bluuebunny/crossref_metadata_2025 huggingface dataset:
An improved procedure for detection of hypoxanthine--guanine phosphoribosyl transferase heterozygotes.
Authors: Page T , Bakay B , Nyhan W L
Published: January 2020

The absence of activity of the enzyme hypoxanthine-guanine phosphoribosyltransferase (EC 2.4.2.8) is known to be the cause of the Lesch-Nyhan syndrome. Previous methods for detection of heterozygous carriers of this genetic defect either are quite time consuming, require specialized equipment, or lack the necessary sensitivity. We present here a method in which thin-layer chromatography and autoradiography are used to assay the activity of this enzyme in individual hair roots collected from the scalp of the possible carrier. This method is fast and sensitive, and requires no specialized equipment.
Use of glycerol-cryoprotected Lactobacillus casei for microbiological assay of folic acid.
Authors: Wilson S D , Horne D W
Published: January 2020

A simple procedure for preparing glycerol-cryoprotected Lactobacillus casei cultures has been developed. L. casei grown in medium supplemented with low concentrations of folic acid (0.3 micrograms/L) is diluted with an equal volume of glycerol (800 mL/L) and stored at -20 degrees C. Growth response of the glycerol-cryoprotected L. casei to low concentrations of folic acid exceeded that of cultures maintained by monthly agar stab transfer. Also, growth for the zero-folate blanks was considerably less for the cryoprotected cultures. Assay of folate in several rat tissues correlated well (r = 0.999) with the standard microbiological assay. The growth rate of the culture depends on the inoculum size, and a heavy inoculum of cryoprotected L. casei may be used to complete the assay after only an overnight incubation.
Improved liquid-chromatography of aspirin, salicylate, and salicyluric acid in plasma, with a modification for determining aspirin metabolites in urine.
Authors: Buskin J N , Upton R A , Williams R L
Published: January 2020

Detailed performance specifications are given for a specific and sensitive liquid-chromatographic assay for aspirin, salicylic acid, and salicyluric acid in plasma. The sensitivity of this method for aspirin (50 micrograms/L) is 10-fold that of previous methods, so that concentrations of aspirin in plasma can now be followed for about four or five half-lives after the peak plasma concentration arising from a single 650-mg dose. In addition, this assay is as sensitive for salicylic and salicyluric acid in plasma as any hitherto. A modification permits measurement of gentisic acid, salicylic acid, and salicyluric acid in urine; further modifications allow indirect measurement of conjugated gentisate and salicylate in urine.
Identification and metabolic implication of 1-deoxyglucose (1,5-anhydroglucitol) in human plasma.
Authors: Yoshioka S , Saitoh S , Fujisawa T , Fujimori A , Takatani O , Funabashi M
Published: January 2020

1-Deoxyglucose (1,5-anhydroglucitol), a metabolite related to diabetes mellitus, was identified in human plasma by gas-liquid chromatography and by gas-liquid chromatography/mass spectrometry. Plasma polyols were accurately determined with a gas-liquid chromatograph equipped with an all-glass capillary column. The plasma content of 1-deoxyglucose in healthy persons varies with age. Although the precise physiological role of 1-deoxyglucose remains obscure, the method described here for determining the minor polyol components of plasma, as well as the findings of 1-deoxyglucose in the plasma of healthy subjects, may be useful for investigating the metabolic roles of 1-deoxyglucose.
Thermal-ionization isotope-dilution mass spectrometry as a definitive method for determination of potassium in serum.
Authors: Gramlich J W , Machlan L A , Brletic K A , Kelly W R
Published: January 2020

Thermal-ionization isotope-dilution mass spectrometry is a highly precise and accurate method for the determination of potassium concentrations in serum. Although not suited for routine use because of the time and expense required, the technique provides an extremely valuable tool for the characterization of reference materials and for evaluating other analytical methods. The technique has recently been used to determine the concentration of potassium in a human serum standard, NBS Standard Reference Material 909. Seven vials of the serum were chemically processed and then analyzed by two spectroscopists independently, using different mass spectrometers. The results confirm previous work that indicates that a precision of 0.1% relative can be routinely achieved. The systematic errors in the method have been thoroughly evaluated. When the precise results are thus corrected, they are essentially bias free and hence definitive.
Direct 125I-radioligand assays for serum progesterone compared with assays involving extraction of serum.
Authors: Ratcliffe W A , Corrie J E , Dalziel A H , Macpherson J S
Published: January 2020

