bluuebunny/crossref_metadata_2025 huggingface dataset:
Seven Different Assays of Hyaluronan Compared for Clinical Utility
Authors: Lindqvist Ulla , Chichibu Kenji , Delpech Bertrand , Goldberg Ronald L , Knudson Warren , Poole A Robin , Laurent Torvard C
Published: January 2020

To compare six assays of hyaluronan (hyaluronic acid; HYA) in serum, developed in different laboratories, we analyzed 10 samples from each of three groups: healthy persons, patients with primary biliary cirrhosis, and patients with rheumatoid arthritis. All the assays are based on the use of affinity proteins specific for HYA, prepared from cartilage or brain tissue, and are analogous to RIA or enzyme immunoassay techniques. The assay results were of the same magnitude. Although statistical analysis indicated that the methods in some cases deviated significantly from one another, this variation was less than the physiological variation in the healthy population. Therefore, the results of clinical investigations in which the various methods have been used are comparable. The analyses have high specificity and sensitivity for primary biliary cirrhosis but are somewhat less suitable for detecting rheumatoid arthritis. A seventh laboratory, which obtained antibodies to HYA, used these in an RIA to analyze a separate series of serum specimens. Results were in agreement with those obtained by one of the other assays.
Polypeptide Nicks Cause Erroneous Results in Assays of Human Chorionic Gonadotropin Free β-subunit
Authors: Kardana Andrew , Cole Laurence A
Published: January 2020

Pregnancy and trophoblastic disease testing laboratories measure human chorionic gonadotropin (hCG; human choriogonadotropin) free beta-subunit to screen for Down syndrome and to diagnose persistent trophoblastic disease (invasive mole and choriocarcinoma). The results from various laboratories, however, vary widely for similar groups of patients. Average concentrations of free beta-subunit reported for persistent trophoblastic disease, for instance, range from 2.6% to 37% of total hCG. On hCG and its free beta-subunit, peptide bonds can be missing between beta-subunit residues 44 and 45 or between residues 47 and 48 (nicked molecules). To explore the possibility that the disparity in the reported concentrations of free beta-subunit was due to differences in recognition of nicked molecules by different antibodies, we generated 0% and 100% nicked beta-subunit standards and investigated their recognition in three separate immunoassays of free beta-subunit. The immunoassay with antibody 1E5 did not recognize nicked beta-subunit (4% cross-reactivity with nicked beta-subunit) and thus detected only intact beta-subunit. The immunoassay with antibody FBT11 gave similar results with nicked and nonnicked thus standards (96% cross-reactivity with nicked beta-subunit), as did the assay with antibody B204 (73% cross-reactivity with nicked beta-subunit). These two immunoassays thus measured total (nicked + nonnicked) beta-subunit. We used the three immunoassays to examine sera from normal pregnancy, Down syndrome pregnancy, hydatidiform mole, and persistent trophoblastic disease, all of which contain nicked and nonnicked beta-subunit molecules. The results obtained resembled the studies with nicked beta-subunit standards. The results from the FBT11 and B204 assays (total beta-subunit) were highest, results from the 1E5 assay (intact molecules only) being as much as 10 times lower. We conclude that nicks in beta-subunit and the extent of recognition of nicked molecules by different antibodies affect the concentrations reported for free beta-subunit.
Ion-Trap Detection of Volatile Organic Compounds in Alveolar Breath
Authors: Phillips M , Greenberg J
Published: January 2020

We describe a method for the collection and microanalysis of the volatile organic compounds in human breath. A transportable apparatus supplies subjects with purified air and samples their alveolar breath; the volatile organic compounds are captured in an adsorptive trap containing activated carbon and molecular sieve. The sample is thermally desorbed from the trap in an automated microprocessor-controlled device, concentrated by two-stage cryofocusing, and assayed by gas chromatography with ion-trap detection. Compounds are identified by reference to a computer-based library of mass spectra with subtraction of the background components present in the inspired air. We used this device to study 10 normal subjects and determined the relative abundance of the volatile organic compounds in their alveolar breath. The breath-collecting apparatus was convenient to operate and was well tolerated by human volunteers.
Fecal Coproporphyrin Isomers in Hereditary Coproporphyria
Authors: Blake Dennis , McManus Julie , Cronin Virginia , Ratnaike Sujiva
Published: January 2020

To see whether the fecal coproporphyrin III:coproporphyrin I (CIII:CI) ratio (determined by HPLC) would be suitable for screening patients at risk of hereditary coproporphyria (HC), we compared such ratios with the lymphocyte coproporphyrinogen oxidase (EC 1.3.3.3) activities (COOX) in 38 subjects from one large family and two smaller families with HC. The CIII:CI ratio was normal (less than 1.3) in adults with normal COOX (greater than 180 nmol/g of protein per hour) and high (greater than 2) in those with low COOX. Results were difficult to interpret in six of 10 children, who had borderline or low COOX but normal fecal CIII:CI ratios. Five subjects with low COOX and abnormal fecal CIII:CI ratios had normal fecal total porphyrin, indicating that the latter investigation alone is inadequate for family studies. The sample for determining the fecal CIII:CI ratio is easier to obtain and the assay is technically less demanding than COOX. We found the fecal CIII:CI ratio suitable for investigation of adults in a family study, but its usefulness in children needs to be established.
Spectrotype Distributions of Circulating IgG from Patients with Systemic Lupus Erythematosus
Authors: Barbi G , Keil L B , Gaito A D , DeBari V A
Published: January 2020

