bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of salivary proteins on binding curves of three radioimmunoassay kits: Amerlex-M progesterone, Amerlex cortisol, and Biodata testosterone.
Authors: Fulton A , Chan S , Coleman G
Published: January 2020

Radioimmunoassay (RIA) is generally used to measure certain salivary hormones because of its high sensitivity. For speed and simplicity, it has been used in the form of "direct" assays, i.e., without first extracting the analyte from its matrix. Investigating the effect of the principal salivary proteins on the binding behavior of three commercial RIA kits, we found that the Amerlex-M [125I]progesterone binding was greatly reduced when alpha-amylase and mucins were added to the binding medium, whereas IgA and IgG were less effective. The Serono Biodata [125I]testosterone binding was unaffected by proteins, while the Amerlex [125I]cortisol binding was decreased by alpha-amylase and mucins. The protein influence was largely eliminated when an extraction step was incorporated. Thus, direct RIA of saliva may be subject to matrix effects, to extents that vary with the kit and that may adversely affect the quality of the assay results.
Hereditary complete deficiency of lactate dehydrogenase H-subunit.
Authors: Joukyuu R , Mizuno S , Amakawa T , Tsukada T , Nishina T , Kitamura M
Published: January 2020

We report the second known case of a patient, a 45-year-old Japanese woman, with hereditary complete deficiency of lactate dehydrogenase (LDH; EC 1.1.1.27) H-subunit. Total LDH activity in her serum was abnormally low (35 U/L, normal reference interval 195-360). LDH activity in her erythrocytes was also low, but all the other glycolytic enzyme activities in her erythrocytes were within normal limits. Electrophoresis of her serum, erythrocytes, lymphocytes, thrombocytes, and saliva showed only one band, the LDH M4 isoenzyme. LDH activities in her saliva and lymphocytes exceeded the reference interval. Her erythrocytes contained fructose 1,6-diphosphate 26 mumol/L (normal range 4-13), dihydroxyacetone phosphate 75 mumol/L (normal range 8-22), glyceraldehyde 3-phosphate 57 mumol/L (normal range 4-14), and pyruvate 45 mumol/L (normal range 31-63). The family study of three generations showed that this deficiency was inherited in an autosomal recessive mode.
Properties of the beta 1- and beta 2-adrenergic receptor subtypes revealed by molecular cloning.
Authors: Frielle T , Caron M G , Lefkowitz R J
Published: January 2020

The beta 1- and beta 2-adrenergic receptor subtypes are biochemically and functionally similar, because both receptors mediate the catecholamine-dependent activation of adenylate cyclase through the GTP-binding protein, Gs. Pharmacologically, the two receptors can be distinguished on the basis of their relative affinities for the agonists epinephrine and norepinephrine as well as their affinities for several selective antagonists. The primary structures of the human beta 1- and beta 2-adrenergic receptors have recently been deduced from the cloning of their genes and (or) cDNAs, revealing high sequence homology and a membrane topography of seven putative transmembrane regions similar to that of rhodopsin. Chimeric beta 1/beta 2-adrenergic receptor cDNAs have been constructed by site-directed mutagenesis and the chimeric RNA transcripts expressed in Xenopus laevis oocytes. The pharmacological properties of the expressed chimeric receptor proteins were assessed by radioligand binding utilizing subtype-selective agonists and antagonists. Apparently, several of the putative transmembrane regions contribute significantly to the determination of subtype selectivity, presumably by formation of a ligand-binding pocket, with determinants for agonist and antagonist binding being distinguishable.
Cell and substrate adhesion molecules in embryonic and neural development.
Authors: Crossin K L
Published: January 2020

Recent studies of cell adhesion molecules and substrate adhesion molecules suggest that many key events related to embryonic and neural pattern formation depend upon cell surface modulation. I briefly review here the evidence supporting this conclusion.
Reliability of the determination of whole-blood oxygen affinity by means of blood-gas analyzers and multi-wavelength oximeters.
Authors: Kwant G , Oeseburg B , Zijistra W G
Published: January 2020

Determination of the oxygen affinity of human whole blood with the help of blood-gas analyzers and multi-wavelength oximeters is compared with an accurate method for recording hemoglobin oxygen dissociation curves (Clin Chem 1982;28:1287-92). P50 (oxygen tension at half saturation; So2 = 50%) and Hill's n (delta log [So2/(1-So2)]/delta log Po2) were determined in blood of 24 healthy donors. Three slightly different procedures were used for determination of P50 on the basis of Po2, pH, Pco2, and So2 measured with each of four different blood-gas analyzer/oximeter combinations. These methods were not able to discriminate between high and low values for P50 within the normal reference interval, but never failed to detect the high oxygen affinity of blood stored for 12 days, reflected in low values of P50. The methods thus proved suitable for detecting clinically significant deviations in oxygen affinity such as occur in patients with, e.g., abnormal hemoglobins, anemias, or carbon monoxide poisoning. Determination of Hill's n by these methods did not produce useful results.
Immunoassay for estrogen receptor does not detect inactivated receptor.
Authors: Molnar L A , Dhirani N , Tran K , McBlain W A
Published: January 2020