We compared two direct radioimmunoassays for progesterone in 50 microL of unextracted serum or plasma with assays involving extraction of serum. The direct assays include the use of either danazol at pH 7.4 or 8-anilino-1-naphthalenesulfonic acid at pH 4.0 to displace progesterone from serum binding-proteins. Progesterone is then assayed by using an antiserum to a progesterone 11 alpha hemisuccinyl conjugate and the radioligand 125I-labeled progesterone 11 alpha-glucuronyl tyramine, with separation by double-antibody techniques. Direct assays with either displacing agent gave good analytical recovery of progesterone added to human serum, and progesterone values for patients' specimens correlated well (r greater than 0.96) with results of assays involving extraction of serum. Precision was similar with each displacing agent over the working range 2.5-100 nmol/L and superior to that of extraction assays. We conclude that these direct assays of progesterone are analytically valid and more robust, precise, and technically convenient than many conventional methods involving extraction of serum.
Improved assessment of plasma lipoprotein patterns. IV. Simple preparation of a lyophilized control serum containing intact human plasma lipoproteins.
Authors: Wieland H , Seidel D
Published: January 2020

Addition of sucrose (413-825 mmol/L) to human serum allows plasma lipoproteins to be lyophilized without denaturation. A control serum so prepared is especially suited for use in monitoring determinations of apolipoproteins and quantitative lipoprotein electrophoresis. It is clear upon reconstitution; hence, the described procedure may also be useful for preparation of control sera for general clinical chemistry without de-lipoproteinization.
Effects of Sample Freezing on Ion-Selective Electrode Determinations of Serum Calcium
Authors: Plant Sandford B , McCarron David A
Published: January 2020

We assessed in a prospective study the effects of freezing, thawing, and prolonged storage of serum samples for ionized calcium determinations with the "Clin ± Ion." Forty-eight serum samples were collected before and during infusions of EDTA and calcium into human subjects. Values for ionized calcium in serum samples stored frozen for one and six months were similar to those determined for fresh specimens. Samples stored for 12 months had significantly (p <0.001) lower values. Thawing and refreezing the sample within the first six months did not alter values. We conclude that serum samples can be frozen and stored for as long as six months without affecting results for ionized calcium determinations.
Fluoroimmunoassay of digitoxin in serum.
Authors: Al-Hakiem M H , Simon M , Mahmod S , Landon J
Published: January 2020

This fluoroimmunoassay for digitoxin in serum involves use of a sheep antiserum to digitoxin coupled to magnetizable solid-phase particles and fluorescein-labeled 3-O-succinyl digitoxigenin as tracer. Sodium salicylate blocks binding of the drug by binding proteins, and endogenous fluorophores and other interfering components in serum samples are reliably and completely removed at the separation and wash steps, which are facilitated by magnetic sedimentation. The method is sufficiently sensitive, precise, and specific for application to routine monitoring of digitoxin therapy, and results correlate closely (r = 0.992) with those of an established radioimmunoassay.
Effect of anti-thyroxin autoantibodies on radioimmunoassay of free thyroxin in serum.
Authors: Konishi J , Iida Y , Kousaka T , Ikekubo K , Nakagawa T , Torizuka K
Published: January 2020

Serum thyroxin was nearly or completely undetectable by radioimmunoassay in an elderly patient with Graves' disease being treated with methimazole. Abnormal binding of thyroxin to antibodies of the IgG variety was shown, the association constant of the complex being 8.0 x 10(8) L/mol and the binding capacity 6.3 nmol/g of IgG. The effect of the antibody on results of radioimmunoassay of free thyroxin was studied with three commercial kits, two of which (Clinical Assays and Damon Diagnostics) gave essentially the same values as did equilibrium dialysis. The third (Amersham International) gave falsely high results because the 125I-labeled thyroxin derivative used in the kit was bound by the autoantibody.
Flameless atomic absorption spectrophotometry of selenium in whole blood.
Authors: Tulley R T , Lehmann H P
Published: January 2020

