bluuebunny/crossref_metadata_2025 huggingface dataset:
Histologic and Laboratory Characteristics of Symptomatic and Asymptomatic Castleman Disease in the Pediatric Population
Authors: Chisholm Karen M , Fleming Mark D
Published: January 2020

Objectives

Compare the morphologic, laboratory, and clinical features of asymptomatic and symptomatic Castleman disease in the pediatric population.

Methods

We reviewed clinical records and histopathology of patients with Castleman disease from 2 pediatric institutions.

Results

Of 39 patients with pediatric Castleman disease, 37 had unicentric disease, all classified with the hyaline vascular variant of Castleman disease, 8 of which were clinically symptomatic. These 8 patients demonstrated abnormal laboratory findings, including microcytic anemia, elevated erythrocyte sedimentation rate and C-reactive protein, and hypoalbuminemia. In addition, histopathologic evaluation showed that the 8 symptomatic cases had more hyperplastic germinal centers, fewer atrophic or regressed germinal centers, fewer mantle zones containing multiple germinal centers, reduced “onion skinning” of mantle zones, and fewer “lollipop” formations compared with the asymptomatic cases.

Conclusions

This series of pediatric Castleman disease showed that lymph nodes from asymptomatic patients generally demonstrated the more classic hyaline vascular histology, whereas those with symptoms could lack or have only focal classic findings. As such, reactive lymph nodes with subtle Castleman-like features should prompt clinical correlation to ensure proper diagnosis.
An Innovative, Prospective, Hybrid Cohort-Cluster Study Design to Characterize Dengue Virus Transmission in Multigenerational Households in Kamphaeng Phet, Thailand
Authors: Anderson Kathryn B , Buddhari Darunee , Srikiatkhachorn Anon , Gromowski Gregory D , Iamsirithaworn Sopon , Weg Alden L , Ellison Damon W , Macareo Louis , Cummings Derek A T , Yoon In-Kyu , Nisalak Ananda , Ponlawat Alongkot , Thomas Stephen J , Fernandez Stefan , Jarman Richard G , Rothman Alan L , Endy Timothy P
Published: January 2020

Difficulties inherent in the identification of immune correlates of protection or severe disease have challenged the development and evaluation of dengue vaccines. There persist substantial gaps in knowledge about the complex effects of age and sequential dengue virus (DENV) exposures on these correlations. To address these gaps, we were conducting a novel family-based cohort-cluster study for DENV transmission in Kamphaeng Phet, Thailand. The study began in 2015 and is funded until at least 2023. As of May 2019, 2,870 individuals in 485 families were actively enrolled. The families comprise at least 1 child born into the study as a newborn, 1 other child, a parent, and a grandparent. The median age of enrolled participants is 21 years (range 0–93 years). Active surveillance is performed to detect acute dengue illnesses, and annual blood testing identifies subclinical seroconversions. Extended follow-up of this cohort will detect sequential infections and correlate antibody kinetics and sequence of infections with disease outcomes. The central goal of this prospective study is to characterize how different DENV exposure histories within multigenerational family units, from DENV-naive infants to grandparents with multiple prior DENV exposures, affect transmission, disease, and protection at the level of the individual, household, and community.
Characteristics of Invasive Pneumococcal Disease Caused by Emerging Serotypes After the Introduction of the 13-Valent Pneumococcal Conjugate Vaccine in England: A Prospective Observational Cohort Study, 2014–2018
Authors: Amin-Chowdhury Zahin , Collins Sarah , Sheppard Carmen , Litt David , Fry Norman K , Andrews Nick , Ladhani Shamez N
Published: January 2020

Background

England is experiencing a rapid increase in invasive pneumococcal disease (IPD) caused by serotypes 8, 12F, and 9N; their clinical characteristics and outcomes have not been described.

Methods

Public Health England conducts national IPD surveillance. Cases due to emerging serotypes were compared with those included in the 13-valent pneumococcal conjugate vaccine (PCV13) and the remaining non-PCV13 serotypes.

Results

There were 21 592 IPD cases during 2014–15 to 2017–18, including 20 108 (93.1%) with serotyped isolates and 17 450 (86.8%) with completed questionnaires. PCV13 serotypes were responsible for 20.1% (n = 4033), while serotype 8 (3881/20 108 [19.3%]), 12F (2365/20 108 [11.8%]), and 9N (1 296/20 108 [6.4%]) were together responsible for 37.5% of cases. Invasive pneumonia was the most common presentation (11 424/16 346 [69.9%]) and, overall, 67.0% (n = 11 033) had an underlying comorbidity. The median age (interquartile range) at IPD due to serotypes 8 (59 [45–72] years) and 12F (56 [41–70] years) was lower than serotype 9N (67 [53–80] years), PCV13 serotypes (68 [52–81] years), and remaining non-PCV13 serotypes (70 [53–82] years). Serotype 9N IPD cases also had higher comorbidity prevalence (748/1087 [68.8%]) compared to serotype 8 (1901/3228 [58.9%]) or 12F (1042/1994 [52.3%]), and higher case fatality (212/1128 [18.8%]) compared to 8.6% (291/3365) or 10.0% (209/2086), respectively.

Conclusions

Serotypes 8 and 12F were more likely to cause IPD in younger, healthier individuals and less likely to be fatal, while serotype 9N affected older adults with comorbidities and had higher case fatality.
Naturally Acquired Protection Against Upper Respiratory Symptoms Involving Group A Streptococcus in a Longitudinal Cohort Study
Authors: Lewnard Joseph A , Whittles Lilith K , Rick Anne-Marie , Martin Judith M
Published: January 2020

Background

Pharyngitis due to group A Streptococcus (GAS) represents a major cause of outpatient visits and antibiotic use in the United States. A leading vaccine candidate targets 30 of the > 200 emm types of GAS. We aimed to assess natural protection conferred by GAS against respiratory symptoms.