Accurate quantification of estrogen receptor (ER) is essential for optimal clinical characterization of individual cases of breast cancer. If breast tumors are mishandled, the relatively labile ER protein may lose its steroid-binding capacity (become inactivated) and not be measurable by the routine steroid-binding assay. We tested whether the commercial enzyme immunoassay of Abbott Laboratories could quantify inactivated ER. Samples of powdered breast tumors from humans were exposed to various temperature and homogenization conditions known to inactivate ER, and any remaining ER was quantified by both the immunoassay and the steroid-binding assay. For all inactivation conditions tested, the two assay methods detected the same proportions of remaining ER. We conclude that the inactivation reaction for ER also alters one or both of the antigenic site(s) necessary for the immunoassay. Hence, for breast tumors mishandled to the extent of inactivating ER, the immunoassay offers no advantage over the more conventional steroid-binding assay for quantifying any remaining ER.
An automated procedure for measuring biotinidase activity in serum.
Authors: Weissbecker K A , Gruemer H D , Heard G S , Miller W G , Nance W E , Wolf B
Published: January 2020

In this automated procedure for quantifying biotinidase activity in human serum, a manual colorimetric method that measures conversion of the enzyme's artificial substrate N-biotinyl p-aminobenzoate was modified for use with a Technicon AutoAnalyzer II. The intra-run replicate precision (CV) was 2.1% and the day-to-day CV was 4.6% for quality-control sera. Results were linearly related to biotinidase activity in serum over the complete range of clinically relevant values, 0.2 to 11.0 U/L. Moreover, results of the automated assay were not significantly different from those of the manual assay. Because the automated procedure is faster and more precise, we recommend it for population-based studies and some screening studies.
A new commercial method for the enzymatic determination of creatinine in serum and urine evaluated: comparison with a kinetic Jaffé method and isotope dilution-mass spectrometry.
Authors: Lindbäck B , Bergman A
Published: January 2020

We evaluated a new, simple, enzymatic kinetic method from Wako Chemicals GmbH in comparison with a kinetic Jaffé method by using isotope dilution-mass spectrometry (ID-MS) as a reference method. An ID-MS-calibrated serum standard was used. Both the enzymatic and the Jaffé method correlated well with ID-MS, except for sera with high concentrations of bilirubin. Ethyl acetoacetate, acetone, and glucose in serum interfered somewhat with the Jaffé method but not with the enzymatic method. We conclude that the present enzymatic method has merit as compared with a Jaffé method for routine work, but is more expensive.
Partial characterization of an abnormal lactate dehydrogenase isoenzyme, LDH-1ex, in serum from a patient with hepatocellular carcinoma.
Authors: Kalpaxis D L , Giannoulaki E E
Published: January 2020

Serum from a patient with hepatocellular carcinoma contained an abnormal isoenzyme of lactate dehydrogenase (LDH; EC 1.1.1.27), LDH-1ex, that on electrophoresis on 10-g/L agarose gel migrated anodally to the LDH-1 band. This isoenzyme was partly purified by ultrafiltration and preparative electrophoresis. Gel chromatography and sodium dodecyl sulfate/polyacrylamide gel electrophoresis studies of the resulting LDH-1ex preparation suggested that this isoenzyme is probably a tetramer made up of four single polypeptide chains (monomers), each having a molecular mass of about 32,000 Da. LDH-1ex was heat stable and reacted more readily with 2-hydroxybutyrate than did the slower migrating LDH-4 and LDH-5 isoenzymes. LDH-1ex showed no activity when lactate was omitted from the substrate solution or replaced by ethanol.
Activities of dipeptidyl peptidase II and dipeptidyl peptidase IV in synovial fluid from patients with rheumatoid arthritis and osteoarthritis.
Authors: Gotoh H , Hagihara M , Nagatsu T , Iwata H , Miura T
Published: January 2020

We examined the activities of peptidases in synovial fluid from patients with rheumatoid arthritis (RA) and osteoarthritis (OA). Dipeptidyl peptidase IV (DPP IV) activity was lower in synovial fluid from patients with RA, in contrast to the increase of DPP II activity in synovial fluid, as compared with OA. The DPP II/DPP IV ratio for synovial fluid was significantly higher in patients with RA than in patients with OA. A significant correlation was observed between the DPP II/DPP IV ratio for synovial fluid from patients with RA and the amount of C-reactive protein reaction. These results may be useful in the diagnosis of joint effusion of unknown origin.
Somatotropin as measured by a two-site time-resolved immunofluorometric assay.
Authors: Strasburger C , Barnard G , Toldo L , Zarmi B , Zadik Z , Kowarski A , Kohen F
Published: January 2020

To date, many of the current criteria for diagnosis of somatotropin (growth hormone, GH) deficiency have been based upon measurement of this hormone by competitive radioimmunoassay (RIA) with use of polyclonal antibodies. In recent years, however, the development of hybridoma technology has led to the generation of various monoclonal antibodies (Mabs) to GH with different affinities and epitope specificities. Subsequently, these reagents have been used in the development of noncompetitive two-site immunometric assays (e.g., immunoradiometric assay; IRMA). In general, the values obtained for serum GH by IRMA have been lower than those obtained by RIA, because of the epitope-specificity profile of the Mabs in the IRMA. Attempting to obtain GH values numerically similar to those by RIA, we used a combination of Mabs to GH in developing and evaluating a two-site time-resolved immunofluorometric assay (IFMA) based on the streptavidin-biotin interaction. Fluorescence is proportional to concentration of analyte and is linearly related to concentration over the range 0.3 to 40 micrograms/L. The assay was satisfactory with respect to sensitivity, accuracy, and precision (CV less than 10% over the entire working range). In addition, the concentration of GH was determined by the IFMA and a competitive RIA in serum obtained from GH deficient and acromegalic patients. The pairing of antibodies in the IFMA gave numerical values that agreed well with those by RIA (r = 0.97; n = 100).
Serum mitochondrial aspartate aminotransferase activity: not useful as a marker of excessive alcohol consumption in an unselected population.
Authors: Schiele F , Artur Y , Varasteh A , Wellman M , Siest G
Published: January 2020

