bluuebunny/crossref_metadata_2025 huggingface dataset:
Liquid-chromatographic measurement of riboflavin in serum and urine with isoriboflavin as internal standard.
Authors: Lambert W E , Cammaert P M , De Leenheer A P
Published: January 2020

We describe how riboflavin can be precisely and accurately measured in serum and urine. A structural analog, isoriboflavin, is used as an internal standard. Urine samples are prepared by adding the internal standard and trichloroacetic acid. For serum, proteins are denatured with trichloroacetic acid. A simple Sep-pak treatment of the supernate removes contaminating interferents. Within- and between-run precision is reflected by respective CVs of 2.2 and 4.9% for urine at 180 micrograms/L and 4.4 and 7.3% for serum at 10 micrograms/L. The standard curve is linear far beyond the concentrations encountered in serum and urine. The detection limit is estimated to be 10 micrograms/L and 1 microgram/L for urine and serum, respectively. The normal reference interval, as determined from 50 results for each matrix, is 36 to 349 micrograms/g of creatinine (mean 112 micrograms/g) for urine and 5.5 to 14.4 micrograms/L (mean 8.8 micrograms/L) for serum. Both distributions are skewed. The method is suited for routine use.
A highly sensitive immunoassay system involving antibody-coated tubes and liposome-entrapped dye.
Authors: O'Connell J P , Campbell R L , Fleming B M , Mercolino T J , Johnson M D , McLaurin D A
Published: January 2020

In this colorimetric immunoassay for digoxin, large, unilamellar phospholipid vesicles approximately 0.2 micron in diameter are loaded with high concentrations of Sulforhodamine B. Digoxigenin coupled to phosphatidylethanolamine, incorporated into the lipid formulation, confers immunological specificity. The liposomes are then used as tracers in simple competitive-binding immunoassays with antibody-coated tubes. Results are amplified by 10(3) to 10(4) of what could be achieved with one label group attached to each hapten, so that the results can be read spectrophotometrically. The stability of the liposomes is excellent. The method should be applicable to measuring a wide variety of analytes.
"The Yellow IRIS" urinalysis workstation--the first commercial application of "automated intelligent microscopy".
Authors: Deindoerfer F H , Gangwer J R , Laird C W , Ringold R R
Published: January 2020

Since 1982, "automated intelligence microscopy" (AIM) has been refined and adapted to perform the portion of the urinalysis profile traditionally done by a microscope. AIM and analytical subsystems measuring relative density and performing dipstick chemistry compose the main elements of "The Yellow IRIS" urinalysis workstation, an attended system for automation and standardization of routine urinalysis. Performance data gathered at three laboratory test sites show AIM to be analytically consistent over the required range of particulate enumeration, and show that it detects 20% more abnormalities than by conventional microscopy, and with greater precision (CVs 5 to 20%). Complete urinalysis, including the microscopic examination, requires little more than 1 min for normal specimens, 3 min for most abnormal specimens. Actual throughput rate varies with the particulate composition of specimens; typically, it averages greater than 30 specimens per hour, a 300% improvement in urinalysis productivity by CAP standards and an almost 500% improvement when typical emergency-use demands are taken into account.
Determinations of immunoglobulins G, A, and M in the Technicon RA-1000.
Authors: Ng R H , Altaffer M , Statland B E
Published: January 2020

We evaluated the Technicon RA-1000 "Random Access" analyzer for the measurements of immunoglobulins G, A, and M in serum by turbidimetry. For various concentrations of serum pools the total CV ranged from 3.0 to 4.5% for IgG, from 3.1 to 3.2% for IgA, and from 4.8 to 5.4% for IgM. The linearity of the standard curves was good in the clinically useful ranges. Results for patients' samples correlated well with those determined with the Beckman Auto ICS (rate nephelometric method). Normal reference intervals (based on data for 200 healthy blood donors) are 7.67 to 16.87 g/L for IgG, 0.87 to 3.89 g/L for IgA, and 0.61 to 2.70 g/L for IgM. The RA-1000 requires only 8 microL for all three tests and can analyze 25 specimens for all three proteins in 37 min. Compared with other methods for immunoglobulin assays, the RA-1000 has higher throughput and offers significant savings in labor and reagent costs.
Determination of total protein in human milk: comparison of methods.
Authors: Keller R P , Neville M C
Published: January 2020

We evaluated four spectrophotometric methods for suitability in determining total protein in human milk samples. Results were compared with those by an established method for the determination of total nitrogen (micro-Kjeldahl). Values for total protein by all four methods significantly correlated with micro-Kjeldahl. For the biuret assay, Lowry-Peterson assay, Bio-Rad Coomassie Blue assay, and Pierce BCA assay, the correlation coefficients were 0.96, 0.97, 0.89, 0.99, respectively. We also assessed the sensitivity of each assay to several purified human milk proteins. The Pierce BCA assay showed the least difference in values among different types of protein. For this reason, and because it showed the greatest precision (with acceptable constant and proportional error), we recommend the BCA method.
The EPOS Automated Selective Chemistry Analyzer evaluated.
Authors: Moses G C , Lightle G O , Tuckerman J F , Henderson A R
Published: January 2020