Methods

In a 5-year study among school-aged children in Pittsburgh, Pennsylvania, pharyngeal cultures were obtained from children at 2-week intervals, and active surveillance was conducted for respiratory illnesses. We assessed protection via the relative odds of previous detection of homologous strains (defined by field-inversion gel electrophoresis banding pattern), emm types, and emm clusters at visits where GAS was detected with symptoms, vs visits where GAS was detected without symptoms. We used a cluster bootstrap of children to adjust estimates for repeated sampling.

Results

At visits where previously detected GAS emm types were identified, we estimated 81.8% (95% confidence interval [CI], 67.1%–91.7%) protection against typical pharyngitis symptoms among children reacquiring the same strain, and 94.5% (95% CI, 83.5%–98.6%) protection among children acquiring a distinct strain. We estimated 77.1% (95% CI, 33.7%–96.3%) protection against typical symptoms among children acquiring partially heterologous emm types belonging to a previously detected emm cluster. Protection was evident after both symptomatic and asymptomatic detections of GAS. We did not identify strong evidence of protection against atypical respiratory symptoms.

Conclusions

Within a 5-year longitudinal study, previous detection of GAS emm types was associated with protection against typical symptoms when homologous strains were subsequently detected. Naturally acquired protection against partially heterologous types suggests that emm type–based vaccines may have broader strain coverage than what has been previously assumed.
Simultaneous determination of uric acid and creatinine in plasma by reversed-phase liquid chromatography.
Authors: Zhiri A , Houot O , Wellman-Bednawska M , Siest G
Published: January 2020

Because numerous substances, endogenous compounds as well as xenobiotics, interfere with determination of uric acid and creatinine, we have devised a more nearly specific method by which we can simultaneously determine uric acid and creatinine in plasma by "high-performance" liquid chromatography. We used a mobile phase of ammonium acetate (30 mmol/L) and methanol (156 mmol/L) at pH 7.0 and a flow rate of 1 mL/min. We used a C18 reversed-phase column, and measured absorbance at 235 nm, the wavelength corresponding to the absorption maximum of uric acid and creatinine. Uric acid and creatinine could be determined in less than 5 min by directly injecting plasma diluted fivefold; use of a precolumn eliminated the need for deproteinization. We evaluated the precision, recovery, linearity, specificity, and limit of detection of the method and we checked for analytical interference by 37 currently used drugs. We compared results with those obtained by routine and reference methods.
Enzyme-antigen immunoassay for human placental alkaline phosphatase in serum and tissue extracts, and its application as a tumor marker.
Authors: Pollet D E , Nouwen E J , Schelstraete J B , Renard J , Van de Voorde A , De Broe M E
Published: January 2020

In this enzyme-antigen immunoassay for human placental alkaline phosphatase (hPLAP; EC 3.1.3.1.) in serum and tissue extracts, polyclonal rabbit antiserum to mouse IgG2b is adsorbed to the wells of a microtiter plate, its excess binding sites are blocked, then it is incubated with murine monoclonal anti-hPLAP and mixed with serially diluted standard or sample antigen. The amount of antigen bound is determined by measuring its enzymic activity. The standard curve is linear for hPLAP concentrations of 0.2 to 1 U/L. The mean within-assay CV was 3.8% (SD 0.9%) for a serum sample and 6.1% (SD 3.0%) for a tissue extract. The respective mean between-assay CVs were: 6.7% (SD 2.0%), and 7.0% (SD 2.0%). Serum hPLAP concentrations, determined in four different dilutions, had a CV of 5.5%. We evaluated the method by standard additions and by comparing dilution curves for purified hPLAP, hPLAP in serum, and hPLAP in tissue extracts. The upper limit of activity in normal subjects was 0.1 U/L for serum samples, and 1.0 mU/g wet weight of tissue for tissue extracts. hPLAP activity was increased in 9.8% of all cancer patients, and in 40% of ovarian cancer patients. Almost half of the tumor biopsies were positive for hPLAP activity, and 94% of the biopsies from ovarian neoplasia had an increased activity of this isoenzyme. Of the nonmalignant tissues examined, normal lung tissue had the highest hPLAP activity.
Highly sensitive solid-phase immunoenzymometric assay for placental and placental-like alkaline phosphatases with a monoclonal antibody and monodisperse polymer particles.
Authors: Millán J L , Nustad K , Nørgaard-Pedersen B
Published: January 2020

We have used a mouse monoclonal antibody (H7) to placental alkaline phosphatase (PLAP, EC 3.1.3.1) in developing an immunoenzymometric assay for PLAP and PLAP-like enzymes. The antibody is bound to sheep anti-mouse IgG (Ab2) covalently coupled to tosylated shell-and-core light (1.07 g/cm3) monodisperse polymer particles. Adding the H7-Ab2-polymer particle suspension to a PLAP-containing sample gives maximal binding of the antigen within 10 min. PLAP and PLAP-like enzymes remain active and bound to the solid-phase throughout all assay manipulations, and can thus be saved for future testing. In testing the enzymes for inhibition by L-Phe, L-Phe-Gly-Gly, L-Leu, and L-homoarginine, the effect of all the inhibitors is fully reversible. The assay is highly versatile, and its sensitivity (routinely 0.05 micrograms/L) can be increased 1000-fold by adjusting the sample volume and incubation time (sample volume is irrelevant between 50 microL and 5 mL). We have measured the basal activities of PLAP in men and women and, by using enzyme inhibitors, have characterized it as corresponding to the PLAP-like phenotypes described in normal human testis.
Zinc content of cellular components of blood: methods for cell separation and analysis evaluated.
Authors: Milne D B , Ralston N V , Wallwork J C
Published: January 2020