Using an immunochemical method, we measured the activity of the mitochondrial isoenzyme (mAST) of aspartate amino-transferase (EC 2.6.1.1, AST) in the serum of 687 subjects attending the Centre for Preventive Medicine for a health examination. The distributions of the activities were asymmetrical, with mean values of 1.8 U/L (SD 2.0) for men and 1.4 U/L (SD 1.6) for women. The average ratio of mitochondrial to total AST activity was 0.051 (range 0-0.42). In this unselected population we found no change in the mitochondrial activity or in the mitochondrial-to-total ratio attributable to alcohol consumption, even in subjects who consumed more than 88 g per day. Of 35 men with an alcohol consumption greater than 88 g/d, 19 had a serum gamma-glutamyltransferase activity of greater than or equal to 60 U/L, 17 had glutamate dehydrogenase values greater than or equal to 5 U/L, and only nine had an mAST activity greater than or equal to 3 U/L (values corresponding to the 80th percentiles of the total population). We conclude that the test is not particularly useful as a screening procedure in an unselected population under present-day conditions of measurement.
Influence of end-stage renal failure on concentrations of free apolipoprotein A-1 in serum.
Authors: Duval F , Frommherz K , Atger V , Drüeke T , Lacour B
Published: January 2020

We determined, immunoturbidimetrically, the concentrations of apolipoprotein A-I (apo A-I) (a) in total serum, (b) in total lipoproteins, and (c) in the fraction of d greater than 1.21 kg/L (free apo A-I) in 31 uremic patients (16 on hemodialysis, HD, and 15 with end-stage renal failure, ESRF) and 14 control subjects. The concentration of free apo A-I in serum was significantly increased in both groups of uremic patients (0.27 +/- 0.07 g/L for HD and 0.22 +/- 0.08 g/L for ESRF, mean +/- SD), in comparison with the control group (0.14 +/- 0.04 g/L). The ratio of total apo A-I to high-density-lipoprotein cholesterol was significantly increased in sera from both uremic groups, whereas the apo A-I/HDL-chol ratio of total lipoproteins was similar in all three groups. No apo A-I could be detected in five ultrafiltrates of plasma, even after 100-fold concentration. Analysis of apo A-I isoforms by isoelectrofocusing revealed a significant relative increase in apo A-I3 and a decrease in apo A-I5 isoform in ESRF patients, but not in HD patients. Finally, we found no relationship between serum TG and free apo A-I concentration.
HPLC measurement of phenylalanine by direct injection of plasma onto an internal-surface reversed-phase silica support.
Authors: Atherton N D
Published: January 2020

Direct-injection analysis for phenylalanine in plasma by isocratic "high-performance" liquid chromatography (HPLC) is described. For chromatographic separation I use an internal-surface reversed-phase silica support. The method involves ultraviolet detection, requires as little as 10 microL of plasma (routinely 20 microL), and supplies results in less than 7.5 min from injection. The intra-batch coefficient of variation for phenylalanine in human plasma samples (197-1104 mumol/L) ranged between 3.0% and 6.0% and the range of linearity was 10-1250 mumol/L for a 20-microL sample injection loop. The upper limit can readily be extended by using a smaller volume loop. Phenylalanine results from direct plasma injection (y) correlated well with those of a protein precipitation/HPLC technique (x): y = 0.931x + 46.4 (r = 0.951, n = 41, range 78-1344 mumol/L).
Specific routine determination of 3'-azido-3'-deoxythymidine (AZT) in plasma by partly automated liquid chromatography.
Authors: Kupferschmidt R , Schmid R W
Published: January 2020

A simple isocratic HPLC method for determining azidothymidine (AZT) in serum and plasma of patients has been developed. The novel, specific, two-step, solid-phase extraction approach used for sample preparation gives a nearly quantitative recovery (95.3%) of AZT from the blood plasma matrix and requires only minimal handling of infectious clinical samples. Automatic "on-line" injection is achieved with an AASP system by switching a small cartridge, which retains the extracted analyte, into the HPLC stream. The overall HPLC procedure shows satisfying reproducibility with low standard deviation (CV: 2.1%). Because of the low detection limit (about 10 ng) and the possibility of concentrating AZT quantitatively in as much as 5 mL of plasma or serum, the method can be used in routine monitoring of AZT as well as in pharmacokinetic studies. Nevertheless, before establishing therapeutic drug monitoring for AZT, it still must be determined at what time after the last AZT dose blood specimens should be drawn for correct therapeutic interpretation of the concentration of AZT measured in blood.
Prostatic inhibin-like peptide quantified in urine of prostatic cancer patients by enzyme-linked immunosorbent assay.
Authors: Teni T R , Bandivdekar A H , Sheth A R , Sheth N A
Published: January 2020