Pathogenic autoantibodies from patients with systemic lupus erythematosus (SLE) may represent a relatively cationic fraction of IgG. We compared the spectrotype distributions of affinity-purified IgG from the sera of 10 SLE patients and 10 age- and sex-matched control subjects. Purified IgG was subjected to isoelectric focusing between pI 3 and 9. No significant difference was observed for pI 6.0-6.5 and 7.5-8.0. However, control subjects had a higher percent of total IgG at 6.5-7.0 (15.4 +/- 4.0 vs 11.1 +/- 2.0, P = 0.008) and at 7.0-7.5 (22.4 +/- 4.8 vs 18.2 +/- 3.8, P = 0.04) whereas SLE patients had a higher percent of total IgG at 8.0-8.5 (24.3 +/- 3.0 vs 20.5 +/- 4.0, P = 0.03) and at 8.5-9.0 (21.9 +/- 5.9 vs 15.1 +/- 3.7, P = 0.006). Spectrotype distributions of circulating IgG from SLE patients are skewed toward higher pI, providing further evidence of proliferation of B-cell clones that express more cationic IgG in patients with SLE. Longitudinal studies of serum IgG from several patients for > 1 year reveal distinct changes in both cationic and anionic clonotypes, suggesting clonal expansion of antiidiotypes to autoantibodies.
Using Changes in Attributable Risk to Predict Long-Term Efficacy of Simvastatin Treatment
Authors: Vermaak W J , Ubbink J B , Ungerer J P , Marais A D , Firth J , Van Lathem J M , Becker P J
Published: January 2020

It is a common perception that coronary heart disease (CHD) deaths will decline by 2% for every 1% decrease in serum total cholesterol (TC). We investigated the changes in serum TC concentrations obtained with simvastatin, an inhibitor of hydroxymethylglutaryl-CoA reductase, in 70 hypercholesterolemic subjects. Although simvastatin reduced serum TC concentrations by 30% after 6 months, calculations from age- and risk-related TC quintiles show that the absolute difference in attributable CHD deaths would be just > 1%. These results are reminiscent of the experience from several large prospective primary drug trials that demonstrated impressive relative benefit ratios but much smaller absolute differences (< 2%) in CHD endpoints between the treated and untreated groups. This epidemiologically based approach of assessing efficacy and potential long-term benefit of a lipid-lowering drug may be more appropriate and potentially less misleading than just the percentage change of TC (or other lipoproteins) so frequently reported in short-term drug trials.
Transferrin Index: An Alternative Method for Calculating the Iron Saturation of Transferrin
Authors: Beilby J , Olynyk J , Ching S , Prins A , Swanson N , Reed W , Harley H , Garcia-Webb P
Published: January 2020

We surveyed 140 clinical chemistry laboratories in Australia to establish which laboratory methods they used to determine serum iron status: 125 measured serum iron (Fe), 85 measured transferrin (TRF), 47 measured total iron-binding capacity (TIBC), and 14 measured both TRF and TIBC. Of the 55 laboratories routinely reporting TRF saturation (TS), 16 calculated TS directly as (Fe/TIBC) x 100, and 9 used [Fe/(TRF x 2)] x 100. Thirty laboratories measured TRF and converted it to an equivalent TIBC concentration; the derived TIBC was then used to calculate TS. We measured iron, TIBC, and TRF concentrations in 94 control subjects, 59 patients with alcoholic liver disease (ALD), and 20 with proven genetic hemochromatosis (GH). TS was compared with a transferrin index (TI = Fe/TRF) to determine whether both methods were sensitive for GH screening and which method gave the fewest false-positive results with discrimination limits of > 55% and > 1.0, respectively. All GH patients were detected by both TS and TI at these limits. One control subject had a TI > 1.0, whereas three control subjects had a TS > 55%. Nine patients with ALD had a TI > 1.0 and 11 ALD patients had a TS > 55%. Some iron-overload patients had lower than expected TS values compared with TI, possibly because of ferritin interference in the TIBC assay. Also, the precision of the TRF assay was better than that of the TIBC assay: CVs of 1.85-3.68% vs 6.17%. We therefore recommend that calculated TI replace TS in screening for iron overload.
Serotonin Concentrations in Plasma and Variations During the Menstrual Cycle
Authors: Hindberg I , Naesh O
Published: January 2020

We used a specific and sensitive radioenzymatic method to establish a reference interval for the concentration of serotonin in platelet-poor plasma in 98 healthy volunteers (49 men, 49 women). The interval was 0-11 nmol/L with a median of 2.8 nmol/L. No difference in concentration in relation to sex or age was observed. In a group of eight very old volunteers (ages 86-92 years), however, concentrations were increased. In addition, we monitored the plasma concentrations of serotonin in 20 healthy women (ages 26-45 years) through two menstrual cycles. Periovulatory and premenstrual concentrations were greater than the serotonin concentration at the start of menstruation.
Two Novel Nonradioactive Polymerase Chain Reaction-Based Assays of Dried Blood Spots, Genomic DNA, or Whole Cells for Fast, Reliable Detection of Z and S Mutations in the α1-Antitrypsin Gene
Authors: Andresen B S , Knudsen I , Jensen P K , Rasmussen K , Gregersen N
Published: January 2020

Two new nonradioactive polymerase chain reaction (PCR)-based assays for the Z and S mutations in the alpha 1-antitrypsin gene are presented. The assays take advantage of PCR-mediated mutagenesis, creating new diagnostic restriction enzyme sites for unambiguous discrimination between test samples from individuals who are normal, heterozygous, or homozygous for the mutations. We show that the two assays can be performed with purified genomic DNA as well as with boiled blood spots. The new assays were validated by parallel testing with a technique in which PCR is combined with allele-specific oligonucleotide (ASO) probes. In all cases tested the results obtained by the different techniques were in accordance. The new assays can be used for prenatal diagnostics and can be performed directly with boiled tissue samples. Because the new assays are easy to perform and reliable, we conclude that they are well suited for routine diagnosis.
Fluorometric Enzyme Immunoassay of Basic Fibroblast Growth Factor with Monoclonal Antibodies
Authors: Kurobe M , Kato A , Takei Y , Hayashi K
Published: January 2020