Platelets, mononucleated cells, polymorphonucleated cells, and erythrocytes were separated from whole blood by use of discontinuous gradients of colloidal polyvinylpyrrolidone-coated silica ("Percoll"). We measured the zinc content of these cells by flame atomic absorption spectrophotometry, using a modified technique for micro-samples that obviated matrix interferences. Thus, results obtained by conventional flame atomic absorption and by the micro-method were identical. Inter-comparisons of separation methods indicated that separation of platelets and mononucleated cells by a two-gradient system of "Ficoll-Hypaque" (a synthetic polymer of sucrose) or Percoll was relatively poor, whereas there was a good separation when a tertiary gradient system of Percoll was used. The apparent zinc content of mononucleated cells depended on the degree of separation from the platelets, with contamination by platelets resulting in artificially high values for mononucleated cells.
Preparation and certification of human hair powder reference material.
Authors: Okamoto K , Morita M , Quan H , Uehiro T , Fuwa K
Published: January 2020

Human hair powder reference material was prepared and certified for elemental composition. Human scalp hair (from Japanese men) was washed in a non-ionic detergent solution in an ultrasonic cleaner, dried, ground in an agate ball mill, sieved through a polyethylene net, blended, bottled (1100 vials, 2 g each), and finally sterilized by 60Co radiation. The prepared material satisfied the homogeneity criteria for a reference material. We determined trace elements by inductively coupled plasma atomic emission spectrometry and atomic absorption spectrometry and investigated matrix interference effects. The material was certified by using the data obtained by various analytical techniques; certified values are provided for Ca, Cd, Cr, Cu, Fe, Hg, K, Mg, Mn, Na, Ni, Sr, and Zn. The elemental composition of this reference material is considered similar to normal values for the male Japanese population.
Immunochemical methods for measurement of transferrin in serum: effects of analytical errors and inappropriate reference intervals on diagnostic utility.
Authors: Bandi Z L , Schoen I , Bee D E
Published: January 2020

We estimated analytical errors of the Calbiochem, Kallestad, Hyland, Meloy, Helena, and Beckman immunochemical methods for serum transferrin. Intermethod biases were determined by analysis of the "Reference Preparation for Serum Proteins" of the College of American Pathologists and by analysis of 106 patients' serum samples. We judged the acceptability of errors by comparing confidence limits for total errors with 1/4 of the normal reference intervals. The transferrin status of each patient's sample was interpreted by comparing the result of each method with the normal reference interval claimed by the corresponding manufacturer. We found that the combined effects of medically unacceptable analytical errors and inappropriate normal intervals caused results of the tested methods not to be interchangeable. The Calbiochem method identified 61 serum samples (57%) as having abnormally high transferrin concentrations. In contrast, for the same specimen, with the Meloy method we found abnormally high transferrin concentrations for only two samples (1.8%).
Solid-phase enzyme-receptor assay (SPERA): a competitive-binding assay for glycopeptide antibiotics of the vancomycin class.
Authors: Corti A , Rurali C , Borghi A , Cassani G
Published: January 2020

A solid-phase enzyme-receptor assay (SPERA) has been developed for glycopeptide antibiotics of the vancomycin class such as teicoplanin, vancomycin, ristocetin, avoparcin, actaplanin, A-47934, A-41030, and A-35512-B. The assay exploits the mechanism of most action of these antibiotics, which is based on their interaction with acyl-D-alanyl-D-alanine, a constituent of the walls of most growing bacterial cells. The antibiotics and enzyme-labeled teicoplanin compete for a synthetic analog of the biological receptor, albumin-epsilon-aminocaproyl-D-alanyl-D-alanine. The various antibiotics produced different competition curves, 50% displacement being obtained with antibiotic concentrations ranging from 0.04 to 4 mg/L, vancomycin and actaplanin being the weakest and strongest competitors, respectively. For teicoplanin in human serum the intra-assay CV was 7.2%, the interassay CV was 11.2%, and the analytical recovery 94%. Teicoplanin concentrations obtained by SPERA (chi) correlated well with those obtained by microbiological assay (y): y = 1.03 chi + 0.053 (r = 0.943; n = 60). We conclude that SPERA is a powerful tool for identification and quantitative detection of glycopeptide antibiotics, even in complex media.
Transcutaneous carbon dioxide and oxygen tension measured at different temperatures in healthy adults.
Authors: Wimberley P D , Grønlund Pedersen K , Olsson J , Siggaard-Andersen O
Published: January 2020