This is a highly specific enzyme-linked immunosorbent assay (ELISA) for measuring prostatic inhibin-like peptide (PIP) in urine, in which we use penicillinase (EC 3.5.2.6) conjugated with PIP and, as solid phase, a polystyrene microtiter plate. We used this ELISA to measure PIP in 24-h urine specimens from men with prostatic cancer (PCa) and from age-matched controls. For prostatic cancer patients the mean +/- SEM urinary PIP of 36.1 +/- 5 micrograms/24 h was significantly (P less than 0.001) lower than the mean of 127.1 +/- 9 micrograms/24 h for the age-matched controls. PIP values for 30 samples measured by both ELISA and RIA correlated well (r = 0.985). We could detect as little as 1.56 ng of PIP in a sample. Analytical recovery of added PIP ranged from 91% to 104%. Mean CVs were 8.9% within-assay and 12.7% between-assay. We believe that this ELISA will be useful in assessing the status of PIP in men with normal and diseased prostates and in examining the function of the hypothalamus-pituitary-prostate axis.
Determination of branched-chain amino acids and tyrosine in serum of patients with various hepatic diseases, and its clinical usefulness.
Authors: Azuma Y , Maekawa M , Kuwabara Y , Nakajima T , Taniguchi K , Kanno T
Published: January 2020

We developed an automated enzymatic method for determination of the branched-chain amino acids (BCAAs; valine, isoleucine, leucine) and tyrosine in serum, and applied it to the clinical evaluation of patients with various hepatic diseases. Analytically, the test results were acceptably precise and reproducible, and correlated well with results obtained with an amino acid analyzer. Clinically, we found that a decrease in BCAAs, an increase in tyrosine, and the BCAAs/tyrosine ratio in serum paralleled the severity of hepatic parenchymal damage. We conclude that this enzymatic determination of BCAAs and tyrosine is simple and convenient enough for routine clinical laboratory use, and that the ratio of BCAAs/tyrosine obtained may be a good indicator of the severity of hepatic disorders.
Radioimmunoassay of osteocalcin with polyclonal and monoclonal antibodies.
Authors: Power M J , Gosling J P , Fottrell P F
Published: January 2020

We have developed and thoroughly validated radioimmunoassays (RIAs) for osteocalcin in plasma and serum, demonstrating independence of volume and determining the recovery of added standard and within- and between-assay precision. Antibodies were raised in rabbits and mice by immunization with either osteocalcin adsorbed to polyvinylpyrrolidone or osteocalcin conjugated to bovine serum albumin. In both species, use of the conjugated osteocalcin was more efficacious. Correlation of the results obtained when groups of plasma or serum samples were analyzed by RIAs with different antibodies (two polyclonal and one monoclonal) indicated that the normal range observed for osteocalcin concentrations in serum depends on the antibody used. These findings may account for the wide range of "normal" osteocalcin values reported by different groups.
A rapid micro-scale procedure for determination of the total lipid profile.
Authors: Wu L L , Warnick G R , Wu J T , Williams R R , Lalouel J M
Published: January 2020

We describe a one-day micro-scale procedure for determining the total lipid profile. Only 0.55 mL of plasma is needed for complete quantification of total cholesterol (TC), triglyceride (TG), and all lipoproteins. After precipitation with dextran sulfate and magnesium, the high-density lipoprotein (HDL) fraction was separated by centrifugation in an Eppendorf microcentrifuge. Very-low-density lipoprotein (VLDL) was separated from low-density lipoprotein (LDL) plus HDL in a Beckman TL 100 ultracentrifuge. TC, TG, and cholesterol in different lipoprotein fractions were measured enzymatically in a Baker "Encore II" automated analyzer. CVs, both within-day and day-to-day, were less than 3% for TG and TC, and less than 5% for HDL-C determinations. CVs for LDL-C and VLDL-C were less than 7.5% and 15%, respectively. Results by our micromethods (n = 66) agreed well with those by the conventional methods used at the Northwest Lipid Research Center, which are standardized against the Reference Methods of the Centers for Disease Control. Coefficients of correlation between the two methods were 0.98 for TC, 1.0 for TG, 0.98 for HDL-C, 0.94 for LDL-C, and 0.96 for VLDL-C. Results of electrophoresis on agarose gel and radioactivity-recovery studies indicate that our micro-centrifugation and slicing procedures result in clean separation of VLDL from other lipoproteins.
Loss of porphyrins from solution during analysis: effect of sample pH and matrix on porphyrin quantification in urine by "high-performance" liquid chromatography.
Authors: Perkins S L , Johnson P M
Published: January 2020

We report the effect of sample matrix and pH on quantification of porphyrins by HPLC with fluorimetric detection. For aqueous solutions of pH less than 2.5, HPLC peak heights of the porphyrins increased with decreasing pH, reaching a plateau at pH less than 1.0. This loss of porphyrins from solutions with pH greater than 1.0 appeared to be due to a combination of microprecipitation and aggregation effects. No such "pH effect" was observed for urine samples supplemented with mixed-porphyrin standards. Addition of trace amounts of albumin to aqueous solutions also decreased these pH-related losses. These findings suggest a porphyrin-protein interaction that prevents microprecipitation and aggregation processes. We conclude that standard solutions of porphyrins for HPLC analysis should be prepared in a urine matrix. If aqueous solutions are used, then the pH must be adjusted to less than 1.0. Urine samples from normal individuals require only adjustment of pH to less than 2 before analysis; however, porphyric urines requiring dilution should be prepared with porphyrin-free urine diluent.
Two whole-blood multi-analyte analyzers evaluated.
Authors: Fleisher M , Gladstone M , Crystal D , Schwartz M K
Published: January 2020