We compared three different strategies for measuring basic fibroblast growth factor (bFGF) by fluorometric enzyme immunoassay (EIA). After optimizing conditions, we found that a primary anti-bFGF MAb directly conjugated with peroxidase gave the best detection limit for recombinant bFGF (approximately 30 ng/L, 3 pg/assay tube) in a two-site sandwich assay. The detection limit of methods based on biotinylated primary MAbs or on secondary antibodies followed by streptavidin-conjugated peroxidase was slightly lower than that of the above method. Using the most sensitive EIA examined in this study, we made a preliminary measurement of immunoreactive bFGF in sera of apparently healthy people and found it to be 190 (SD 32) ng/L (n = 48), in agreement with an earlier reported value (30-206 ng/L). Also, the concentration of immunoreactive bFGF in sera was above normal in 19 of 31 patients with stomach cancer.
Abbott IMX Evaluated for Assay of Prostate-Specific Antigen in Serum
Authors: Dnistrian A M , Schwartz M K , Smith C A , Nisselbaum J S , Fair W R
Published: January 2020

We evaluated a new fully automated procedure for quantitative measurement of prostate-specific antigen (PSA) by the Microparticle Enzyme Immunoassay (MEIA) technology developed for the Abbott IMx automated immunoassay system. The performance characteristics of the Abbott IMx PSA assay (y) were evaluated and compared with those of the Hybritech Tandem-E PSA assay (x), a solid-phase two-site immunoenzymometric assay. PSA values for both assays were well correlated (r = 0.99); regression analysis yielded the equation y = 0.92x - 0.23 micrograms/L. The Abbott assay proved reliable and reproducible, as shown by the intra- and interassay coefficients of variation (2.0-3.4% and 3.1-4.7%, respectively). The assay gave a linear standard curve up to 100 micrograms/L and was very sensitive (detected PSA < 0.1 microgram/L). This analytical sensitivity was comparable with that of the Tandem-E PSA assay. Overall, the IMx PSA assay demonstrated the accuracy, precision, linearity, and intermethod correlation required for monitoring patients with prostate cancer.
Washing Erythrocytes to Remove Interferents in Measurements of Zinc Protoporphyrin by Front-Face Hematofluorometry
Authors: Hastka J , Lasserre J J , Schwarzbeck A , Strauch M , Hehlmann R
Published: January 2020

Zinc protoporphyrin (ZPP) is determined by hematofluorometry of whole blood to detect iron deficiency in blood donors. In hospitalized patients, ZPP did not correlate with established markers of iron status. We performed 4500 ZPP measurements with the Aviv front-face hematofluorometer in samples from 475 patients and measured ferritin, transferrin saturation, hemoglobin, and erythrocyte indices. We found that the fluorometric determination is affected by substances dissolved in plasma but that this interference can be eliminated by using washed erythrocytes. In validation tests the within-day variation was < 3.5%; the day-to-day variation was < 6.8%. In 130 healthy persons without iron deficiency, ZPP was < or = 40 mumol/mol heme, which we consider a normal value. Mean ZPP in 46 iron-deficient patients was 256 (SD 105) mumol/mol heme (correlation with ferritin: -0.73; with hemoglobin: -0.85; P < 0.001). When washed erythrocytes are used, the hematofluorometric determination of ZPP is sensitive and specific for detecting iron deficiency in otherwise healthy individuals and hospitalized patients.
Clinical Evaluation of a Pancreatic Lipase Mass Concentration Assay
Authors: van Ingen H E , Sanders G T
Published: January 2020

An immunoactivation assay for determining pancreatic lipase mass concentration was clinically evaluated and compared with results obtained by measuring total amylase and pancreatic amylase activity. A group of 30 patients with pancreatitis was compared with a control group of 32 patients in which this disease was suspected but excluded. Both lipase mass concentration and pancreatic amylase activity exhibit good sensitivity (0.93 each) and specificity (0.94 and 0.97, respectively) at cutoff concentrations of 200 micrograms/L and 200 U/L, respectively. The median increase in lipase mass concentration (37.1 times the upper limit of the reference interval) in the pancreatitis group was higher than that for either total amylase or pancreatic amylase activity (5.94 and 14.5 times, respectively) but showed a similar time to peak value. We conclude that the lipase assay is the method of choice for diagnosing pancreatitis.
Bone-Resorption Markers Galactosyl Hydroxylysine, Pyridinium Crosslinks, and Hydroxyproline Compared
Authors: Bettica P , Moro L , Robins S P , Taylor A K , Talbot J , Singer F R , Baylink D J
Published: January 2020

We compared the clinical performances of four bone-resorption (BR) assays (hydroxyproline, HYP; galactosyl hydroxylysine, GHYL; deoxypyridinoline, DPD; and pyridinoline, PYD) in subjects with different BR rates: normal (adult men and premenopausal women), mildly increased (postmenopausal osteoporotic women), high (Paget disease patients), and very high (children). The discrimination power (Z score) and the accuracy (estimated by receiver-operating characteristic analysis) for GHYL, DPD, and PYD were compared with those for HYP. Discrimination power and accuracy were similar for high- and very-high-BR groups for all four assays. However in the mildly increased-BR group, DPD, GHYL, and PYD showed a higher discrimination power and accuracy than did HYP. The clinical performances of HYP, DPD, GHYL, and PYD are comparable for large changes in BR. For modest changes, DPD, GHYL, and PYD are more accurate and have a higher discrimination power than does HYP.
Failure of Microchromatographic Measurement of Fetal Hemoglobin in β0 Thalassemia-Hereditary Persistence of Fetal Hemoglobin
Authors: Krauss J S , Jonah M H , Devoe L D , Pantazis C G
Published: January 2020