Transcutaneous carbon dioxide tension (tc-pco2) at 37, 39, 41, 43, and 45 degrees C, and transcutaneous oxygen tension (tc-po2) at 41, 43, and 45 degrees C were measured simultaneously in 10 healthy adults during hyperventilation and inhalation of O2/CO2 gas. Nine electrodes were applied to each subject: Five CO2 electrodes, one O2 electrode, and three combined O2/CO2 electrodes. The CO2 electrodes had negligible temperature coefficients in the calibration gases, but the O2 electrodes showed an increase in po2 of 4.5% per degree C. With skin application, tc-pco2 increased approximately 4% per degrees C between 37 and 45 degrees C, which is close to the anaerobic temperature coefficient of pco2 in blood. The tc-po2 increases on the skin with increasing temperature appeared to be more dependent on changes in blood flow in skin, but in the temperature range 43 to 45 degrees C, tc-po2 showed the expected decrease in the temperature coefficient with increasing po2. The correlation between transcutaneous and capillary pco2 was close at all transcutaneous electrode temperatures, even 37 degrees C, provided the skin was preheated (via the electrode) to 45 degrees C. For tc-po2, an electrode temperature of at least 43 degrees C was necessary to produce a reasonable correlation between tc-po2 and capillary po2. The combined O2/CO2 electrodes measured slightly higher pco2 values than the single CO2 electrodes, but there were no differences in po2 readings, stabilization time, imprecision, or electrode drift between the two electrode types. The imprecision (CV, %) of tc-pco2 and tc-po2 measurements was approximately twice that of the corresponding capillary blood-gas measurements.
Changes in the ratio of lactate dehydrogenase isoenzymes 1 and 2 during the first day after acute myocardial infarction.
Authors: Jablonsky G , Leung F Y , Henderson A R
Published: January 2020

It is known that the ratio of isoenzyme 1 to total lactate dehydrogenase (LD, EC 1.1.1.27) in serum is increased in all patients with acute myocardial infarction within 24 h of the infarct. We now show that the LD-1/LD-2 ratio for serum more promptly indicates acute myocardial infarction, being for most patients equivalent to measurement of creatine kinase (EC 2.7.3.2) isoenzyme 2 (CK-2, CK-MB) in serum. Of 128 patients with a confirmed diagnosis of myocardial infarction, 66 had normal values for all "cardiac" enzymes at the time of admission, but greater than 75% of them showed a parallel increase in values for CK-2 and the LD-1/LD-2 ratio. Of the 26 patients who had one or more abnormal values for cardiac enzymes on admission, 95% showed a parallel increase in CK-2 and the LD-1/LD-2 ratio, the median time for the beginning of these changes being 9 h from the onset of chest pain. The remaining 36 patients were excluded from the study because CK-2 decreased after admission or because the time of onset of chest pain was uncertain.
Lack of specificity of current anti-digoxin antibodies, and preparation of a new, specific polyclonal antibody that recognizes the carbohydrate moiety of digoxin.
Authors: Thong B , Soldin S J , Lingwood C A
Published: January 2020

Current immunoassays for digoxin do not distinguish digoxin from its glycosidic metabolites. We have synthesized a novel digoxin/bovine serum albumin conjugate via reductive ozonolysis of the lactone ring such that the carbohydrate moiety of digoxin remains intact. Antibodies raised against this conjugate show minimal cross reactivity to digoxigenin, bisdigitoxide, monodigitoxide, digoxigenin, and digitoxin. With this antibody, digoxin can be measured in the presence of these metabolites.
Monoclonal antibodies can precipitate low-density lipoprotein. II. Radioimmunoassays with single and combined monoclonal antibodies for determining apolipoprotein B in serum of patients with coronary artery disease.
Authors: Marcovina S , Kottke B A , Mao S J
Published: January 2020

We have established four lines of monoclonal antibodies against human low-density lipoproteins (LDL) that, mixed in equal proportions, can precipitate LDL in gel and so can be used for apolipoprotein (apo) B determination in plasma. One monoclonal antibody (clone A), with a relatively low binding affinity to LDL (ka = 0.6 X 10(9) L/mol) and recognizing only two species of apo B, significantly underestimated the concentration of apo B in 74 patients with and 27 without coronary artery disease (CAD). High-affinity monoclonal antibody C (Ka = 3.8 X 10(9) L/mol), which recognized all four apo B species, gave the same value for apo B as determined with the mixture of monoclonal antibodies. The latter results (by radioimmunoassay, y) correlated well with those by radial immunodiffusion (chi): y = 0.994 chi + 0.003 (r = 0.987). The CAD patients showed an increased concentration of apo B as compared to the angiographically documented CAD-negative patients. Except for the values determined by clone B (p = 0.07), the increase was statistically significant (p = 0.002-0.018) for values determined by use of the other clones or their mixture.
Chemical nature of a synthetic bilirubin conjugate and its reactivities in the total and direct reactions by the Jendrassik-Gróf method.
Authors: Doumas B T , Wu T W , Poon K C , Jendrzejczak B
Published: January 2020

Using liquid chromatography, nuclear magnetic resonance spectroscopy, and elemental analyses, we verified that a commercially available synthetic bilirubin conjugate is predominantly a ditaurobilirubin (DTB) disodium salt. The Jendrassik-Gróf total bilirubin (TBIL) method quantitatively measures unconjugated bilirubin (Bu) and the Bu-equivalent content in DTB, from which we infer that the azopigments of Bu and DTB have identical molar absorptivities, which do not change in the presence of either serum or serum albumin of human or bovine origin. However, based on the ratio of direct bilirubin (DBIL) to TBIL, the DBIL reaction in dilute HCI is incomplete (even up to 20 min), with lower yields in a matrix of bovine serum albumin than in human serum. By contrast, the DBIL reaction in acetate buffer (pH 4.75) is quantitative for DTB in human serum (or its albumin), but less so in bovine serum (or its albumin). DTB is water soluble, is relatively stable when lyophilized with human serum, and is determined with such high precision in the above-mentioned endpoint assays that it may be a suitable surrogate calibrator for conjugated bilirubin.
Improved assay of antithrombin-III. Effects of certain additives on thrombin and on chromogenic peptide substrates.
Authors: Gram J , Jespersen J
Published: January 2020