We evaluated two multi-analyte analyzers, the NOVA Stat Profile 1 (SP 1) and the NOVA Stat Profile 5 (SP 5). The SP 1 measures pH, pCO2, pO2, sodium, potassium, ionized calcium, and hematocrit in heparinized whole blood; the SP 5 determines all these analytes, plus chloride (not evaluated because it was unavailable at the time this study was initiated) and glucose. Interassay precision for pH, pCO2, pO2, sodium, potassium, ionized calcium, and hematocrit (all on the SP 1) and glucose (on the SP 5) was excellent, the respective CVs (%) being: less than 0.006, 2.1, less than or equal to 3.0, less than or equal to 0.5, less than or equal to 1.7, less than or equal to 2.1, less than or equal to 3.9, and less than or equal to 1.2. Correlation with results obtained with Corning's Model 178, NOVA Biomedical's Model 6, Beckman's Astra 8, and Roche's Cobas-Bio was also excellent (r greater than 0.975 for all analytes, r for hematocrit 0.865). Temperature-stability studies on whole-blood specimens maintained at 1 degree C indicated that results for all measured analytes were essentially uncharged for at least 2 h, except for potassium, which increased by 13% in 2 h. At 22 degrees C, values for pH, pCO2, pO2, and glucose changed significantly within 2 h. Advantages of the NOVA Stat Profile series include decreased specimen requirements (250 microL), analysis time (72-90 s), turnaround time (about 4.5 min), and overall cost of operation.
Functional and immunological concentrations of alpha 2-macroglobulin compared in plasma of subjects without alpha 1-protease inhibitor deficiency.
Authors: Gaillard M C , von Arb M , da Costa A , Kilroe-Smith T A
Published: January 2020

Alpha 2-Macroglobulin concentrations as measured by a functional assay and by laser nephelometry with antibody precipitation were compared for 67 outpatients at a respiratory clinic and 30 blood-donor control subjects with no respiratory disease. In both groups, concentrations of alpha 1-protease inhibitor in plasma, as measured by laser nephelometry, were within the normal reference interval. A reasonable correlation (r = 0.70) was obtained between results of functional assays and of laser nephelometry for the control individuals. For patients with pulmonary disease the correlation between the two assay methods was poorer (r = 0.53). We conclude that the functional assay of plasma is the better method for detecting abnormal concentrations of alpha 2-macroglobulin in patients with lung diseases.
Rhabdomyolysis in a case of free-base cocaine ("crack") overdose.
Authors: Jandreski M A , Bermes E W , Leischner R , Kahn S E
Published: January 2020

This is the case of a 27-year-old black man who was admitted to Loyola University Medical Center after a one-time experience of smoking free-base ("crack") cocaine. Clinical manifestations of the resulting cocaine intoxication were rhabdomyolysis, acute renal failure, and transient liver failure. This patient came to our attention because of the striking alterations in his blood-chemistry values, which indicated acute tissue damage, and his remarkable recovery within 96 h. We discuss the dramatic changes in the laboratory findings and the clinical course of this patient.
The application of expert systems in the clinical laboratory.
Authors: Winkel P
Published: January 2020

An "expert system" consists of a knowledge base containing information of a general nature and an inference system that receives data from the user and applies the knowledge base to produce advice and explanations. An expert system stripped of its knowledge base (a tool) may be used to build new expert systems. Existing systems relevant for laboratory medicine are reviewed. The role in the laboratory of expert systems and their integration and evaluation are discussed.
Calculated bicarbonate or total carbon dioxide?
Authors: O'Leary T D , Langton S R
Published: January 2020

To test the relationship pK' = 6.103 + log[HCO3calc] - log[HCO3meas], we used a Corning 168 blood-gas analyzer to analyze 500 blood samples for pH and PCO2, from which we calculated a value for bicarbonate. We also analyzed 500 venous blood samples, collected simultaneously, for potentiometric total carbon dioxide with the Ektachem 700 analyzer. In a similar study of 415 arterial and venous blood samples, we determined total carbon dioxide colorimetrically with the SMA 6/60 analyzer. The coefficients of determination (r2) found for the difference observed between the calculated and measured bicarbonate values vs the pK' in the two studies were 0.86 and 0.96, respectively. The results also confirmed the positive bias caused by organic acids in the Ektachem method for total carbon dioxide. Analysis of the SMA 6/60 results indicated a significant decrease of the pK' in patients classified as having a metabolic acidosis.
Radioimmunoassay of 5-methoxytryptophol in plasma.
Authors: Skene D J , Churchill A , Raynaud F , Pevet P , Arendt J
Published: January 2020

5-Methoxytryptophol (ML) is synthesized by the pineal gland, but no radioimmunoassay has been described for its routine measurement in human plasma. We have developed and validated such an RIA. The assay is sensitive (detecting as little as 8 ng/L) and specific, and requires no extraction stage. A preliminary study of healthy volunteers showed an intra-individual variation in plasma ML that was independent of the sex of the subject and the time of daytime collection. Investigation of the 24-h pattern of ML in seven men revealed a low-amplitude daily rhythm (P less than 0.03). Mean concentrations of ML between noon and midnight significantly exceeded those between 0030 and 1130 hours in each individual (P less than 0.05). This assay is practical and convenient, and it should greatly assist in investigation of factors affecting concentrations of ML in human plasma.
Use of purified lyophilized human lactate dehydrogenase isoenzyme 5 in a study of measuring lactate dehydrogenase activity.
Authors: Smith D A , Moses G C , Henderson A R
Published: January 2020