We report microchromatographic measurement of fetal hemoglobin (HbF) proportions in a 36-year-old African-American multigravida woman. At 34 weeks she delivered a 630-g male infant who subsequently did well. Hemoglobin electrophoresis of the hemolysate revealed nearly 100% HbF without HbA, an extremely unusual naturally occurring sample. Family studies revealed a combination of hereditary persistence of fetal hemoglobin (HPFH) and beta zero-thalassemia minor. Southern blot technique confirmed heterozygous alpha 2 thalassemia and HPFH but failed to identify the beta thalassemic lesion. The absence of HbA and the very high amounts of HbF led us to measure HbF by several methods to confirm the accuracy of microchromatography of HbF at values approaching 100%. HPLC revealed a 14% F1 suggestive of microchromatographic underestimation due to glycated HbF. We conclude that cation-exchange microchromatography and the Betke method of alkali denaturation underestimate HbF values as they approach 100% and do not recommend these procedures in this rare situation.
Creatine Kinase MB Isoforms in Patients with Skeletal Muscle Injury: Ramifications for Early Detection of Acute Myocardial Infarction
Authors: Wu A H , Wang X M , Gornet T G , Ordóñez-Llanos J
Published: January 2020

We measured total creatine kinase (CK), CK-MB isoenzyme, and the MB isoforms in 202 serum and plasma samples from nine groups of patients and normal individuals: 39 with acute myocardial infarction (MI), divided according to time between the onset of chest pain and blood collection (1-6 h, 7-12 h, and 13-48 h); 26 with chest pain for whom an MI was ruled out, sampled at admission; 17 undergoing bypass surgery or cardiac catheterization, sampled within 6 h after either procedure; 17 with acute skeletal muscle injury, sampled within 8 h after injury; 30 marathon runners immediately after a race; 17 runners and other athletes > 12 h after training or a race; 12 with cerebral injury or seizures, sampled at admission; 8 with closed head injury, sampled at admission; and 38 normal subjects. CK-MB (relative index) and MB isoforms (MB2/MB1) were respectively increased in 15% and 75% of MI patients 1-6 h after onset, 94% and 94% after 7-12 h, and 88% and 8% after 12 h, and in 87% and 82% of cardiac surgery patients. MB isoforms were increased in most patients with acute skeletal muscle trauma and in subjects examined after exercise, but were within normal limits in patients for whom MI was ruled out, patients with cerebral trauma, and normal individuals. The relative index of MB/total CK was normal in essentially all individuals in the last groups, including those with acute skeletal muscle trauma. We concluded that the CK-MB isoform ratio is increased in both acute skeletal muscle injury and MI. The isoform ratio is most useful for distinguishing recent from old (> 12 h) injury.
Fluorescence Polarization Immunoassay and HPLC Assays Compared for Measuring Monoethylglycinexylidide in Liver-Transplant Patients
Authors: Chen Y , Potter J M
Published: January 2020

We compared fluorescence polarization immunoassay (FPIA, x) and HPLC (y) for measuring monoethylglycinexylidide (MEGX) concentrations in 119 serum samples from 61 liver-transplant donors and recipients. The correlation between the two methods was y = 1.48 micrograms/L + 0.8x (r = 0.89). The bias (mean difference) was 12 micrograms/L (0.055 mumol/L) through the MEGX concentration range measured (0-250 micrograms/L, 0-1.136 mumol/L). We observed a major difference between the two methods in samples from four recipients and one donor. Cross-reactivity in FPIA with lignocaine and two of its metabolites (glycinexylidide and 2,6-xylidine) was < 3%. Samples with high bilirubin concentrations (> 200 mumol/L) required dilution before assay of MEGX by FPIA. Although there was an increase in apparent MEGX concentrations in some samples with increased bilirubin concentrations, the relationship was not constant. Increased plasma concentrations of cholesterol and triglyceride resulted in relatively small increases in apparent MEGX concentrations.
Performance of an Enzyme-Linked Immunosorbent Assay System for Antibodies to Hepatitis C Virus with Two New Antigens (c11/c7)
Authors: Saito M , Hasegawa A , Kashiwakuma T , Kohara M , Sugi M , Miki K , Yamamoto T , Mori H , Ohta Y , Tanaka E
Published: January 2020

We developed an enzyme-linked immunosorbent assay (ELISA) system for antibodies to the hepatitis C virus (HCV), using two new recombinant antigens (c11 and c7) derived from the HCV genome. The performance of this ELISA system (Imucheck HCV Ab) was examined. The CV values for both intra-assay precision and reproducibility of identifying HCV antibody in the panel sera ranged from 3.5% to 6.4%. The blood elements in serum and anticoagulants did not interfere in this ELISA system. The specificity of Imucheck HCV Ab to samples from patients with non-A, non-B (NANB)-type chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma was 93.7%, 93.5%, and 81.4%, respectively. These results are more sensitive than those obtained by the first-generation anti-HCV ELISA system. In the samples from patients with NANB-type acute hepatitis, Imucheck HCV Ab enabled detection of HCV antibodies at an early stage. This system increased the sensitivity for blood donor screening and for monitoring patients with acute hepatitis.
Determination of β2-Microglobulin in Serum by a Microparticle-Enhanced Nephelometric Immunoassay
Authors: Viedma J A , Pacheco S , Albaladejo M D
Published: January 2020