When manual assays of antithrombin-III with use of different synthetic chromogenic peptide substrates are modified for a centrifugal analyzer, the downward deflection of the reference curve at the zero point raises a serious problem, because the position of the zero value is included in the calculation of the reference curve. Complete linearity was obtained by addition of polyethylene glycol 6000 to the thrombin solution and Tween 80 to the substrate solution in final concentrations of 1 g/L and 100 mg/L, respectively. The additives increased the concentration of active thrombin in the solutions by preventing its adsorption and aggregation, and the solubility of the substrates in aqueous media is increased. The precision of the optimized method, tested in routine assays during three months, yielded an estimated day-to-day CV of 3.4%.
Estriol and its conjugates in late pregnancy determined by extraction with Carbopack B and liquid chromatography with fluorometric detection.
Authors: Andreolini F , Borra C , Caccamo F , Di Corcia A , Nicoletti I , Samperi R , Improta F
Published: January 2020

We report a method for measuring estriol and its intact conjugates in urine, serum, and amniotic fluid. A single assay can be done within about 50 min, eight samples assayed in less than 5 h. A 70-microL urine sample is diluted and the estriol conjugates are adsorbed from it onto graphitized carbon black (Carbopack B, Supelco). After two washings, the analytes are desorbed with chloroform/methanol (60/40, by vol) containing tetrapropylammonium bromide. After solvent evaporation, the residue is redissolved in 100 microL of water/acetonitrile and 20 microL is injected into the chromatograph. Or 1 mL of serum or 0.5 mL of amniotic fluid is deproteinized with cold methanol, then passed through the Carbopack column. After three washings, the estriol and its conjugates are desorbed and treated as for urine. Mean analytical recoveries of the analytes in any of these body fluids were within about 92-98%, except for estriol-3-sulfate-16 alpha-glucuronide in serum (mean recovery 88.3%). The limit of sensitivity is well below the concentrations of clinical interest, and the method is not susceptible to substantial interferences.
Solid-phase extraction and liquid chromatography for improved assay of cyclosporine in whole blood or plasma.
Authors: Kabra P M , Wall J H , Blanckaert N
Published: January 2020

In this simple, precise, accurate, and specific isocratic liquid chromatographic procedure for determining cyclosporine, the cyclosporine is extracted from 1 mL of whole blood or from plasma, with 500 micrograms of cyclosporin D added per liter as internal standard, by elution from a Bond-ElutTM C18 extraction column with 300 microL of a mixture of ethanol and tetrahydrofuran. A 100-microL aliquot of the eluate, injected onto a cyano-phase analytical column, is eluted with a mixture of acetonitrile and pH 7.0 phosphate buffer at a flow rate of 1.0 mL/min and at 50 degrees C. Detection is at 210 nm. The chromatography is complete in less than 14.0 min. The method can measure less than 10.0 micrograms/L. Analytical recovery of cyclosporine added to whole blood ranged from 99 to 109% for concentrations up to 2000 micrograms/L. Between-run CVs ranged from 6.4 to 6.6%. None of numerous drugs and steroids tested interfered. Results by radioimmunoassay exceeded by 20 to 350% those measured by the present method.
Studies directed toward labeling analysis of angiotensin II in plasma.
Authors: Emanuel R L , Joppich-Kuhn R , Williams G H , Giese R W
Published: January 2020

We assay a 1-mL plasma sample containing angiotensin II (103 pg by radioimmunoassay) for the hormone by the following sequence of steps: add 125I-labeled val5-angiotensin II as an internal standard, extract on a C18 Sep Pak column, extract on an antibody affinity column, label the extract with an 125I Bolton-Hunter reagent, separate on a Bio Gel P2 column, and repetitively separate on a reversed-phase "high-performance" liquid-chromatographic column, detecting the eluting compounds by counting radioactivity. The fact that we measured 46 pg of angiotensin II-like substance per milliliter in a sample of pooled plasma is encouraging for the further development of this methodology. In particular, replacing the radioisotope with a more suitable chemical label such as an electrophoric (electron-capturing) release tag should be useful.
Five methods for measuring low-density lipoprotein cholesterol concentration in serum compared.
Authors: Hoffmann G E , Hiefinger R , Weiss L , Poppe W
Published: January 2020

Five methods for the quantification of low-density-lipoprotein cholesterol were compared: ultracentrifugation, electrophoresis, precipitation with polyvinyl sulfate or heparin, and an indirect calculation procedure (Friedewald formula). Excellent agreement of results was obtained with all procedures for 49 of 51 sera. Discrepancies were as much as 1.69 g/L for the remaining two cases, which contained appreciable amounts of "floating" beta-cholesterol as detected with a combination of ultracentrifugation and electrophoresis.
Estimation of the response-error relationship in immunoassay.
Authors: Sadler W A , Smith M H
Published: January 2020

Estimation of the response-error relationship in immunoassay provides a weighting function for the main analysis, and may in general be essential to ensure statistically valid data reduction. In this study we generated 50,000 sets of simulated radioimmunoassay response data with a computer, using five response-error functional forms that are commonly assumed. Parameters were estimated by three least-squares regression methods and three that are modifications of a maximum-likelihood method. Two likelihood estimators that require significantly different computing times were shown to be virtually indistinguishable, statistically more efficient than least-squares estimators, and-in contrast to least-squares estimators-to guarantee positive predicted variances in the range of the data.
Particle-counting immunoassay of a fetuin-like antigen in serum and cerebrospinal fluid.
Authors: Fagnart O C , Cambiaso C L , Sindic C J , Masson P L
Published: January 2020