We evaluated the analytical performance of the EPOS (Eppendorf Patient Oriented System) Automated Selective Chemistry Analyzer, using the following tests for serum analytes: alanine and aspartate aminotransferases, lactate dehydrogenase, creatine kinase, gamma-glutamyltransferase, alkaline phosphatase, and glucose. Results from the EPOS correlated well with those from comparison instruments (r greater than or equal to 0.990). Precision and linearity limits were excellent for all tests; linearity of the optical and pipetting systems was satisfactory. Reagent carryover was negligible. Sample-to-sample carryover was less than 1% for all tests, but only lactate dehydrogenase was less than the manufacturer's specified 0.5%. Volumes aspirated and dispensed by the sample and reagent II pipetting systems differed significantly from preset values, especially at lower settings; the reagent I system was satisfactory at all volumes tested. Minimal daily maintenance and an external data-reduction system make the EPOS a practical alternative to other bench-top chemistry analyzers.
Biological variance of cholinesterase and 5'-nucleotidase in serum of healthy persons.
Authors: Moses G C , Tuckerman J F , Henderson A R
Published: January 2020

We measured cholinesterase (EC 3.1.1.8) and 5'-nucleotidase (EC 3.1.3.5) activities in serum of 24 healthy laboratory staff during 12 months. Overall mean activities ranged from 5.3 to 13.4 kU/L for cholinesterase and 5.4 to 9.8 U/L for 5'-nucleotidase. Cholinesterase activity was significantly (p less than 0.01) higher for men than for women. 5'-Nucleotidase activity was significantly (p = 0.01) higher for subjects 40 years or older than for those younger than 40, but was not different with respect to sex or time of year. Average intra- and interindividual variances (SD2) were 0.38 and 2.69 for cholinesterase and 1.41 and 0.97 for 5'-nucleotidase, respectively. Intra- to interindividual standard deviation ratios were 0.38 for cholinesterase and 1.21 for 5'-nucleotidase. Average within-run analytical variances were 0.13 and 0.3 (4% and 13% of total variance) for cholinesterase and 5'-nucleotidase, respectively. The importance of these findings in regards to diagnostic interpretation of serum cholinesterase and 5'-nucleotidase results is discussed.
In vitro oxidation of uric acid in serum by methylene blue.
Authors: Andranovich T , Kelner M J
Published: January 2020

Methylene blue oxidizes uric acid to allantoin in vitro, analogous to the reaction between uric acid and phosphotungstate, although the rate is considerably slower. The reaction requires oxygen and produces hydrogen peroxide. As little as 10 micromol of methylene blue per liter in a serum sample will produce a measurable difference in the apparent uric acid concentration within 4 h at room temperature. Methylene blue may be administered for various medical reasons. If uric acid is to be measured in such sera, the procedure should be performed within 30 min, whatever method of analysis is used.
Clinical and analytical evaluation of kits for measurement of creatine kinase isoenzyme MB.
Authors: Koch T R , Mehta U J , Nipper H C
Published: January 2020

We studied the analytical and clinical performance of six methods for creatine kinase (EC 2.7.3.2) isoenzyme MB (CK-MB): three immunoassays (Behring, Hybritech, and International Immunoassay Labs); one immunoinhibition assay (Roche); one immunoinhibition/column method (Du Pont); and one electrophoretic method (Beckman). Between-day precision for all kits was poor at the upper reference limit. All methods gave results linearly related to CK-MB concentration and all were free from CK-MM, CK-BB, and adenylate kinase interference. Only the Du Pont method was adversely affected by atypical isoenzymes. For diagnosis of acute myocardial infarction in a coronary care population (n = 40; prevalence = 45%), all methods were approximately 95% efficient, when appropriate reference criteria were used. Some manufacturers fail to provide data for an appropriate (acutely ill, non-infarct) reference population; decreased diagnostic specificity may result from use of reference ranges based on results for healthy subjects. Expression of CK-MB as a percent of total CK degrades efficiency unless total CK is markedly increased.
Factors affecting determination of delta-aminolevulinate by use of Ehrlich's reagent.
Authors: Tomokuni K , Hirai Y
Published: January 2020

We investigated the formation of delta-aminolevulinic acid (ALA)-pyrrole and subsequent color development used in determining ALA in urine. Of three condensation reagents examined--acetylacetone, ethyl acetoacetate, and methyl acetoacetate--the last was the best. When the ALA-pyrrole is reacted with Ehrlich's reagent in ethyl acetate according to one procedure, the optimum concentration of perchloric acid in the reagent is 0.5 to 1.0 mol/L. Its optimum concentration is 1.0 to 2.3 mol/L when the reaction takes place in water.
Digoxin--issues and controversies.
Authors: Soldin S J
Published: January 2020

This review deals briefly with the clinical pharmacology of digoxin and reviews in some depth the problems inherent in methods of digoxin measurement. Digoxin-like factors (materials that cross react in digoxin immunoassays) are discussed, and some current evidence suggesting the existence of a new hormone, endoxin (endogenous digoxin), is summarized.
Concentration of glucocorticoid receptor sites in normal human lymphocytes.
Authors: Steiner A E , Wittliff J L
Published: January 2020

We used a whole-cell glucocorticoid receptor assay to examine characteristics of the glucocorticoid receptor in the lymphocytes of normal human donors. We measured binding of [3H]dexamethasone to the lymphocytes of four different donors on several different occasions; the variation about the mean for the assays was +/- 15%. Whole-cell assays in 15 normal subjects showed a mean value of 6.18 fmol/10(6) cells or 3722 sites per cell, with a somewhat higher level in men (7.67 fmol/10(6) cells, or 4620 sites per cell) than women (4.48 fmol/10(6) cells, or 2698 sites per cell). We saw no correlation between donor age and receptor values, in either group. Assays in which we used [3H]prednisolone demonstrated similar binding properties as with [3H]dexamethasone. The mean glucocorticoid receptor value for normal human T-cells from three donors was 2.52 fmol/10(6) cells, or 1518 sites per cell.
Specific interaction between antidigoxin antibodies and digoxin-like immunoreactive substances in cord serum.
Authors: Scherrmann J M , Sandouk P , Guedeney X
Published: January 2020