This fully automated nephelometric immunoassay to quantify beta 2-microglobulin in human serum measures the light-scattering signal produced by agglutination of commercially available latex microparticles (diameter 0.1 micron) coated with specific F(ab')2 against beta 2-microglobulin. The calibration curve, generated by serial dilutions of a beta 2-microglobulin standard of known concentration, is used to calculate beta 2-microglobulin concentrations in serum samples by the logit-log function and linear-regression analysis. The assay range (sample dilution 400-fold) extends from 0.3 to 40.0 mg/L. No antigen excess appears at beta 2-microglobulin concentrations up to 320 mg/L. Within-run CVs ranged from 1.0% to 3.4%, and between-days from 1.2% to 2.8%. Total imprecision (CV) was < 5%. Analytical recovery averaged 99.5% +/- 2.8%. Rheumatoid factor, complement, bilirubin (up to 340 mumol/L), and hemoglobin (up to 2.0 g/L) do not interfere. Strongly turbid lipemic samples must be cleared before analysis. Standard curve linearity was very good in samples covering the clinical useful range of concentrations. Results of the method correlated well with those of radioimmunoassay and microparticle enzyme-linked immunoassay (r = 0.979 and 0.975, respectively). The reference interval (nonparametric estimation) in apparently healthy adults (n = 303) was 0.87 (0.80-0.94) to 2.42 (2.28-2.45) mg/L; the median value was 1.54 mg/L.
Measurement of Free Desipramine in Serum by Ultrafiltration with Immunoassay
Authors: Hursting M J , Clark G D , Raisys V A , Miller S J , Opheim K E
Published: January 2020

We developed an ultrafiltration method for assaying free desipramine (DMI) in serum. An ultrafiltrate of DMI-containing serum was prepared by centrifugation through an Amicon Centrifree micropartition filter. Syva DMI solid-phase extraction (SPE) columns were used to extract the DMI from the serum and ultrafiltrate. The Syva monoclonal EMIT assay was used to quantify the DMI in the extract. In some experiments, the percent free DMI was quantified with radioactivity. Nonspecific losses of DMI in serum to the ultrafilter system were low (recoveries > 91%). Extraction of [3H]DMI from phosphate-buffered saline (to mimic serum ultrafiltrate) with the Syva SPE system was quantitative (recoveries of 98.4% +/- 4.6%). Free DMI concentrations, derived from serum containing 2.5-2500 micrograms/L DMI, were determined by ultrafiltration; results agreed well with values determined by equilibrium dialysis, the average percent of free DMI being 18.4% +/- 0.25% and 15.9% +/- 0.51%, respectively. To increase the sensitivity of the free DMI assay in the therapeutic range (total DMI 125-300 micrograms/L), we increased fourfold the ultrafiltrate volume applied to the SPE column. For free DMI at 11-130 micrograms/L, the within-run and between-run CVs for the ultrafiltration method were < 9% and < 15%, respectively. Binding of DMI to serum proteins decreased over the pH range 6.0-8.0, although temperatures between 20 and 28 degrees C did not affect binding. The ultrafiltration assay is fast, accurate, simple, and adaptable to standard laboratory instrumentation.
Missense Mutations of the Tissue-Nonspecific Alkaline Phosphatase Gene in Hypophosphatasia
Authors: Henthorn P S , Whyte M P
Published: January 2020

Hypophosphatasia is an inborn error of metabolism that is characterized clinically by defective bone mineralization and biochemically by deficient activity of the tissue-nonspecific isoenzyme of alkaline phosphatase (TNSALP) in serum and in tissues. Clinical severity is extremely variable, ranging from death in utero to pathologic fractures first presenting in adulthood. Severe forms of the disease are inherited in an autosomal recessive fashion; the modes of transmission of mild forms are uncertain. Deficiency of TNSALP activity in this condition suggests that mutations in the TNSALP "candidate" gene are the primary defects. This hypothesis was supported in 1988 by the demonstration, in one inbred infant, that an identical missense mutation in both alleles of the gene encoding TNSALP caused lethal hypophosphatasia. Here we summarize the work leading to that discovery and discuss the recent identification of additional missense mutations in the TNSALP gene associated with the entire clinical spectrum of hypophosphatasia.
Phospholipase Resistance of the Glycosyl-Phosphatidylinositol Membrane Anchor on Human Alkaline Phosphatase
Authors: Wong Y W , Low M G
Published: January 2020

Alkaline phosphatase (ALP) is attached to the cell surface in mammalian tissues via a glycosyl-phosphatidylinositol (GPI) anchor and can be released from the membrane by GPI-specific phospholipases. In a range of cultured human cell lines, however, the sensitivity of ALP to phospholipases was observed to be variable in magnitude (approximately 20-90%). The mechanism of phospholipase resistance was explored with phospholipases of different bond specificities. The results suggest that phospholipase resistance is the result of acylation of the inositol ring in the GPI anchor. The occurrence of phospholipase-resistant forms of ALP may have important implications for the in vivo release and disposition of plasma ALP.
Alkaline Phosphatase Isoenzyme Patterns in Malignant Disease
Authors: Van Hoof V O , Van Oosterom A T , Lepoutre L G , De Broe M E
Published: January 2020