A fetuin-like antigen was detected (smallest concentration detectable: 5 micrograms/L) by particle-counting immunoassay in 2% (13/641) of consecutive patients' sera but not in sera from 80 healthy blood donors, 40 neonates, or 40 pregnant women. The relation of the presence of detectable antigen to patients' diagnosis is not yet clear. However, in the group with cancer (154), it was found only in two of four patients with nephroblastoma and in three of five with tumors of tissue derived from the neurological crest: retinoblastoma (1/1), neuroblastoma (1/3), and medulloblastoma (1/1). Serum specimens from 422 patients with neurological disorders showed the antigen at a concentration greater than 5 micrograms/L in cases of neurosyphilis (5/11), peripheral neuropathy (12/38), Guillain-Barré syndrome (7/27), and multiple sclerosis (74/184). When we assayed 232 specimens of cerebrospinal fluid from the same neurological patients, we found the antigen in two cases of multiple sclerosis (6 and 15 micrograms/L) and in one case of Guillain-Barré syndrome (54 micrograms/L).
Free thyroxin measured in dried blood spots: results for 10 000 euthyroid and 29 hypothyroid newborns.
Authors: Travert G , Lemonnier F , Fernandez Y
Published: January 2020

We measured free thyroxin (FT4) in dried blood samples from 10 000 euthyroid and 29 hypothyroid newborns (three with transient hypothyroidism and 26 permanent). In euthyroids, the mean FT4 concentration was 16.9 (SD 4.5) pmol/L. Preterm babies had low concentrations of FT4, and values were closely related to birth weight when less than 2500 g. Assay of blood-spot FT4 appears to be specific as a test for hypothyroidism screening, yielding 0.42% false positives when a mean minus 2 SD cutoff value (8 pmol/L) was used. Correcting FT4 for birth weight further decreased the false-positive rate, to 0.07%. In all three newborns with transient hypothyroidism, FT4 was less than 8 pmol/L. In 25 of 26 permanent hypothyroids, FT4 ranged from undetectable to 5 pmol/L; in the 26th infant, who had a large ectopic gland, it was 8.2 pmol/L. We believe that FT4 assay offers an attractive improvement over total T4, whether performed as the first screening test or as a confirmatory test in thyrotropin screening programs.
Radioenzymatic microassay for simultaneous estimations of dopamine, norepinephrine, and epinephrine in plasma, urine, and tissues.
Authors: Hussain M N , Benedict C R
Published: January 2020

With this sensitive and specific radioenzymatic assay picogram quantities of norepinephrine, epinephrine, and dopamine can be simultaneously determined directly in plasma, urine, and tissue. We discuss in detail the steps that are critical for success of the assay, as well as the rationale behind the use of specific reagents. The assay is appropriate for use in any routine clinical chemistry laboratory. This assay is sensitive to approximately 1 pg of catecholamine. As compared with a commercially available method, this assay is significantly more economical and less time consuming.
Evaluation of a commercial immunoenzymometric assay for alpha-fetoprotein.
Authors: Hullin D A , Woodhead J S
Published: January 2020

A two-site immunoenzymometric assay (Abbott Diagnostics) for alpha-fetoprotein (AFP) in maternal serum and amniotic fluid has been evaluated for its suitability as a screening test for open neural tube defects. In a retrospective study based on 190 pregnancies of known outcome, performance of the kit in measuring both serum and amniotic fluid AFP correlated well with that of an in-house radioimmunoassay. Of 39 pregnancies associated with open neural tube defects, only four would not have been detected by the use of sequential measurement of serum and amniotic fluid AFP (also essentially in agreement with results obtained by the RIA). We conclude that this immunoassay could form the basis for a screening program for antenatal detection of open neural tube defects.
An automated liquid-chromatographic system for convenient determination of glycated hemoglobin A1c.
Authors: Saleh A K , Moussa M A
Published: January 2020

For quantification of glycated hemoglobin (HbA1c), we compared an automated system (Pharmacia Fast Protein Liquid Chromatography), which separates and determines HbA1c, with a commercial disposable-minicolumn kit (Boehringer-Mannheim), which separates total HbA1. We studied 41 diabetic women and 79 apparently normal women on their first postparturition day. The automated method was more precise (within-run CV 0.98-4.16%) than the kit method (within-run CV 3.67-7.77%). Results by the two methods correlated well for both control (p less than 0.001) and diabetic (p less than 0.05) groups. Values for HbA1c correlated significantly with fasting blood-glucose concentrations in controls and diabetics (intraclass correlation coefficient rI = 0.822 and 0.851, respectively, p less than 0.001) as well as with 1- and 2-h values for glucose after a 75-g glucose load in the control group (rI = 0.649 and 0.846, respectively, p less than 0.001). For HbA1 no such significant correlation was found except with values for fasting blood glucose in diabetics (rI = 0.745, p less than 0.001). Evaluation of HbA1c was a more sensitive index of glycemic status. The automated system is convenient, reliable, and easily operated.
Rapid measurement of free anticonvulsant drugs by direct liquid chromatography of serum ultrafiltrates.
Authors: Bock J L , Ben-Ezra J
Published: January 2020