In this method of analysis for selenium in whole blood by flameless atomic absorption spectrophotometry, 1 mL of sample is first digested with perchloric and nitric acids. After reduction and neutralization, the sample is reacted with 2,3-diaminonaphthalene, and the product is extracted into toluene. Twenty microliters of the extract is injected into the graphite furnace of a flameless atomic absorption spectrophotometer, along with 20 microliters of a 1.0 g/L cupric nitrate solution. Blood-based standards are used to establish the standard curve. The amount of selenium required to give an absorbance of 0.0044 is 5.3 micrograms/L. Precision is good, recovery excellent. The extract is stable for 24 h.
Simple, reliable chromatographic measurement of oxalate in urine.
Authors: Di Corcia A , Samperi R , Vinci G , D'Ascenzo G
Published: January 2020

In this assay for oxalate in urine, oxalate is adsorbed from the urine onto graphitized carbon black (Carbopack B). After desorption and removal of the solvent, oxalic acid is gas-chromatographically measured after being derivatized with BF3/methanol. Chromatography is on Carbopack B/polyethylene glycol (Mr 20000), 93.7/6.3 by weight. The lower limit of detection of urinary oxalate is about 3 mg/L (CV less than or equal to 3.6%). A series of oxalate determinations in 24-h urine samples of 15 subjects gave a mean of 47.1 (SD 15.4) mg/24 h, with an analytical recovery of 97.4% (SD 3.0%, range 93.6-102.4%). Total analysis time for one sample is about 2 h.
Determinations of amylase isoenzymes in serum by use of a selective inhibitor.
Authors: Huang W Y , Tietz N W
Published: January 2020

A simple, rapid screening procedure for determining the relative amounts of pancreatic (P)- and salivary (S)-type amylase in serum is presented, involving incorporation of a selective inhibitor (from wheat-germ) into commercially available BMD Single-Vial Amylase and Beckman Enzymatic Amylase-DS procedures for manual and automated isoamylase measurements. Optimal concentrations of inhibitor inhibit the S-type amylase fraction by 87-88%. In contrast, the pancreatic fraction is inhibited by only 19% in either the manual or automated methods. The degree of inhibition is constant for amylase activities up to at least 520 U/L. Use of the ratio (P-amylase/total amylase activity) X 100 helps differentiate between hyperamylasemia caused by S-type or P-type amylase. In preliminary studies, patients with pancreatitis showed a ratio greater than 70%.
Enzymic and chemical-extraction determinations of free fatty acids in serum compared.
Authors: Demacker P N , Hijmans A G , Jansen A P
Published: January 2020

We compared an enzymic test kit for determination of free fatty acids in serum (NEFA C-test, WAKO) with two modifications of a chemical extraction procedure, I (Clin. Chim. Acta 43: 317-320, 1973) and II (Clin. Chim. Acta 80: 327-332, 1977). All three procedures are specific for long- and medium-chain fatty acids. Short-chain fatty acids, some keto acids, and phospholipids did not interfere. Added fatty acid was quantitatively accounted for in all methods. Results obtained with the enzymic method and II did not differ significantly, whereas the results by I were about 10% lower. The concentration of NaCl in the copper reagent, but not the kind of solvent used to dissolve the standard, influenced the accuracy of the chemical methods. In the enzymic procedure, hydrolysis of triglycerides during incubation is unlikely to be the reason for too-high values. The precision of all three procedures is acceptable for use in clinical laboratories.
Evaluation of three minicolumn procedures for measuring hemoglobin A1.
Authors: Hammons G T , Junger K , McDonald J M , Ladenson J H
Published: January 2020