Digoxin-like immunoreactive substances (DLIS) have been quantified by two different digoxin radioimmunoassays (RIA) in 47 cord sera. The mean DLIS value (in digoxin equivalents) ranged from 0.960 (SD 0.184) to 0.181 (SD 0.104) nmol/L between the two different kits and different lot numbers of the reagents. One of the RIA methods showed an obvious lot-to-lot effect. The use of a longer incubation interval and a higher incubation temperature markedly decreased cross reactions with DLIS. The effect of modifying the incubation conditions in RIA is similar to that described for assays of steroids because the dissociation rates of the immunocomplex play a critical role. Data suggest a specific interaction between DLIS and digoxin antibodies. Control of the incubation conditions is recommended, to decrease or increase the amount of the DLIS in cord serum specimens.
Determination of urinary lysozyme for potential detection of tubular dysfunction in diabetic nephropathy.
Authors: Shima K , Hirota M , Fukuda M , Tanaka A
Published: January 2020

Seeking to study whether measurement of lysozyme (EC 3.2.1.17) in urine by a reliable radioimmunoassay can provide a suitable index of renal tubular function and how lysozymuria develops in temporal relation to proteinuria in diabetic nephropathy, we have compared the urinary excretion of lysozyme and beta 2-microglobulin with the 15-min excretion rate of phenolsulfonphthalein in 39 patients with Type 2 (non-insulin-dependent) diabetes and investigated the temporal relation between the onset of lysozymuria and proteinuria in 15 patients with Type 1 (insulin-dependent) diabetes. The concentrations of lysozyme and beta 2-microglobulin in urine increased in proportion to the decrease in the rate of excretion of phenolsulfonphthalein in these patients. The coefficient of correlation between lysozyme concentration and the 15-min excretion rate of phenolsulfonphthalein (r = -0.70) was higher than that between beta 2-microglobulin concentration and the 15-min excretion rate of phenolsulfonphthalein (r = -0.46). Abnormally high lysozymuria, suggesting the existence of tubular dysfunction, was demonstrated in six of the patients with Type 1 diabetes who showed no proteinuria or only a slight increase in urinary protein excretion. Lysozymuria may thus be added to a list of the indicators for diabetic nephropathy.
Measurement of hemoglobin A1c by a new liquid-chromatographic assay: methodology, clinical utility, and relation to glucose tolerance evaluated.
Authors: Jeppsson J O , Jerntorp P , Sundkvist G , Englund H , Nylund V
Published: January 2020

A chromatographic method for determining glycated hemoglobin (Hb A1c) by use of a new monodisperse cation-exchanger has been investigated. Hb A1c was separated from other "minor hemoglobins": Hb F, Hb A3 (the glutathione adduct), and the acetaldehyde adduct in alcoholics. The method was fully automated and a single column could be used for more than 1000 runs. The normal reference interval was 4.0-5.2%; the interval for diabetic outpatients was 5.6-12.4%. Within-run and the between-run CVs were less than 0.9% and 1.7%, respectively. Carbamylation in uremic patients who were undergoing hemodialysis increased the proportion of Hb A1c to 1%. Hb A1c results were compared with results from glucose tolerance tests. In our study, Hb A1c less than 5.5% excluded diabetes: subjects with Hb A1c greater than 6.2% showed diabetes. If blood sampled during fasting had been screened with determinations of glucose and Hb A1c, only 20% of referred subjects would have needed an oral glucose tolerance test for diagnosis of diabetes.
External quality assessment of assays of lead in blood.
Authors: Bullock D G , Smith N J , Whitehead T P
Published: January 2020

The operation and results of the United Kingdom External Quality Assessment Scheme for Lead in Blood, which currently has about 140 participants within and outside the U.K. are described. The choice of specimens, scheme design, and scoring of performance are discussed, as is the validity of the consensus values used as a basis for the scoring. There has been a continued improvement in performance as assessed by this program, some of which appears to have been stimulated by the introduction of Variance Index scoring. The relative performance of the methods for assaying lead in blood is also examined.
Evaluation of a quantitative immunoturbidimetric assay for rheumatoid factors.
Authors: Melamies L M , Ruutsalo H M , Nissilä H
Published: January 2020

A quantitative immunoturbidimetric assay for rheumatoid factors (RF) is described, based on the immunoprecipitation between aggregated human IgG and rheumatoid factors in serum. The resulting turbidity is measured photometrically at 340 nm. The method is standardized against the WHO international reference preparation and the results are expressed in international units per milliliter (int. units/mL). Results correlate well with those by different latex-agglutination techniques (r = 0.80-0.96). The correlation with Waaler-Rose test modifications were 0.75 and 0.92. The within-run and between-run coefficients of variations were respectively from 1.2 to 2.6% and 1.3 to 1.9% for high and low RF concentrations. Quantitative and reproducible results, together with high throughput of samples and compatibility with most clinical chemistry analyzers and photometers, make this new assay well suited for routine screening and monitoring of rheumatoid factors.
Direct chemiluminescence immunoassay for estradiol in serum.
Authors: De Boever J , Kohen F , Usanachitt C , Vandekerckhove D , Leyseele D , Vandewalle L
Published: January 2020