In this simultaneous assay of unbound phenobarbital, phenytoin, primidone, and carbamazepine, serum ultrafiltrate is prepared by centrifugation in disposable filter units (Centrifree Micropartition System, Amicon Corp.) and injected directly onto a reversed-phase liquid-chromatography column. Drugs in the effluent are detected by absorbance at 210 nm. The measured concentrations were not critically dependent on the exact conditions of ultrafiltration (rotor angle, speed or time of centrifugation, amount of sample filtered). Pooled serum containing all four drugs gave reproducible results in repeated analyses during several days. For comparison we assayed 113 clinical specimens for one or more of these drugs by this method, and for the total (bound plus unbound) concentration of each drug by immunoassay. For each drug there was a nearly linear relationship between the two types of measurements, except for six specimens that had unusually high fractions of unbound drug. This assay appears highly suitable for routine determination of the free, biologically active fraction of anticonvulsant drugs, possibly as an alternative to measurements of total drug concentrations.
Accuracy and precision of five analog radioimmunoassays for free thyroxin compared.
Authors: Gow S M , Kellett H A , Toft A D , Beckett G J
Published: January 2020

We compared the precision of free thyroxin (FT4) measurements by kits involving analog RIA and the use of antibody-coated tubes (Becton Dickinson and Coat-A-Count), magnetic separation (Amerlex Magnetic and Corning Magic), or centrifugation of antibodies linked to solid beads (Amerlex). Results of kits with magnetic separation were the most reproducible. Amerlex, Amerlex Magnetic, and Becton Dickinson kits gave values comparing best with those obtained by direct equilibrium dialysis. Coat-A-Count and Corning Magic results differed significantly from dialysis values, both for patients' samples and kit standards. The kits had equal diagnostic efficiency in patients with suspected thyroid disease. On measurement of FT4 some patients were reclassified from "subclinical thyroid disease" to "overt disease." Most patients with triiodothyronine thyrotoxicosis had increased FT4. Several kit values were low for pregnant women and patients with nonthyroidal illness but the Amerlex and Amerlex Magnetic assays had fewer low results. The Amerlex Magnetic FT4 assay gave the best precision, agreement with the reference method, and diagnostic efficiency.
"Carryback": effect of viscous liquid controls on the preceding sample analyzed with the SMA II continuous-flow analyzer.
Authors: Winter S D , Kerr C , McAvoy M , Gallomore M D , Smith C , Lepoff R B
Published: January 2020

We systematically studied the "carryback" effect of ethylene glycol-based controls on the preceding sample on an SMA II continuous-flow analyzer. Including Beckman Level 1 unassayed liquid control material as a sample lowered the 12 analyte values of the preceding sample by an average of 2.7%, Level 2 (the most viscous) by an average of 4.4%, and Level 3 by 3.2%. Water-reconstituted lyophilized control material caused no carryback effect, but lyophilized control reconstituted with 330 mL/L ethylene glycol decreased the preceding sample's results by 4.1% (average carryback). We believe that carryback is caused by the drag placed on the sample line by a viscous sample, which decreases the volume of the preceding sample that is delivered to the reagent or pre-dilution mixing coils. Our findings were confirmed on another SMA II. Limited study of a SMAC analyzer gave inconclusive results, but further evaluation of continuous-flow systems for carryback is warranted. Carryback substantially increases total analytical variability.
Monoclonal antibody-based solid-phase immunoenzymometric assays for quantifying human immunoglobulin G and its subclasses in serum.
Authors: Papadea C , Check I J , Reimer C B
Published: January 2020

We developed quantitative immunoenzymometric assays for human IgG and its subclasses by using monoclonal antibodies, an avidin-biotin detection system and, as the calibrant, the U.S. National Reference Preparation for Specific Human Proteins. The assays are sensitive (detecting as little as 6 micrograms/L), precise (average inter-assay CV less than 11%), and vary linearly with concentrations over a five- to 10-fold range, depending on the monoclonal antibody. We evaluated 22 different monoclonal antibodies, many of which remained highly reactive when immobilized in wells of microtiter plates coated with bovine serum albumin-glutaraldehyde to "capture" total IgG or subclasses of IgG in the sample. We demonstrated the specificity of the most reactive antibodies by using a panel of 20 purified myeloma proteins. The sum of IgG subclass concentrations correlated well (r = 0.84, p less than 0.001) with the total IgG measured in sera from 63 apparently healthy adults (26 men, 37 women). We estimated 95 percentile reference intervals for the immunoglobulins in these subjects and determined the following mean percentage distributions of IgG subclasses: IgG1 49, IgG2 33, IgG3 9, and IgG4 7. The availability of these assays should facilitate studies of the clinical significance of the subclasses.
Free thyroxin and free triiodothyronine as measured by equilibrium dialysis and analog radioimmunoassay in serum of patients taking phenytoin and carbamazepine.
Authors: Liewendahl K , Tikanoja S , Helenius T , Majuri H
Published: January 2020

We measured free thyroxin (FT4) and free triiodothyronine (FT3) in serum of patients taking the anti-epileptic drugs phenytoin and carbamazepine, both by equilibrium dialysis procedures and analog-type radioimmunoassays. By either assay, the mean concentration of FT4 was significantly decreased in patients receiving either drug, whereas their FT3 concentrations were normal or only slightly decreased. Adding therapeutic concentrations of these drugs in vitro to control sera had a small or no incremental effect on FT4 and FT3 as measured by either method, but adding greater concentrations of the drugs in vitro markedly increased the concentrations of the free hormones. These results indicate that the main mechanism of the decrease in concentrations of free thyroid hormones in serum during therapy with anticonvulsant drugs is not the displacement of hormones from their binding to plasma proteins. We also determined, using a new and sensitive immunoradiometric assay, that patients taking carbamazepine, but not those taking phenytoin, had significantly less thyrotropin in the serum.
Hyperamylasemia, specific pancreatic enzymes, and hypoxanthine during recovery from diabetic ketoacidosis.
Authors: Møller-Petersen J , Andersen P T , Hjørne N , Ditzel J
Published: January 2020