We examined the stability of human lactate dehydrogenase (EC 1.1.1.27) isoenzyme 5--purified to a specific activity of about 400 kU/g--when lyophilized in a buffered, stabilized matrix of bovine albumin. This isoenzyme was prepared with a final activity of about 500 U/L and stored at -20, 4, 20, 37, and 56 degrees C for as long as six months. This isoenzyme decayed with approximate first-order kinetics, with an estimated half-life at -20 degrees C of about 475 years. Stability of reconstituted samples stored at 20 or 4 degrees C was poor, suggesting that the reconstituted material should be used without delay; material stored at -20 degrees C showed excellent stability for 15 days. We propose that such preparations might be further investigated as standards for use in electrophoresis of lactate dehydrogenase isoenzymes.
Four sensitive thyrotropin assays critically evaluated and compared.
Authors: Sgoutas D S , Barton E G , Hammarstrom M , Pete P M , Sgouta S A
Published: January 2020

We compared four sensitive procedures for thyrotropin (TSH)--Corning's Magic-Lite, ElectroNucleonics' Delfia, Baxter's Stratus, Nichols' Allegro--for their ability to completely discriminate TSH concentrations in sera in euthyroidism, hyperthyroidism, and hypothyroidism. We evaluated the analytical and clinical performance of these procedures according to previously published criteria. All procedures we examined fulfilled the criterion stipulating less than 1% overlap between the assay variation at the lower normal range limit and the assay sensitivity. Both measurements were made with an interassay CV not exceeding 10% to 15%. All procedures produced results that correlated well for specimens with TSH concentrations of greater than 0.4 milli-int. unit/L, and all four procedures clearly distinguished hypothyroid from euthyroid subjects. In a hyperthyroid-euthyroid comparison, three of the procedures, the Magic-Lite, Delfia, and Allegro, differentiated the two with 97% accuracy, the Stratus procedure with only 90% accuracy. The procedures appeared to differ even more in the measurement of TSH in serum of patients undergoing suppressive treatment with thyroid hormones and in hospitalized nonthyroidally ill patients. The observed differences among procedures were thought to be related in part to a matrix serum effect, which is accentuated in samples from hospitalized patients.
Use and applications of nucleic acid probes in the clinical laboratory.
Authors: Mifflin T E
Published: January 2020

The use and availability of nucleic acid probes are significantly greater than they were three to five years ago. This increased utilization has resulted because of numerous changes in molecular genetic procedures. First, the sources of probes have shifted from cloned genomic and cDNA fragments to single-strand oligonucleotides and RNA or "riboprobes." These newer probes offer defined composition and capability to create separate sense and anti-sense strands. Second, an increased variety of labeling techniques provide more ways to incorporate isotopic and nonisotopic species into these nucleic acid fragments than before. Third, the advent of nucleic acid amplification forecasts the ability to detect sub-attomol quantities of target nucleic acids, which could not be detected by classical hybridization methods. Fourth, more-sensitive signaling systems (e.g., chemiluminescence) now available will also contribute to the lowering of detection limits. Fifth, alternative sample-processing methods will shorten the time required to perform these assays. Finally, coupling these newer molecular probe procedures with automation will lead to improved precision and reliability and promote their introduction into the clinical laboratory as routine procedures. Additional factors such as clinical correlation, cost analysis, etc. will stimulate clinical laboratory personnel to participate further in development and use of these procedures.
Quantitative assays based on the use of replicatable hybridization probes.
Authors: Lomeli H , Tyagi S , Pritchard C G , Lizardi P M , Kramer F R
Published: January 2020

Amplifiable hybridization probes--molecules with a probe sequence embedded within the sequence of a replicatable RNA--will promote the development of sensitive clinical assays. To demonstrate their utility, we prepared a recombinant RNA that contained a 30-nucleotide-long probe complementary to a conserved region of the pol gene in human immunodeficiency virus type 1 (HIV-1) mRNA. Test samples were prepared, each containing a different number of HIV-1 transcripts that served as simulated HIV-1 mRNA targets. Hybridizations were carried out in a solution containing the chaotropic salt, guanidine thiocyanate. Probe-target hybrids were isolated by reversible target capture on paramagnetic particles. The probes were then released from their targets and amplified by incubation with the RNA-directed RNA polymerase, Q beta replicase (EC 2.7.7.48). The replicase copied the probes in an exponential manner: after each round of copying, the number of RNA molecules doubled. The amount of RNA synthesized in each reaction (approximately 50 ng) was sufficient to measure without using radioisotopes. Kinetic analysis of the reactions demonstrated that the number of HIV-1 targets originally present in each sample could be determined by measuring the time it took to synthesize a particular amount of RNA (the longer the synthesis took, the fewer the number of targets originally present). The results suggest that clinical assays involving replicatable hybridization probes will be simple, accurate, sensitive, and automatable.
Patient-side immunoassay system with a single-use cartridge for measuring analytes in blood.
Authors: Gibbons I , Gorin M , Yassinzadeh Z , Peterson P , Besemer D , Dillon K , Burd T , Hillman R , Smoluk G , Cobb M
Published: January 2020

We describe an immunoassay system suited to patient-side assay of therapeutic drugs and blood proteins. The system consists of an electronic monitor and single-use plastic cartridges containing dry reagents and liquid diluents. The monitor is turned on by insertion of a cartridge. To run the test, the user applies an unmeasured drop of blood to the cartridge when prompted by the monitor. All subsequent steps are performed without further user intervention and results are provided in less than 3 min. The system hemolyzes and precisely dilutes the blood. Hemoglobin concentration is measured, then the diluted blood is precisely diluted further and mixed with two dry reagents. The drug concentration is measured by a turbidimetric latex agglutination inhibition reaction. Theophylline and hemoglobin assay results for clinical samples correlate well with results of widely used comparison methods.
Diagnosis of recessive X-linked ichthyosis: quantitative HPLC/mass spectrometric analysis of plasma for cholesterol sulfate.
Authors: Shackleton C H , Reid S
Published: January 2020