In this simple, reliable, fast solid-phase chemiluminescence immunoassay for directly measuring (i.e., without prior extraction) estradiol-17 beta in serum, a monoclonal antibody is used that binds estradiol with high affinity (Ka = 10(10) L/mol), and does not bind other steroids tested, the highest cross reactivity observed being 0.1% for estradiol-17 alpha. In this system the monoclonal antibody is bound to the wells of microtiter plates via a second antibody directed against the monoclonal antibody. Fifty microliters of serum and estradiol-displacing agents are added, followed by 100 pg of estradiol-isoluminol conjugate, and the label is measured by luminometry after the binding reaction. The sensitivity of the assay is 180 pmol per liter of serum, and the effective working range at less than or equal to 10% CV is 270 to 6700 pmol/L. Analytical recovery of added estradiol averaged 99.7% (SD 6.5%). Within- and between-assay CVs ranged between 5 and 12.7%. Thirty-five unknown serum samples can be assayed within 4 h. Results correlated well with those obtained with a direct RIA: r = 0.94 (n = 149). This assay opens new perspectives for chemiluminescence immunoassays.
Total urinary estriol determined by "on-line" liquid chromatography with ultraviolet detection.
Authors: Schöneshöfer M , Dhar T K , Ioanides D
Published: January 2020

We report a fully automated method for the specific assessment of total estriol in urine. Acid-hydrolyzed urine is concentrated on a short reversed-phase column and prepurified with methanolic Tris buffer, the pH of the buffer increasing from 7 to 11. The anionic form of the estriol molecule is eluted with methanolic NaOH. By acidification in a mixing chamber, estriol in its neutral form is focused on the top of a second short reversed-phase column, effectively chromatographed on an analytical column, and quantified by ultraviolet absorbance at 278 nm. Losses of estriol throughout the total procedure are negligible and thus external calibration is feasible for quantification. Analytical recoveries for estriol-supplemented urines ranged from 98.3 to 105%. Replicate analyses of a urine containing 68 mumol of estriol per liter gave a CV of 2.76%. As little estriol as 2 mumol/L can be detected. Results from pregnancy urines correlated well with those of radioimmunoassay. The method is especially suited for clinical emergencies in a routine laboratory.
Quantification of bone alkaline phosphatase in serum by precipitation with wheat-germ lectin: a simplified method and its clinical plausibility.
Authors: Behr W , Barnert J
Published: January 2020

We report an easy, rapid method for quantifying bone isoenzyme of alkaline phosphatase (EC 3.1.3.1., ALP) in serum. The original method described by Rosalki and Ying Foo (Clin Chem 1984;30:1182-6) was somewhat simplified. In contrast to their results, we found that bone ALP is precipitated quantitatively by wheat-germ lectin. To check the clinical plausibility of the method, we used samples from several comparison groups (blood donors, children, pregnant women, patients with neoplasms but without skeletal involvement) and a large number of patients suffering from bone diseases and diseases of the liver and biliary tree. Measured activities of bone ALP nearly always correlated with the clinical diagnosis. Only patients with hepatitis often had pathological bone activities not in accord with the other findings. Possible reasons for this observation are discussed.
Extending the linear range for kinetic reactions by considering the relationship between enzyme activity of the reagent and measurement intervals.
Authors: Goren M P , Davis J E
Published: January 2020

We show that the rate-concentration curve is sigmoidal for enzyme-catalyzed procedures that are commonly applied to rapid automated analyzers. Linear data can be obtained by judicious selection of the reagent enzyme activity (Vm) and the measurement interval (t1 to t2). For determination of substrate concentrations much less than Km, conditions that balance linearity with sensitivity and accuracy are obtained when Vm/Km = [ln (t2/t1)]/(t2-t1). We also present theoretical and experimental data that show the linear range can be extended to concentrations exceeding the Km value. We illustrate the application of theoretically appropriate conditions by analysis of procedures reported to be optimized. Familiarity with these concepts can obviate laborious and potentially misleading experimentation.
Simultaneous liquid-chromatographic determination of 3,4-dihydroxyphenylglycol, catecholamines, and 3,4-dihydroxyphenylalanine in plasma, and their responses to inhibition of monoamine oxidase.
Authors: Eisenhofer G , Goldstein D S , Stull R , Keiser H R , Sunderland T , Murphy D L , Kopin I J
Published: January 2020

This is a reversed-phase liquid-chromatographic method, with electrochemical detection, for simultaneously measuring, in plasma, the concentrations of the catecholamine precursor dihydroxyphenylalanine (DOPA); the endogenous catecholamines norepinephrine, epinephrine, and dopamine; and the deaminated catecholamine metabolites dihydroxyphenylacetic acid (DOPAC) and dihydroxyphenylglycol (DHPG). We used this method to assess effects of monoamine oxidase (EC 1.4.3.4) inhibition in humans. Plasma DHPG concentrations as determined by the present method (mean 826, SEM 61 ng/L) were similar to those found by other methods. Inhibition of monoamine oxidase (by administering deprenyl or tranylcypromine) decreased plasma DHPG by greater than 65%, plasma DOPAC by greater than 50%, and plasma DOPA by about 20%, without consistently affecting norepinephrine or epinephrine. Simultaneous measurement of DOPA, catecholamines, and DHPG may be useful for examining the synthesis, release, and intraneuronal metabolism of norepinephrine. The assay method is rapid, reliable, and simple, and it provides a more comprehensive assessment of noradrenergic nervous function than does measurement only of catecholamines.
Measurements of serum gamma-seminoprotein and prostate specific antigen evaluated for monitoring carcinoma of the prostate.
Authors: Siddall J K , Shetty S D , Cooper E H
Published: January 2020