Early treatment of patients with malignant disease and liver or bone metastasis may increase their survival time. We have used the activity patterns of liver and bone isoenzymes of alkaline phosphatase (ALP), separated by agarose gel electrophoresis, to detect early metastasis. We studied ALP isoenzyme patterns in a background population of 101 patients with no evidence of any disease that might influence this pattern; a healthy reference population (n = 330); and the following three groups of patients: 143 with malignant disease, 47 with nonmalignant liver disease, and 22 with nonmalignant bone disease. Cutoff and predictive values of liver ALP, high-molecular-mass (high-M(r)) ALP, and bone ALP were established for detecting liver and bone metastasis. The positive predictive value of liver and high-M(r) ALP was higher than that of total ALP in detecting liver metastasis, but liver and high-M(r) ALP did not enable us to differentiate between malignant and nonmalignant liver disease. Total ALP activity was of slightly more value than liver and high-M(r) ALP in enabling us to rule out liver metastasis. From bone ALP activity we could not distinguish between nonmalignant bone disease and bone metastasis. The negative predictive value of bone ALP in the diagnosis of bone metastasis was low, but its positive predictive value was high and superior to that of total ALP.
Cluster Analysis in Diagnosis
Authors: Vogt Wolfgang , Nagel Dorothea
Published: January 2020

The purpose of this paper is to survey the usefulness of cluster analysis in the special case of diagnoses. This complex topic is restricted, however, to the application on laboratory characteristics, separately or in connection with clinical data. The article is subdivided into three parts: (a) the fields of a possible use of cluster analysis, detection of diseases or subgroups of diseases, and data reduction by detection of structures; (b) a brief mathematical description of hierarchical and partitioning classification techniques (as a crucial point, the problems associated with these methods are discussed); (c) a critical review of 24 publications of the past 10 years concerning cluster analysis and diagnoses.
Increased plasma amylase in the family of a patient with 3-hydroxy-3-methylglutaryl-coenzyme A lyase deficiency
Authors: Plöchl E , Colombo J P , Wermuth B , Gibson K M
Published: January 2020

A patient with 3-hydroxy-3-methylglutaryl-CoA lyase (HMG-CoA lyase, EC 4.1.3.4) deficiency presented consistently above-normal values of plasma amylase (EC 3.2.1.1). Activities measured were in the lower normal range in family members not proven heterozygotes and in the upper normal range in the proven heterozygotes. Heterozygosity was proven by intermediate HMG-CoA lyase activities determined in cultured fibroblasts and in lymphocytes in the parents and the paternal grandmother. Because all of the family members had diseases of the pancreas, colon, and liver, we question whether the heterozygote state contributes to the impaired function of these organs. Our findings of significantly increased amylase activities in the heterozygotes and the patient, in comparison with the other family members, support this hypothesis.
Laboratory Monitoring of Nutritional Status in Burn Patients
Authors: Rettmer R L , Williamson J C , Labbé R F , Heimbach D M
Published: January 2020

Most nutrition laboratory testing relies on serum concentrations of ingested nutrients, their coenzymes, proteins, or lipids. Alternatively, functional tests measure a specific physiological process or biochemical reaction. We compared these two approaches to nutritional assessment in intensive-care burn patients, in whom the serum concentrations of transthyretin (prealbumin), albumin, transferrin, carotene, retinol, ascorbic acid, copper, cholesterol, iron, and calcium were all below established reference ranges. In contrast, serum triglyceride concentrations were often above the reference range. Functional tests for thiamin, riboflavin, pyridoxine, and iron (by zinc protoporphyrin/heme ratio) in these patients all showed normal values. Dietary intake, weight trends, and nitrogen balances all indicated that these patients' estimated caloric and protein needs had been met. These findings suggest that static measurements of serum concentrations may be unreliable indicators of nutritional status in burn patients.
Determination of Serum Glucose by Isotope Dilution Mass Spectrometry: Candidate Definitive Method
Authors: Magni F , Paroni R , Bonini P A , Kienle M G
Published: January 2020

We report a rather simple method to determine glucose concentration in serum, using isotope dilution mass spectrometry and [13C6]glucose as internal standard. The procedure involves a single step of sample purification and the conversion of the analyte into its aldononitrile pentaacetate. The between-day and within-day contribution to total variance for a single measurement was determined by assaying Standard Reference Material (SRM) 909 serum. The method was then applied to measurement of glucose concentration in three lyophilized sera: SRM 909 and two other commercially available sera. In the two studies, the concentration of SRM 909 serum was found to be 0.8% above and 0.3% below the reported value (6.25 mmol/L), respectively; the overall coefficient of variation for determinations in all sera ranged from 0.37% to 0.56%. The precision and the accuracy of the method satisfy the requirements for a Definitive Method.
Endogenous Digitalis-Like Factors in Human Milk
Authors: Clerico A , Paci A , Del Chicca M G , Biver P , Giampietro O
Published: January 2020

We measured the concentration of endogenous digitalis-like factors (EDLFs) in milk or colostrum of women during nursing on different days after delivery. EDLF concentrations were assayed by a solid-phase RIA involving antidigoxin antibodies and by a radioreceptor assay (RRA) involving human placenta Na+/K(+)-ATPase. The mean (SD) EDLF concentrations as measured by RIA were 35.6 (19.4) ng of digoxin equivalents per liter in milk samples (n = 37) and 61.3 (12.5) ng/L in colostrum samples (n = 5); the mean EDLF concentration as measured by RRA in milk samples (n = 11) was 573 (717) ng/L (range 0-2098). EDLF concentration in milk is greater than circulating concentrations in healthy adults but is comparable with serum concentration in the third trimester of pregnancy. In milk and serum samples (n = 8) collected at the same time, heating and (or) extracting with Sep-Pak C18 cartridges before the RIA produced significantly different EDLF values from those in untreated serum (P less than 0.001) and milk (P = 0.035). EDLF in milk appeared to be not bound or weakly bound to milk protein, as indicated by the fact that boiling did not increase the digoxin-like immunoreactivity.
Determining Zinc Coproporphyrin in Maternal Plasma--a New Method for Diagnosing Amniotic Fluid Embolism
Authors: Kanayama N , Yamazaki T , Naruse H , Sumimoto K , Horiuchi K , Terao T
Published: January 2020