A specific, accurate liquid-chromatographic/mass-spectrometric (HPLC/MS) method for measurement of cholesterol sulfate in plasma from normal individuals and patients with recessive X-linked ichthyosis (RXLI) is described. The method is superior to previously described techniques because it measures the analyte intact rather than after hydrolysis. Traces of free cholesterol in the sample analyzed do not add to the measured result. We used either [13C2]cholesterol sulfate or [2H6]cholesterol sulfate as internal standards, which we add to plasma before extraction. Use of such standards makes quantitative extraction unimportant. We use a single solid-phase extraction (SPE) C18 cartridge for plasma extraction. After the cartridge is washed with methanol/ammonium acetate solutions, the fraction containing the steroid sulfates is eluted with methanol, evaporated, and subjected to HPLC/MS analysis, wherein the molecular anions of analyte and internal standards are monitored. The peak ratio gives the cholesterol sulfate concentration directly. Using this method, we have diagnosed 24 patients with RXLI. Their concentration of cholesterol sulfate ranged between 41.7 and 185.3 mumol/L (mean 93.85, SD 31.2 mumol/L). In normal individuals (n = 9) the mean cholesterol sulfate concentration was 2.77 mumol/L (SD 0.62, range 2.05-3.95 mumol/L). The instrumentation required is complex, but the assay is simple: sample preparation takes about 30 min; mass spectrometry, 10 min. About 0.1 mL of plasma is required for cholesterol sulfate measurement in RXLI patients and 0.5-1 mL in normal individuals.
Time-resolved immunofluorometric assay of human pancreatic isoamylase in serum, with use of two monoclonal antibodies.
Authors: Diamandis E P , Papanastasiou-Diamandi A , Lustig V , Khosravi M J , Tan A
Published: January 2020

This new method for determining pancreatic isoamylase (EC 3.2.1.1) in serum involves two monoclonal antibodies: one immobilized in a microtitration well (the capture antibody), the other biotinylated. After the sample is incubated with the two antibodies, the captured immunocomplex is quantified by adding streptavidin labeled with a europium chelator and measuring the specific Eu3+ fluorescence in a time-resolved mode. Three assay protocols are proposed, involving incubation times of 90, 45, or 25 min. The assay has low (0.005%) cross-reactivity with the salivary isoenzyme. Analytical performance was satisfactory. Results correlate well with results obtained by measuring total amylase activity or by measuring pancreatic isoamylase activity after immunoinhibition. Unlike numerous current amylase assays, this method measures enzyme mass rather than enzyme activity. Potentially, this is a highly specific assay.
Rapid screening for deficiency of alpha 1-proteinase inhibitor.
Authors: Lloyd C , Travis J
Published: January 2020

This rapid screening procedure for detection of low but functional elastase-inhibitory activity in human plasma is based on the fact that incubation of excess porcine pancreatic elastase (EC 3.4.21.36) with plasma results in formation of a complex with active alpha 1-proteinase inhibitor (alpha 1PI, also called alpha 1-antitrypsin). In normal individuals all of the elastase is complexed, leaving no free enzyme to hydrolyze the elastase substrate, and the reaction mixture remains clear. Because individuals homozygous for the Z allele have relatively low concentrations of alpha 1PI, their plasma cannot complex all of the elastase in the assay. The uncomplexed enzyme hydrolyzes the elastase-specific p-nitroanilide substrate, producing a yellow reaction mixture. Use of this simple assay for early screening of individuals for alpha 1PI deficiency may substantially decrease the number of untreated cases of familial emphysema, a disorder that develops as a result of a genetically derived proteinase-proteinase inhibitor imbalance.
Fast, stable method for harvesting leukocytes for measuring intracellular magnesium.
Authors: Frandsen N J , McNair P
Published: January 2020

A new method for isolating blood leukocytes and measuring intracellular leukocyte magnesium is based on three successive centrifugations (1300 x g, 5 min, 22 degrees C), with no use of Ficoll, followed by two washes. We express the leukocyte magnesium concentration in micromoles of magnesium per gram of protein. Magnesium is measured by atomic absorption spectrophotometry, protein by the classic Lowry method. With the method 8% to 28% of the leukocytes are isolated and 0% to 0.025% of the erythrocytes. Of the isolated leukocytes 86% to 93% are viable. The magnesium concentration is constant at centrifugal forces greater than 1000 x g, at a centrifugation time of 5 to 15 min, and with use of two or three washes. However, the cells lose 1% of their magnesium content per hour when samples are left at 22 degrees C. The temperature influences this. In blood samples from 98 volunteer blood donors, mean leukocyte magnesium concentrations were 26.4 (SD 3.9) mumol/g. There was no significant correlation between the magnesium concentration and the relative amount of neutrophils.
dbPepNeo: a manually curated database for human tumor neoantigen peptides
Authors: Tan Xiaoxiu , Li Daixi , Huang Pengjie , Jian Xingxing , Wan Huihui , Wang Guangzhi , Li Yuyu , Ouyang Jian , Lin Yong , Xie Lu
Published: January 2020