We have compared the concentrations in serum of gamma-seminoprotein (gamma-SM) and prostate specific antigen (PSA), two antigens of prostatic origin that are synthesized independently of prostatic acid phosphatase (PAP, EC 3.1.3.2), to assess their potential in monitoring prostatic cancer. At presentation, 27/30 (90%) patients with metastases had a PSA concentration greater than 10 ng/mL, and 29/30 (97%) a gamma-SM concentration greater than 10 ng/mL; 21/61 (34%) with disease but without metastases had an abnormal content of PSA, and 23/61 (38%) an abnormal gamma-SM. Concentrations of PSA and gamma-SM were significantly correlated (r = 0.68, p less than 0.001). In 20 patients without metastases followed longitudinally, the median concentrations of gamma-SM, PSA, and PAP in the 13 patients who developed bony metastases or showed signs of local spreading of the tumor were 58 ng/mL, 34 ng/mL, and 2.1 U/L, respectively. The corresponding median values in the seven patients who remained clinically stable were 2.5 and 3.9 ng/mL, and 2.3 U/L. We conclude that either PSA or gamma-SM can warn of disease progression when PAP activities are still within normal limits.
Evaluation of an enhanced luminescence assay for alpha-fetoprotein.
Authors: John R , Henley R , Shankland D
Published: January 2020

We evaluated a two-site enhanced luminescence immunoenzymometric assay (Amerlite; Amersham International) for alpha-fetoprotein (AFP) in maternal serum and amniotic fluid. The assay is rapid, involving two incubations totalling 4 h. The working range of the assay for serum AFP is 5.5 to 750 kilo-int. units/L (CV less than 10%), with a sensitivity of detection of 0.2 kilo-int. unit/L. The regression equation for the Amerlite assay (y) and our in-house RIA procedure (x) was y = 0.816x + 2.9 (n = 142, r = 0.96). Analytical recovery of added AFP (code 72/227) at three concentrations was 86.7%. Serum AFP concentrations were measured at 15 to 18 weeks of gestation in subjects with normal pregnancies and in subjects whose pregnancies resulted in open neural tube defects; all of the latter had serum AFP concentrations greater than 2.5 multiples of the median. We find the Amerlite system to be an efficient, reliable system for screening for open neural tube defects without use of hazardous radioactive labels.
6 beta-hydroxycortisol in serum and urine as determined by enzyme immunoassay on microtitre plates.
Authors: Zhiri A , Mayer H A , Michaux V , Wellman-Bednawska M , Siest G
Published: January 2020

We modified our previous enzyme immunoassay (Clin Chim Acta 1986;157:267-76) for estimating 6 beta-hydroxycortisol (6 beta-OHF) to provide a more routine procedure for measuring it in urine and to increase sensitivity for its measurement in serum. Precision, analytical recovery, specificity, and detection limit are reported. 6 beta-OHF and 17-hydroxycorticosteroids in 24-h and 4-h (08:00-12:00 h) urine collections from healthy volunteers showed no significant day-to-day differences in excretion rate for either collection period. Mean values for 6 beta-OHF in serum of men, women, and women taking oral contraceptives showed no significant differences among these groups.
Age dependence of tissue plasminogen activator concentrations in plasma, as studied by an improved enzyme-linked immunosorbent assay.
Authors: Rånby M , Bergsdorf N , Nilsson T , Mellbring G , Winblad B , Bucht G
Published: January 2020

A procedure for improving the specificity of enzyme-linked immunosorbent assays (ELISA) was devised, based on addition of antigen-specific or non-immune immunoglobulins to the citrated plasma sample and defining the difference in assay response between these two mixtures as the antigen-specific part of the response. When applied to measurement of tissue plasminogen activator (t-PA; EC 3.4.21.31) antigen in plasma, this procedure resulted in elimination of the overestimates obtained in a large proportion (10-20%) of patients' samples when assayed according to the conventional ELISA technique. Basal t-PA concentrations in plasma were found to be highly age-dependent, normal values being about 3 micrograms/L for adults near 30 years of age and about 10 micrograms/L for those over 60. Patients with gallbladder stone disease had increased mass concentrations of t-PA in plasma, even when corrected for the age effect; patients with multi-infarct dementia did not.
Validation of a semi-automated technique for measuring lactate in whole blood.
Authors: Weil M H , Leavy J A , Rackow E C , Halfman C J , Bruno S J
Published: January 2020

An electrode-based lactate analyzer was recently developed to facilitate "stat" laboratory measurement of lactate in whole blood. The results obtained with this technique were compared with those of our continuous-flow enzymatic method for 172 analyses of samples from 25 pigs; 88 analyses of samples from 12 normal human volunteers before, during, and after exercise; and 91 analyses of samples from 80 critically ill patients. This comparison revealed strongly significant linear correlations: r = 0.992 for the porcine analyses and r = 0.994 for the combined human analyses. In analytical-recovery studies with pooled porcine plasma with an initial lactate concentration of 3.85 mmol/L, 94.3% of added lactate (1, 5, and 10 mmol/L) was accounted for in the electrode technique, 96% in the reference technique. We conclude that the electrode-based lactate analyzer is rapid, precise, and accurate for measurement of lactate in whole-blood samples.
Enhanced luminescence immunoassay: evaluation of a new, more sensitive thyrotropin assay.
Authors: John R , Henley R , Chang D , McGregor A M
Published: January 2020