We measured the concentration of zinc coproporphyrin I (ZnCP-I), a characteristic component of meconium, in maternal plasma by fluorometry after HPLC. We obtained plasma samples from 89 women: 35 at weeks 10-40 of normal pregnancy, 41 shortly after normal delivery, 4 from patients with amniotic fluid embolism (AFE), and 9 from non-AFE patients with intra- or postpartum shock caused by genital bleeding. The plasma ZnCP-I concentration was 97 (SD 83, range 38-240) nmol/L in the AFE patients, 11 (SD 9.2) nmol/L in the non-AFE patients, 12 (SD 7.9) nmol/L during normal pregnancy, and 26 (SD 10) nmol/L shortly after normal delivery. We suggest that measuring ZnCP-I in maternal plasma by fluorometry on HPLC is a rapid, noninvasive, and sensitive method for diagnosing AFE and propose 35 nmol/L as the cutoff value for the ZnCP-I concentration in maternal plasma for the diagnosis of AFE.
Micral-Test Strips Evaluated for Screening for Albuminuria
Authors: Marshall S M , Shearing P A , Alberti K G
Published: January 2020

We have evaluated Micral-Test, an immunochemical strip test specific for albumin, as a screening tool for slight ("micro") albuminuria. First morning urine samples containing albumin concentrations (by radioimmunoassay) of 0.4-440 mg/L were collected from 112 diabetic patients. The Micral-Test result for each sample was assessed by one observer. All 34 samples having albumin concentrations greater than or equal to 20 mg/L and 71 of 78 samples less than 20 mg/L were correctly identified, giving 100% sensitivity and 91% specificity. Six samples were measured 10 times by one observer: three samples were read consistently; one, albumin concentration 86 mg/L, was read as 50 and 100 mg/L; and two, albumin concentrations 32 and 38 mg/L, were read as 20 and 50 mg/L, respectively. Contact with urine for 2 s rather than the recommended 5 s resulted in an underestimation of the albumin concentration in 13 of 35 samples (Z = -3.18, P = 0.001), as did taking readings earlier than the recommended 5 min (Z = -3.92, P less than 0.001). Six observers independently performed Micral-Test measurements on 10 samples. Eight samples were correctly classified as greater than or equal to 20 or less than 20 mg/L by all observers, but two (albumin concentrations 25 and 18 mg/L) were misclassified by at least one observer. The Micral-Test is a sensitive and specific screening tool, but is semiquantitative and critically time dependent.
Benzocaine-Adultered Street Cocaine in Association with Methemoglobinemia
Authors: McKinney C D , Postiglione K F , Herold D A
Published: January 2020

A 27-year-old man with a history of cocaine abuse ingested a large quantity of street cocaine in an apparent suicide attempt. Shortly thereafter, he developed tonic-clonic seizures and became cyanotic. An arterial blood gas sample, collected in the emergency department, appeared chocolate-brown and showed pO2 279 mmHg, pCO2 53 mmHg, and pH 7.15. Hemoglobin spectral analysis revealed significant methemoglobinemia (37%). Subsequent gas-chromatographic and mass-spectral analysis of urine confirmed the presence of cocaine (106 mg/L), benzoylecgonine (94 mg/L), and other metabolites. Further testing revealed the presence of benzocaine, a compound known to produce methemoglobinemia. A powder submitted as the "cutting" substance was shown to be benzocaine. When confronted with a possible cocaine overdose (particularly by ingestion), the physician should consider the possible clinical effects of adulterants, especially local anesthetics such as benzocaine.
Simple and Sensitive Determination of Urea in Serum and Urine
Authors: Orsonneau J L , Massoubre C , Cabanes M , Lustenberger P
Published: January 2020

In this method for serum and urinary urea determination, the same reagent is used without predilution of urine samples. The method is based on the pH increase resulting from the ammonia released by urease hydrolysis of urea. o-Cresolphthalein complexone is used to monitor the pH change colorimetrically. Urea concentration and absorbance at 570 nm are linearly related for concentrations as great as 600 mmol/L for urine samples and 100 mmol/L for serum. There are no clinically significant interferences from physiological substances or drugs, and precision and accuracy are excellent (CV approximately 2%, except at very low concentrations in serum; analytical recovery was 99% in urine, 100% in serum). Results by this method (y) and by the Astra method (x) for urine correlated well (y = 0.991x - 2.87, Sy/x = 9.21, r = 0.994), as did the results by this method and by the total enzymatic method (x') for serum (y = 1.002x' + 0.192, Sy/x' = 0.598, r = 0.997). This method is applicable to automated as well as manual instruments, and one-reagent or two-reagent formats can be used.
Profile of Renal Permselectivity by Simultaneous Enzyme-Linked Immunosorbent Assay of Albumin, Transferrin, IgG, and α1-Microglobulin with a New Microplate Reader
Authors: Magnotti R A , Eberly J P , Khoury P R , Daniels S R , Drozda D J , Turner A M , Dolan L M
Published: January 2020

A competitive enzyme-linked immunosorbent assay (ELISA) is described for determining a renal permselectivity profile involving the urinary proteins albumin, transferrin, IgG, and alpha 1-microglobulin (alpha 1-m). The ELISA reader uses a computer-controlled array of multiplexed light-emitting diode (LED)-photodiode pairs for rapid measurements of absorbance on microplates. A 3,3'-dimethylnaphthidine reagent adapts the 3,5,3',5'-tetramethylbenzidine chromophore to monochromatic LED emission at 550 nm. We applied this ELISA to the determination of renal permselectivity in healthy children and young adults and in children with insulin-dependent diabetes mellitus. The geometric means (and SD) of protein excretion rates in a group of 85 normal subjects were as follows: albumin, 3.5 micrograms/min (1.83); transferrin, 173 ng/min (2.76); IgG, 1.11 micrograms/min (2.22), and alpha 1-m, 0.98 microgram/min (2.36).
Estimation of Reference Ranges: How Many Subjects are Needed?
Authors: Lott J A , Mitchell L C , Moeschberger M L , Sutherland D E
Published: January 2020

