Twenty-four drugs and nine biological variables showed no significant interference on the assay. Study of the concentration- and temperature-dependent distribution of FK506 shows that the drug is largely bound to erythrocytes (ratio of blood to plasma concentrations is 10-40); as the erythrocytes become saturated, more of the drug is found in the plasma. Plasma concentrations may vary according to the blood temperature. We conclude that whole blood should be used for FK506 monitoring, as it is for monitoring cyclosporine.
The freckle plot (daily turnaround time chart): a technique for timely and effective quality improvement of test turnaround times
Authors: Pellar T G , Ward P J , Tuckerman J F , Henderson A R
Published: January 2020

Test turnaround times are often monitored on a monthly basis. However, such an interval usually means that not all causes for delay in test reporting can be unequivocally identified for institution of remedial action. We have devised a daily chart--the freckle plot--that graphically displays the test turnaround times by laboratory receipt time. Different symbols are used to designate specimens reported within the test's turnaround time limit, those within 10 min beyond that limit, and those well outside the limit. These categories are adjustable to suit different limits of stringency. Freckle plots are produced on a daily basis and can be used to track down causes for test delays. Using the 1-h turnaround time "stat" potassium test as a model, we found 16 causes for test delay, of which 9 were potentially remediable. By applying these remedies, we were able to increase test compliance, in the day shift, from 91.5% (95% confidence interval 88.8%-93.7%) to 97.6% (95% confidence interval 96.4-98.55%), which is significant at P < 10(-7). This daily plot is a useful quality assurance tool, supplementing the more conventional tests used to ensure laboratory quality improvement.
Stability of intact chorionic gonadotropin (hCG) in serum, liquid whole blood, and dried whole-blood filter-paper spots: impact on screening for Down syndrome by measurement of free beta-hCG subunit
Authors: Spencer K , Macri J N , Carpenter P , Anderson R , Krantz D A
Published: January 2020

The use of multiple maternal serum biochemical markers in screening for Down syndrome is gaining worldwide acceptance. We sought to study the impact of the potential instability of intact human chorionic gonadotropin (hCG) on free beta-hCG subunit, a marker that has recently been used successfully in such screening. We found that, in practice, any changes in free beta-hCG due to the instability of intact hCG do not inhibit the effectiveness of free beta-hCG as a marker for Down syndrome. This was proven by controlled laboratory experiments at various stress temperatures, freeze-thaw studies, and analysis of a large set of screening data with particular reference to time in transit for individual samples. Data from controlled dissociation studies demonstrate that any apparent increase in free beta-hCG due to the instability of intact hCG cannot be attributed simply to the dissociation of intact hCG. Finally, for large-scale mass population screening in areas of the world where transport delays, safety concerns, and high temperatures preclude the shipment of liquid whole blood, dried whole-blood spots in filter paper provide a suitable delivery system with many advantages.
Comparison of hypertonic and isotonic reference electrode junctions for measuring ionized calcium in whole blood: a clinical study
Authors: Masters P W , Payne R B
Published: January 2020

We measured ionized calcium concentrations in whole blood from 91 patients who had no clinical or biochemical evidence of disturbed calcium homeostasis and who had a wide range of serum albumin concentrations. We used both a standard Ciba-Corning 634 analyzer, which has a membrane-restricted saturated KCl reference electrode bridge, and a modified instrument with a 150 mmol/L NaCl bridge. After adjusting the externally standardized values from each instrument for their least-squares regressions on pH, there was a significant correlation between ionized calcium and albumin only with the standard analyzer. In contrast, only values from the modified instrument correlated with serum chloride; this was not explained by ionic strength or organic anion interferences. We conclude that there is unlikely to be any major advantage in using a membrane-restricted isotonic NaCl reference electrode for in vitro clinical measurements, although it may be of value for in vivo monitoring. The importance of measuring serum albumin when using most commercial ionized calcium analyzers is emphasized.
Simultaneous determination of free testosterone and testosterone bound to non-sex-hormone-binding globulin by equilibrium dialysis
Authors: Barini A , Liberale I , Menini E
Published: January 2020

We describe a procedure based on equilibrium dialysis that allows the simultaneous determination of free testosterone and testosterone bound to non-sex-hormone-binding globulin (non-SHBG) in plasma. After saturating SHBG with 5 alpha-dihydrotestosterone (DHT) according to a technique recently described, the percentage of free testosterone in the treated and the untreated samples is measured by equilibrium dialysis with use of a semiautomated instrument that allows rigorous standardization of the experimental conditions. The present method is simpler and faster than the previously described technique in which, after the saturation of SHBG with DHT, the unbound fractions were measured by centrifugal ultrafiltration dialysis. The method is also reproducible and suited for the analysis of a large number of samples. The technique has been applied to the determination of the fractional distribution of testosterone in plasma pools from normally menstruating, pregnant, and postmenopausal women and from normal men.
Whole-blood cyclosporin G in renal transplant recipients determined by two immunoassays and liquid chromatography
Authors: McBride J H , Kim S S , Danovitch G M , Rodgerson D O , Reyes A F , Ota M K
Published: January 2020