We measured thyrotropin (TSH) with an enhanced luminometric assay ("Amerlite"; Amersham International). The detection limit of the assay is 0.02 milli-int. unit/L. Within-assay precision was 6.7 and 7.8% at 3.77 and 12.1 milli-int units/L, respectively, and between-assay precision was almost identical, whether singleton or duplicate samples were assayed. TSH measured in 132 euthyroid subjects ranged from 0.06 to 4.13 milli-int. units/L (mean 1.52, SD 0.86). Similar concentrations were found in 20 healthy pregnant women and 19 of 20 healthy post-menopausal women (one of whom had undetectable TSH). In 17 patients with primary hypothyroidism, TSH concentrations ranged from 9.34 to greater than 200 milli-int. units/L; and in 53 of 59 patients with hyperthyroidism, TSH concentrations were undetectable, ranging in the remaining six from 0.03 to 0.06 milli-int. unit/L. Results for TSH in 28 patients stimulated with thyroliberin were consonant with the results of the thyroliberin test in 25 cases. Thus, for most patients, measurement of a basal TSH concentration evidently will predict their thyroidal status and also the response to thyroliberin, but a few will require additional tests of thyroid function.
Discrepancy between uptake of serum triiodothyronine as measured by albumin and charcoal methods in nonthyroidal illnesses.
Authors: Konno N , Taguchi H , Hagiwara K , Taguchi S , Hatsukaiwa Y , Kumagai M
Published: January 2020

Triiodothyronine (T3) uptake as determined with albumin (AT3U) and with charcoal (CT3U) as binders were compared for patients with low-T3 syndrome. Both T3U values were identical in normal subjects and in patients with thyroidal disorders, pregnancy, low thyroxin-binding globulin (TBG), or acute hepatitis with high concentration of TBG. In the low-T3 syndrome, AT3U was significantly higher than CT3U (38.5 +/- 7.2% vs 29.4 +/- 4.9%, mean +/- SD, n = 37, p less than 0.001), and this difference increased as TBG and albumin concentrations decreased. Decreasing the concentration of TBG and albumin in serum by dilution increased the AT3U more than the CT3U value. Adding albumin to serum decreased AT3U, but not CT3U, when serum TBG and albumin concentrations were low. The free-thyroxin (T4) index as calculated by the AT3U ratio and T4 more accurately reflects the concentration of free T4 in low-T3 syndrome than does the index calculated from the CT3U ratio and T4. Thus AT3U is determined primarily by changes in TBG and albumin concentrations, so that the free-T4 index based on the AT3U ratio and T4 may be more useful than the CT3U method for evaluating free-T4 concentrations in low-T3 syndrome, which frequently has concomitant decreases in TBG and albumin concentrations.
"Fragmented" isoenzymes of alkaline phosphatase in the diagnosis of transient hyperphosphatasemia.
Authors: Schoenau E , Herzog K H , Boehles H J
Published: January 2020

We describe the appearance of "fragmented" isoenzymes of serum alkaline phosphatase (EC 3.1.3.1) in two cases of transient hyperphosphatasemia. We determined the isoenzymes by liquid chromatography, then characterized them by heat inactivation, inhibition with 5 mmol/L L-phenylalanine solution, and electrophoresis on cellulose acetate membranes. We suspect that a virus-induced decrease in clearance of the enzyme from serum is responsible for a similar increase of bone and liver isoenzyme activities and for the presence of these fragmented isoenzymes in transient hyperphosphatasemia.
Multiple myeloma with serum IgM kappa and Bence Jones lambda biclonal gammopathy.
Authors: Graziani M S , Lippi U
Published: January 2020

We report a rare finding: IgM kappa and Bence Jones lambda double gammopathy in serum of a 80-year-old man with untreated symptomatic multiple myeloma. The unusual findings are confined to the laboratory studies demonstrating also a Bence Jones lambda proteinuria, high erythrocyte sedimentation rate (113 mm/h), and anemia. The synthesis of the different light chains seems to occur in separate cellular clones.
Improved fluorescence polarization assay for use in evaluating fetal lung maturity. I. Development of the assay procedure.
Authors: Tait J F , Franklin R W , Simpson J B , Ashwood E R
Published: January 2020

We describe a fluorescence polarization assay for use in predicting fetal lung maturity, which is suitable for the TDx Analyzer (an automated fluorescence polarimeter). The assay requires 0.5 mL of amniotic fluid and approximately 30 min, and involves the fluorophore 1-palmitoyl-2(6-[(7-nitro-2, 1, 3-benzoxadiazol-4-yl)amino]caproyl) phosphatidylcholine. Reproducible polarization measurements depend on proper regulation of incubation time and temperature, but variations in the concentrations of fluorescent probe and amniotic fluid have little effect on measured polarization and therefore little effect on assay precision. Working solutions of the fluorescent probe are stable for at least nine months when stored at -20 degrees C and pH 5. Interferences include erythrocytes, serum, bilirubin, meconium, and lidocaine.
Improved fluorescence polarization assay for use in evaluating fetal lung maturity. III. Retrospective clinical evaluation and comparison with the lecithin/sphingomyelin ratio.
Authors: Ashwood E R , Tait J F , Foerder C A , Franklin R W , Benedetti T J
Published: January 2020

We clinically evaluated, retrospectively, our improved fluorescence polarization assay for fetal lung maturity. The procedure requires 0.5 mL of amniotic fluid and a standard clinical laboratory fluorescence polarimeter (TDx Analyzer, Abbott Laboratories). We measured the L/S ratios for 93 freshly collected amniotic fluids, uncontaminated with blood or meconium, collected within three days of delivery. The fluids were stored frozen for eight to 32 months, then thawed and assayed for net fluorescence polarization. Fourteen of the infants developed respiratory distress syndrome; five, transient tachypnea of the newborn; and 74, no respiratory distress. The polarization assay and lecithin/sphingomyelin ratio had equivalent receiver operating characteristic curves, indicating no difference in their clinical performance. Although a prospective study with fresh amniotic fluid specimens will be necessary to establish a definitive reference range, the present study shows that this assay can be used to rapidly predict fetal lung maturity.
Apolipoprotein E quantified by enzyme-linked immunosorbent assay.
Authors: Bury J , Vercaemst R , Rosseneu M , Belpaire F
Published: January 2020