Neoantigens can function as actual antigens to facilitate tumor rejection, which play a crucial role in cancer immunology and immunotherapy. Emerging evidence revealed that neoantigens can be used to develop personalized, cancer-specific vaccines. To date, large numbers of immunogenomic peptides have been computationally predicted to be potential neoantigens. However, experimental validation remains the gold standard for potential clinical application. Experimentally validated neoantigens are rare and mostly appear scattered among scientific papers and various databases. Here, we constructed dbPepNeo, a specific database for human leukocyte antigen class I (HLA-I) binding neoantigen peptides based on mass spectrometry (MS) validation or immunoassay in human tumors. According to the verification methods of these neoantigens, the collection of peptides was classified as 295 high confidence, 247 medium confidence and 407 794 low confidence neoantigens, respectively. This can serve as a valuable resource to aid further screening for effective neoantigens, optimize a neoantigen prediction pipeline and study T-cell receptor (TCR) recognition. Three applications of dbPepNeo are shown. In summary, this work resulted in a platform to promote the screening and confirmation of potential neoantigens in cancer immunotherapy.

Database URL: www.biostatistics.online/dbPepNeo/.
Accelerometer-Measured Sedentary and Physical Activity Time and Their Correlates in European Older Adults: The SITLESS Study
Authors: Giné-Garriga Maria , Sansano-Nadal Oriol , Tully Mark A , Caserotti Paolo , Coll-Planas Laura , Rothenbacher Dietrich , Dallmeier Dhayana , Denkinger Michael , Wilson Jason J , Martin-Borràs Carme , Skjødt Mathias , Ferri Kelly , Farche Ana Claudia , McIntosh Emma , Blackburn Nicole E , Salvà Antoni , Roqué-i-Figuls Marta
Published: January 2020

Background

Sedentary behavior (SB) and physical activity (PA) are important determinants of health in older adults. This study aimed to describe the composition of accelerometer-measured SB and PA in older adults, to explore self-reported context-specific SB, and to assess sociodemographic and functional correlates of engaging in higher levels of SB in participants of a multicenter study including four European countries.

Method

One thousand three hundred and sixty community-dwelling older adults from the SITLESS study (61.8% women; 75.3 ± 6.3 years) completed a self-reported SB questionnaire and wore an ActiGraph accelerometer for 7 days. Accelerometer-determined compositional descriptive statistics were calculated. A fixed-effects regression analysis was conducted to assess the sociodemographic (country, age, sex, civil status, education, and medications) and functional (body mass index and gait speed) correlates.

Results

Older adults spent 78.8% of waking time in SB, 18.6% in light-intensity PA, and 2.6% in moderate-to-vigorous PA. Accelerometry showed that women engaged in more light-intensity PA and walking and men engaged in higher amounts of moderate-to-vigorous PA. Watching television and reading accounted for 47.2% of waking time. Older age, being a man, single, taking more medications, being obese and overweight, and having a slower gait speed were statistically significant correlates of more sedentary time.

Conclusions

The high amount of SB of our participants justifies the need to develop and evaluate interventions to reduce sitting time. A clinically relevant change in gait speed can decrease almost 0.45 percentage points of sedentary time. The distribution of context-specific sedentary activities by country and sex showed minor differences, albeit worth noting.
A-optimal design method to determine the regularization parameter of coseismic slip distribution inversion
Authors: Wang Leyang , Gu Wangwang
Published: January 2020

The key to the inversion of a coseismic slip distribution is to determine the regularization parameters. In view of the determination of regularization parameters in seismic slip distribution inversion, the A-optimal design method is proposed in this paper. The L-curve method and A-optimal design method are used to design simulation experiments, and the inversion results show that the A-optimal design method is superior to the L-curve method in determining the regularization parameters. These two methods are also used to determine the regularization parameters of the L'Aquila and Lushan earthquake slip distribution inversions, and the results are consistent with those of other research conducted at home and abroad. Compared with the L-curve method, the A-optimal design method has the advantages of a high accuracy that does not rely on the data fitting accuracy.
Rules of Engagement: Implementing Student-Centered Learning in Breast Imaging
Authors: Jordan Sheryl G , Deans Elizabeth C , Johnson Meredith , Lee Sheila S , Beck Dallaghan Gary L
Published: January 2020

In response to changing learner needs, our institution launched a new translational curriculum wherein basic sciences and clinical skills were integrated, longitudinal patient care experiences provided, and earlier opportunities in specialty fields introduced to better inform residency program decisions. Medical students taking the breast imaging elective were enrolled in a breast imaging immersive experience designed to meet the School of Medicine’s core competencies. In focusing the elective on a narrow range of specialized topics and skills, we labeled this experience the Breast Imaging Boot Camp. Outcome data from March 6, 2017, to April 26, 2019, have been analyzed for this report. The modifications made to the elective include: e-mailing a detailed orientation document to students prior to the start of the rotation; assigning students to diagnostic, procedural, and screening roles; the attendance of students at introductory radiology symposia; students’ weekly attendance at institutional multidisciplinary and divisional breast radiologic–pathologic correlation conferences; student self-study assignments using faculty-vetted resources; student participation in breast biopsy simulation and small parts ultrasound laboratories; the development of a student-centric radiology educational website; and student-directed publishing of digital case files. Medical student feedback and course analytics indicated superior course evaluation scores reinforced by narrative feedback. In website domain utilization data, the breast file domain is the most viewed subspecialty domain. The Breast Imaging Boot Camp is a successful curriculum. Its novelty lies in both its integrated approach to medical student core competencies and its clinician educators’ use of today’s student-favored teaching tools.