We measured Na, K, Cl, glucose, hemoglobin, erythrocytes, and hematocrit in the serum or blood from approximately 800 male and 200 female second-year medical students in an effort to define the size of an acceptable reference population. Using the data from the men and Monte Carlo simulations of 10-400 samples, each carried out 5000 times, we found that for most of the above tests, approximately 200 people are required for stable lower (2.5%) and upper (97.5%) reference limits to be obtained. This agrees with the 198 subjects required by strictly statistical criteria to define the same limits with a 99% confidence level.
Cholesterol and Apolipoprotein D in Gross Cystic Disease of the Breast
Authors: Sánchez L M , Díez-Itza I , Vizoso F , López-Otín C
Published: January 2020

Cholesterol and apolipoprotein D (apo D) concentrations were measured in cyst fluids and sera from 66 women with gross cystic disease of the breast. Intracystic cholesterol concentrations are about twofold greater than those found in serum, whereas apo D, the major cyst fluid protein, is present in concentrations as much as 1000-fold greater than found in serum. We classified the cyst fluids into two groups by K+/Na+ ratio and albumin concentration and measured intracystic cholesterol and apo D concentrations in each group. Type I cysts had an average content of cholesterol (12.7 mmol/L) moderately lower than that in Type II cysts (17.6 mmol/L). By contrast, the average concentration of apo D in Type I cyst fluids (15.1 g/L) was slightly higher than that in Type II cysts (13.7 g/L). The absence of a correlation between cholesterol concentration and apo D concentration and the results from previous binding analysis suggest that this protein is not involved in the accumulation of cholesterol in breast-cyst fluid.
Monoclonal Immunoradiometric Assay and Polyclonal Radioimmunoassay Compared for Measuring Neuron-Specific Enolase in Patients with Lung Cancer
Authors: Body J J , Paesmans M , Sculier J P , Dabouis G , Bureau G , Libert P , Berchier M C , Raymakers N , Klastersky J
Published: January 2020

Neuron-specific enolase (NSE) is the most sensitive and specific tumor marker for small-cell lung cancer (SCLC). We evaluated a new monoclonal IRMA (Sangtec) for NSE and compared it with a polyclonal RIA (Pharmacia) in patients with SCLC or other lung cancers (NSCLC). We measured NSE concentrations in 100 healthy subjects (NI group), 100 patients with benign pulmonary diseases (BPD group), and 194 patients with advanced lung cancer (97 SCLC and 97 NSCLC). Intra- and interassay CVs were less than 7% for both assays, and dose-dilution curves paralleled their respective standard curves. Values measured by both assays were highly correlated in all groups. NSE concentrations were significantly (P less than 0.001) lower by IRMA than by RIA in NI and BPD groups. The upper 95th percentile values for NSE in the NI group were 11.7 micrograms/L in the RIA and 9.2 micrograms/L in the IRMA. In NSCLC, the values were significantly (P less than 0.05) lower by IRMA but the percentage of subjects with increased values was higher (vs the NI group, 31% for RIA and 44% for IRMA, P less than 0.005). Diagnostic sensitivity for SCLC was improved with IRMA: 83% of values with RIA and 93% with IRMA were increased above the NI group values (P less than 0.005); the corresponding values for SCLC vs BPD were 81% and 89% (P less than 0.05). NSE values measured in 39 patients with SCLC after chemotherapy were more often increased and were significantly higher with the IRMA than with the RIA (P less than 0.005).
Excess Osmolal Gap in Diabetic Ketoacidosis Explained
Authors: Davidson D F
Published: January 2020

Even in the absence of detectable ethanol or visible lipemia, a large plasma osmolal gap is the usual finding in cases of diabetic ketoacidosis. This gap decreases to an insignificant value within 20 h of treatment. Detailed biochemical analysis of six cases showed that, on average, the gap could be almost wholly accounted for by an increase in acetone, a decrease in the plasma water fraction, and smaller increments in amino acids and glycerol. Calculated plasma osmolality can seriously underestimate the true value in diabetic ketoacidosis, and so some previously observed anomalies may be explained.
Characterization of Cryoglobulins by Immunoblotting
Authors: Musset L , Diemert M C , Taibi F , Thi Huong Du L , Cacoub P , Leger J M , Boissy G , Gaillard O , Galli J
Published: January 2020

We adapted an immunoblotting technique for the immunochemical characterization of cryoglobulins. We first compared the results from 157 samples with results obtained by immunofixation and immunoelectrophoresis and then with clinical observations in 125 cases. Full identification was possible in 98% of the cases by immunoblotting, in 54% by immunofixation, and in 28% by immunoelectrophoresis. Novel microheterogeneity aspects were observed by immunoblotting and immunofixation in 13% and 6% of the cases studied, respectively. Our results confirm the frequency of autoimmune and infectious diseases associated with mixed cryoglobulinemias, whereas a lymphoproliferative syndrome was observed only in cases of cryoglobulinemia with a monoclonal constituent or a microheterogeneity aspect, which was not always revealed by immunofixation or immunoelectrophoresis. The pathophysiological importance and classification of microheterogeneity aspects is not clear. These observations justify using a sensitive and specific method for identifying cryoglobulins, even when present at low concentrations, and call for long-term studies of these patients.