We developed a sensitive, specific "sandwich"-type enzyme-linked immunosorbent assay in which affinity-purified antibodies are used for coating and also for preparing an antibody-peroxidase conjugate to quantify apolipoprotein E in serum and in its lipoprotein fractions. This technique is rapid (results within 5 h), precise (mean intra- and interassay CVs 4.3 and 8.2%, respectively), accurate, and simple to perform. Sample pretreatment did not enhance apo E immunoreactivity. For 47 normolipidemic subjects, the mean apo E concentration was 38.11 (SD 11.1) mg/L. Above-normal apo E concentrations were measured in all types of hyperlipoproteinemia, especially types III and V. Apo E correlated with triglyceride (r = 0.58) and cholesterol concentrations (r = 0.60). As determined by gel filtration, hypertriglyceridemia was associated with a redistribution of apo E towards the triglyceride-rich lipoprotein fractions, which contained 77.1% (SD 16.8%) of total plasma apo E in Fredrickson type V patients, compared with 12.5% (SD 6.3%) in normal persons.
Creatine kinase: reassessment of optimal concentrations for adenosine-5'-diphosphate and magnesium.
Authors: Lahet C , Vialle A , Maire I , Parrin B , Steghens J P , Mathieu M
Published: January 2020

Whereas univariate studies led to an European agreement for the choice of optimal reagent concentrations of 2 mmol/L for ADP and 10 mmol/L for Mg2+ for determining creatine kinase (EC 2.7.3.2) activity in serum, whatever its isoenzyme pattern, the results of our bivariate study led us to recommend higher optimal concentrations: 4.1 to 4.7 mmol/L for ADP and 22 mmol/L for Mg2+. The zone of maximal activity was in fact a broad plateau such that more than 99% of maximal enzyme activity was attained at ADP concentrations between 3 and 5 mmol/L and Mg2+ concentrations between 17 and 26 mmol/L. Under these new conditions the maximum activity measured was modestly increased (about 10%) over the previously recommended method but the assay could be expected to be more resistant to the variations of ADP and Mg2+ concentrations. It may become necessary to modify the European recommended method.
Differentiating muscle damage from myocardial injury by means of the serum creatine kinase (CK) isoenzyme MB mass measurement/total CK activity ratio.
Authors: el Allaf M , Chapelle J P , el Allaf D , Adam A , Faymonville M E , Laurent P , Heusghem C
Published: January 2020

We immunoenzymometrically measured creatine kinase (CK) isoenzyme MB in extracts of myocardium and in homogenates of five different skeletal muscles. CK-MB concentrations in the former averaged 80.9 micrograms/g wet tissue; in the skeletal muscles it varied widely, being (e.g.) 25-fold greater in diaphragm than in psoas. CK-MB in skeletal muscles ranged from 0.9 to 44 ng/U of total CK; the mean for myocardium was 202 ng/U. In sera from 10 trauma and 36 burn patients without myocardial involvement, maximum ratios for CK-MB mass/total CK activity averaged 7 (SEM 1) ng/U and 18 (SEM 6) ng/U, respectively. Except for an infant (220 ng/U), the highest ratio we found for serum after muscular damage was 38 ng/U. In contrast, the mean maximum ratio determined in 23 cases of acute myocardial infarction exceeded 200 ng/U. Among seven determinations performed 8 to 32 h after onset of symptoms, each infarct patient demonstrated at least one ratio greater than or equal to 110 ng/U. Ratios observed after infarct were unrelated to treatment received during the acute phase. We propose a CK-MB/total CK ratio of 80 ng/U as the cutoff value for differentiating myocardial necrosis from muscular injury.
Laboratory tests in the differential diagnosis of hyperamylasemia.
Authors: Tietz N W , Huang W Y , Rauh D F , Shuey D F
Published: January 2020

We evaluated the clinical utility of some recently developed laboratory methods, including total amylase by three methods; isoamylase by inhibition and isoelectric focusing; lipase by pH-Stat and turbidimetry; and immunoreactive trypsin. All methods correlated highly positively with hyperamylasemia due to primary acute pancreatitis. Pancreatic-type isoamylase determinations have the greatest clinical usefulness, because total amylase, lipase, and immunoreactive trypsin are increased in a relatively high percentage of other abdominal diseases. Increases in the last-mentioned enzymes in nonpancreatic abdominal disease may be the result of injuries to the pancreas secondary to the primary disease, which are being detected with these highly sensitive methods. Because of the high clinical sensitivity of lipase and immunoreactive trypsin determinations, a normal result tends to exclude acute pancreatitis. Hyperamylasemia seen in lung carcinoma is due to increase in an amylase isoenzyme similar to the salivary-type amylase. The method for pancreatic-type isoamylase based on selective inhibition is satisfactory for routine clinical laboratory use.
Screening and quantification of hypnotic sedatives in serum by capillary gas chromatography with a nitrogen-phosphorus detector, and confirmation by capillary gas chromatography-mass spectrometry.
Authors: Soo V A , Bergert R J , Deutsch D G
Published: January 2020

We describe a quantitative screen for hypnotic-sedative drugs in which we use capillary gas chromatography with a nitrogen-phosphorus detector (GC/NPD) as the primary method and capillary gas chromatography-mass spectrometry (GC-MS) for confirmation. GC retention times of the acid-extracted underivatized drugs were stable (CVs less than 1%), and the detector response varied linearly over a 20-fold concentration range with a mean correlation coefficient for 11 drugs of 0.989. The limits of detection were satisfactory (0.5 mg/L in a 0.5-mL serum sample and 1-microL injection volume), as were precision (average CV 5.2% within day, 6.4% between day). The complementary use of capillary GC-MS not only unambiguously confirms presumptive peaks identified by GC, but also prevents reports of false positives and identifies compounds not included in the quantitative GC screen that may be listed in the GC-MS library.
Spectrophotometry of carnitine in biological fluids and tissue with a Cobas Bio centrifugal analyzer.
Authors: Cederblad G , Harper P , Lindgren K
Published: January 2020