Cyclosporin G (CsG) is less nephrotoxic than cyclosporin A (CsA) and is undergoing clinical trials for use as an immunosuppressive agent after renal transplantation. Three assays for whole-blood CsA-HPLC, RIA (INCSTAR, Cyclo-Trac SP), and FPIA (Abbott TDx)--were adapted for use with CsG and were assessed for analytical suitability and to determine which assay was capable of deriving CsG values rapidly after transplantation. The assays were acceptable in terms of sensitivity, linearity, analytical recovery, and precision. When considering blood samples (n = 107) from renal transplant recipients receiving a low dose of CsG (5 mg/kg per day) and a high dose (10 mg/kg per day), we obtained the following correlation data: RIA = 0.974HPLC + 27.89 (r = 0.9798, Sy/x = 39.24); FPIA = 0.964HPLC + 33.59 (r = 0.9819, Sy/x = 36.66); and FPIA = 0.977RIA + 9.50 (r = 0.9894, Sy/x = 28.12). The FPIA of CsG is recommended as the most rapid method, although it is the most expensive. HPLC, RIA, and FPIA were capable of accurately deriving projected CsG concentrations at various stages of the clinical trial when the low- and high-dose regimes were tapered over a period of 16 weeks.
Electrochemical homogeneous enzyme immunoassay of theophylline in hemolyzed, icteric, and lipemic samples
Authors: Yao H , Jenkins S H , Pesce A J , Halsall H B , Heineman W R
Published: January 2020

We demonstrate here an electrochemical homogeneous enzyme immunoassay for theophylline, which can be performed in hemolyzed, lipemic, and icteric samples. The assay used an unmodified Syva EMIT theophylline kit. One of the enzymatic reaction products, NADH, reacted with 2,6-dichloroindophenol (DCIP) to reduce DCIP to DCIPH2, which was detected electrochemically with flow-injection analysis. The inter- and intraassay coefficients of variation of this manual technique were < 9% at theophylline concentrations of 14 to 34 mg/L. The CVs were 9-15% at low concentrations (6.3 mg/L), which is below the therapeutic range. Analytical recoveries were 91-97% for normal serum and 92-111% for hemolyzed, icteric, or lipemic sera. The measured concentrations (y) were compared with those obtained by the fluorescence polarization immunoassay (x); a scatter plot of the results showed a linear relationship of y = 1.00 x - 0.57 mg/L (r = 0.966, Sy/x = 1.51). This alternative way to measure the serum concentration of theophylline overcomes the shortcomings of spectrophotometric methods, by which it is difficult to measure theophylline in severely hemolyzed, icteric, or lipemic sera.
Time-resolved immunofluorometry and other frequently used immunoassay types for follicle-stimulating hormone compared by using identical monoclonal antibodies
Authors: Madersbacher S , Shu-Chen T , Schwarz S , Dirnhofer S , Wick G , Berger P
Published: January 2020

The influence of assay design and quantification system on assay performance was investigated by developing, optimizing, and comparing a time-resolved immunofluorometric assay (IFMA), an immunoenzymometric assay (IEMA), an immunoradiometric assay (IRMA), and a competitive radioimmunoassay (RIA), all performed with the same monoclonal antibodies (MCA) directed against human follicle-stimulating hormone (hFSH). The lowest detection limit (2 ng/L for hFSH-I-3, corresponding to 2.5 mIU of 1st International Reference Preparation of hFSH 78/549 per liter), the widest measuring range (2-160,000 ng/L), and the greatest signal-to-noise ratio (13,000:1 at 160,000 ng/L) were obtained in the IFMA. For analysis of serum samples from 101 male (ages 2-91 years) and 99 female (ages 2-90 years) individuals at a single dilution, 100% of samples were within the measuring range of the IFMA, whereas only 87%, 55%, 32%, and 8% of the sera were for the IRMA, the IEMA evaluated with double-wavelength measurement, the conventional IEMA, and the competitive MCA-based RIA, respectively. These studies demonstrate clear advantages of the IFMA in sensitivity and assay range, which allows reliable and cost- and time-effective determination of hFSH in individuals from infancy to senescence.
Stable liposomes for assays of human sera
Authors: Ishimori Y , Rokugawa K
Published: January 2020

We report a novel homogeneous immunoassay system involving protein-bearing liposome-encapsulated carboxyfluorescein as a release marker. We applied this system to determine protein antigens, e.g., ferritin, in human serum samples by a sandwich-type assay. Liposomal lysis was observed in many samples, even though no second antibody was added to the reaction mixture. We demonstrated that the functional groups used to immobilize an antibody on liposomes are related to this phenomenon. Stable liposomes in human sera were prepared by incorporating bromoacetyl groups instead of the dithiopyridyl groups used previously. A good correlation (y = 0.98x - 8.81, r = 0.98, Sx/y = 66.9, range approximately 10-2000 micrograms/L) with results by RIA was obtained in the ferritin measurement of 53 patients' sera by using these liposomes.
Tolerance limits for short-term analytical bias and analytical imprecision derived from clinical assay specificity
Authors: Klee G G
Published: January 2020

I propose a method for defining tolerance limits for assay bias and assay imprecision, based on the effects of these tolerance limits on the clinical specificity of the assay. An analytical "error budget" is defined as the squared sums of the imprecision and bias errors. The maximum limit for this error budget is set at a value corresponding to a 50% increase in the false-positive rate for classifying healthy subjects. For gaussian distributions with +/- 2 SD used as decision limits, this error budget equates to 0.45 SD of combined within-person and between-person biological variation (SDBiol). To provide reasonable power for bias detection in an assay, I recommend that the SD of the assay be kept at less than half the bias limit. Then, for the gaussian distribution, the maximum bias limit should be < 0.36 SDBiol and the SD of the assay should be < 0.18 SDBiol. Procedures are provided for using the same principles to define tolerance limits for decision limits other than +/- 2 SD and for nongaussian distributions.