The contribution of the exocrine pancreas to hyperamylasemia in diabetic ketoacidosis was investigated by measuring total amylase, salivary and pancreatic isoamylases, cathodic trypsin-like immunoreactivity, and pancreatic lipase in 12 consecutive patients recovering from diabetic ketoacidosis. Hyperamylasemia was present in six of the patients [50%; expected incidence: 21.1-78.9% (95% confidence limits)]--in five with simultaneously increased activities of all three specific pancreatic enzymes, and in one with only increased salivary isoamylase. The serum concentration of hypoxanthine--an indicator of the cellular energy state--was above normal in all patients at admission. We found no differences in concentrations of hypoxanthine in serum of patients with or without hyperamylasemia or in patients with or without increases in the specific pancreatic enzymes. In none of the patients was the clinical course or the time--concentration curves of the pancreatic enzymes consistent with acute pancreatitis. The pathogenic mechanism leading to hyperamylasemia in diabetic ketoacidosis remains uncertain.
Effects of dextran on five biuret-based procedures for total protein in serum.
Authors: Barnes D B , Pierce G F , Lichti D , Landt M , Koenig J , Chan K M
Published: January 2020

We evaluated the effect of dextran on values for total protein in serum as measured by the biuret method with five widely used automated instruments: the American Monitor Parallel; the Du Pont aca II; the Roche Cobas-Bio; the Kodak Ektachem 400; and the Beckman Astra 8. Dextran concentrations as great as 25 or 30 g/L had relatively little or no influence on total protein measurements by the latter three instruments. Dextran concentrations exceeding 6 g/L caused falsely low results with the aca, whereas the Parallel gave falsely high results when the dextran concentration exceeded 2 g/L. The aca total protein procedure could be protected from the interference by dextran concentrations up to 30 g/L by injecting 0.4-0.8 mL of ethylene glycol directly into the reagent pack before sampling. However, we could not eliminate the interference with the Parallel procedure by any simple means; we thus recommend that it not be used for measuring total protein in serum samples from patients who are being treated with dextran.
Anti-IgG combined with rate nephelometry for measuring polyethylene glycol-precipitated circulating immune complexes.
Authors: Feldkamp C S , Levinson S S , Perry M , Amin V
Published: January 2020

Immune complexes from serum were assayed for IgG by a simple rate-nephelometric method after extraction with polyethylene glycol that removes monomeric IgG. A 30-min preincubation of the extracted material in reaction buffer before the anti-IgG is introduced eliminates falsely increased values owing to precipitation that increased baseline light scatter in the reaction buffer. We found good parallelism in the reaction of anti-IgG with the IgG calibrator, aggregated human globulin, or endogenous immune complex. Thus IgG can be used for calibration in place of aggregated human globulin, greatly simplifying the assay. A good correlation was found between the present assay and the C1q-binding test (r = 0.83). The present assay is both sensitive and reproducible. The extraction and assay are straightforward and can be completed in a single morning after an overnight precipitation. The reagents for extraction are easily prepared and inexpensive, and the materials for assay are available in kit form. We believe this approach to be well suited for many clinical laboratories to measure circulating immune complexes.
A monoclonal-antibody enzyme immunoassay for detection of hepatitis B surface antigen with use of a biotin-avidin system.
Authors: Liu Y S , Green A
Published: January 2020

In this solid-phase two-site enzyme immunoassay for hepatitis B surface antigen (HBsAg), three monoclonal anti-HBs are used: 5D3 (IgM) is immobilized on plastic beads; 5C3 (IgG2a) and 5C 11 (IgG1), labeled with biotin, are used as the first conjugate. Horseradish peroxidase covalently linked to avidin is the second conjugate. First, serum or plasma is incubated with the antibody-coated bead and biotin-labeled antibodies, simultaneously, at 45 degrees C for 1 h ("stat" procedure), 3 h ("standard" procedure), or 18 h ("overnight" procedure), during which HBsAg forms a complex with the solid-phase antibody and the biotinylated antibodies. The enzyme-conjugated avidin is then bound to the biotin on the antigen-antibody complex at 45 degrees C for 15 min ("stat") or 30 min (standard and overnight procedures). The beads are incubated with enzyme-substrate solution (H2O2 and o-phenylenediamine). Color developed is measured at 492 nm. All procedures satisfied third-generation HBsAg tests required by the FDA Office of Biologics, being sensitive both to ad and ay subtypes in subnanogram amounts. The assay is reactive with adw2, adw4, adr, ayw2, ayw3, and ayr subtypes and can detect viral determinants in HBsAg-anti-HBs immune complex form. Thus it provides a sensitive, simple, and reproducible alternative to radioimmunoassay.
Apparent binding of ionized calcium by various buffers.
Authors: Crowell J A , Bowers G N
Published: January 2020

Using different substance concentrations in an aqueous 1.25 mmol/L solution of CaCl2 plus NaCl to a final solution ionic strength of 160 mmol/L, we tested six buffers for their effect on measurements of ionized calcium (Ca2+). Measured Ca2+ decreased with increasing ionic strength and pH. Increasing concentrations of Tris caused a positive Ca2+ electrode bias; the other five buffers caused a negative Ca2+ electrode bias with increasing concentration. 4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid ("HEPES") at 10 mmol/L maintained pH 7.39 in the aqueous reference solutions containing 0.75 to 2.50 mmol of CaCl2 per liter, and showed a linear Ca2+ response among three different suppliers of Ca2+ instruments; it is thus a suitable buffer for a reference solution.