A spectrophotometric method for carnitine has been adapted to the Cobas Bio centrifugal analyzer. The addition of carnitine to a system containing carnitine acetyltransferase (EC 2.3.1.7) and acetyl-CoA gives rise to the formation of CoA. The system is coupled to 5,5'-dithiobis(2-nitrobenzoate) (DTNB). Assay response varied linearly with concentration of carnitine over a wide concentration range. The total CV was 5.5% for a carnitine concentration in serum of 58.0 mumol/L. Analytical recovery of carnitine added to a serum sample was 93%. No interference was found in icteric, not grossly hemolyzed, lipemic, or uremic sera. Comparison with a radioenzymatic method showed that results correlated well (r greater than 0.965) but the present method gave values proportionally greater by 10 to 25% for samples of plasma, dialysis fluid, urine, and muscle tissue. Advantages over the original spectrophotometric assays involving DTNB include low reagent costs, rapidity, simplicity, and reproducibility. However, this modification is not as sensitive and probably not as specific as the radioenzymatic methods.
Clustering of serum calcium and magnesium concentrations in siblings.
Authors: Payne R B , Jones D P , Walker A P , Evans R T
Published: January 2020

Concentrations of both serum calcium, adjusted for albumin, and serum magnesium of siblings from 23 families were found to cluster around different concentrations within the normal reference interval. Variation between families accounted for 37% of the total variation in calcium and 28% of that in magnesium. A disturbance of serum calcium or magnesium homeostasis in an individual resulting in an altered serum concentration that remains within the reference interval might be recognized by examining values in siblings.
Is standardization more important than methodology for assay of total protein in cerebrospinal fluid?
Authors: Gerbaut L , Macart M
Published: January 2020

Four manual micromethods for protein determination, two turbidimetric (trichloroacetic and sulfosalicylic acid-sodium sulfate) and two colorimetric (Lowry and Coomassie Brilliant Blue--sodium dodecyl sulfate, CBB-SDS) were used to compare the standard curves for total protein (0.30 to 3 g/L) produced with three reference materials: bovine serum albumin, human serum albumin, and diluted human serum. We measured the apparent protein content of a sample of pooled human cerebrospinal fluid by all four methods and with use of all three standards. The only reference material that gave similar results with all four methods was diluted human serum; the CBB-SDS was the only method that gave identical results with all three reference materials. We then measured the protein concentration of 28 individual cerebrospinal fluid samples by the four methods, with diluted human serum as standard. Results by all methods correlated well, but only the sulfosalicylic acid and the CBB-SDS methods gave equivalent results. We conclude that the choice of standard is more important than the method used. However, the CBB-SDS method may be the preferred method because it produced identical standard curves with all three protein standards.
The alkaline reducing activity of glycated serum proteins and its relevance to diabetes mellitus.
Authors: Johnson R N , Baker J R
Published: January 2020

We investigated the ability of the fructosamine assay to detect glycation of serum proteins. We incubated both whole human serum and serum protein fractions in vitro with [14C]glucose, and analyzed for reducing activity and for uptake of 14C by protein. In all experiments, the reducing activity increased linearly with time for seven days and was correlated with 14C uptake (r = 0.94-0.98). Protein ketoamines were about fivefold more actively reducing than equimolar concentrations of deoxymorpholinofructose, the fructosamine standard, which explains why values for fructosamine in serum are higher than the expected concentration of protein ketoamines. We also used [14C, 2-3H]glucose to assess the contribution of the aldimine component to 14C uptake. Whole human serum and albumin incubated with [14C, 2-3H]glucose showed little uptake of 3H in relation to 14C. We conclude that glycated protein can be simply and reliably quantified by the fructosamine assay, and we discuss the relevance of this conclusion to the monitoring of diabetes.
Relation between plasma creatinine and body size.
Authors: Swaminathan R , Ho C S , Chu L M , Donnan S
Published: January 2020

We studied the relationship between creatinine concentration in plasma and body size in 673 healthy individuals (385 women, 288 men). Weight, height, and skinfold thickness were measured and blood was sampled (after a short fast) for estimations of urea and creatinine in plasma. Lean body mass was calculated according to Durnin and Wormsley (Br J Nutr 1974; 32:77-97). Concentrations of urea and creatinine in plasma were significantly lower in women than in men and were significantly correlated with age in both sexes. For neither sex was there a significant correlation between lean body mass and creatinine concentration, even after we took age into account either by multiple linear-regression analysis or by considering subjects at decade intervals.
Liquid-chromatographic analysis for esmolol and its major metabolite in urine.
Authors: Achari R , Drissel D , Hulse J D
Published: January 2020

We describe a simple, reproducible liquid-chromatographic method for determination of esmolol (a short-acting beta blocker) and its major metabolite in human urine. Esmolol is extracted from urine at a pH of 8.4 into methylene chloride; the more polar metabolite remains in the aqueous phase. We then measure esmolol with a muBondapak C18 column and measure ultraviolet absorbance at 229 nm; the metabolite is analyzed with a Spherisorb phenyl column, with absorbance measured at 280 nm. The average extraction recoveries of esmolol and the metabolite were 95 and 92%, respectively. Standard curves were linear and reproducible for esmolol from 0.025 to 5 mg/L and for the metabolite from 1 to 250 mg/L. Within-day CVs for both compounds were less than 6%.