We have evaluated three commercially available column-chromatographic methods (Isolab, Helena, and Bio-Rad) for the determination of "fast" hemoglobin (HbA1). All three methods correlated with HbA1c measurements by "high-performance" liquid chromatography for 121 samples from diabetic patients, with the Isolab method showing the highest correlation (r = 0.921). The Isolab and Helena methods gave results that were linear with proportions of HbA1 as great as 30%; results by the Bio-Rad method were slightly nonlinear at values greater than 15%. The Isolab method showed better within- and between-assay precision (CV) than the other two methods and was considered the simplest to perform by each of four different technologists. We recommend use of the Isolab method over the other two tested and believe that this procedure will be valuable for monitoring long-term glycemic control in diabetic patients.
Methylmalonic acidemia with the unusual complication of severe hyperglycemia.
Authors: Boeckx R L , Hicks J M
Published: January 2020

We describe a case of neonatal methylmalonic acidemia with the unusual complication of severe, insulin-resistant hyperglycemia. Methylmalonic acidemia, an inherited metabolic disease affecting the catabolism of propionic acid, is manifested by persistent metabolic acidosis, urinary excretion of large amounts of methylmalonic acid, and occasionally by hypoglycemia. Severe and persistent metabolic acidosis and hyperglycemia, despite large doses of insulin, were observed in this infant, who excreted large amounts of methylmalonic acid. The diagnosis of methylmalonic acidemia was confirmed by gas chromatography-mass spectroscopy, but the patient died before the defect in glucose tolerance could be delineated. We hypothesize that, in addition to the methylmalonic acidemia, the patient may have had an insulin-receptor defect, which was manifested as an inappropriate response to endogenous and exogenous insulin.
Organic electrochemical techniques having potential clinical application.
Authors: Durst R A , Blubaugh E A , Bunding K A , Fultz M L , MacCrehan W A , Yap W T
Published: January 2020

The Organic Electrochemistry Group at the National Bureau of Standards is pursuing several avenues of research of potential application to problems of clinical chemists. With one development, electrochemical detectors for liquid chromatography, organomercury species can be determined in biological tissues and other matrices. Spectroelectrochemistry is being used to characterize the redox behavior of metal complexes of bleomycin, an antitumor drug. Chemically modified electrodes are being developed as selective electrocatalytic sensors for organohalogen compounds and may lead to new sensors for clinically important analytes. Surface-enhanced Raman spectroscopy is helping characterize the polymer films used to modify the electrode surfaces. Another sensor is being developed for the detection of carboxylic acids: after the photocatalytic oxidation of the acids at a semiconductor electrode, the carbon dioxide produced is subsequently determined with a flow-through gas-sensing electrode. Finally, mathematical modeling may provide a better understanding of the fundamental processes involved in several of the above techniques.
Polymer-membrane electrode-based potentiometric sensing of ammonia and carbon dioxide in physiological fluids.
Authors: Meyerhoff M E , Fraticelli Y M , Greenberg J A , Rosen J , Parks S J , Opdycke W N
Published: January 2020

Several new electrochemical devices designed for detecting ammonia and carbon dioxide in physiological samples have in common the elimination of the fragile glass pH internal electrode used in the conventional Severinghaus-design potentiometric gas sensors. In one new approach, the pH glass electrode can be replaced with a neutral carrier-based polymer-membrane pH electrode. Alternatively, to improve response characteristics and selectivity, we have used ion-selective polymer membranes responsive to ammonium and carbonate as internal sensing elements in conjunction with appropriate internal buffer reagents. The use of inner polymer membranes rather than glass membranes enables the fabrication of essentially disposable devices having rapid response times. In addition, we have constructed automated continuous-flow NH3- and CO2-sensing systems involving novel tubular forms of the polymer-membrane electrodes and a simple flow-through gas dialysis arrangement. The polymer electrode-based systems described offer many unique and practical advantages over existing gas-sensing units.
Effect of temperature on the p50 value for human blood.
Authors: Samaja M , Melotti D , Rovida E , Rossi-Bernardi L
Published: January 2020

We investigated the effect of temperature (19, 30, 37, and 43 degrees C) on the p50 value for normal human blood at pco2 = 5.72 kPa (43 mmHg), at various pHs (range 7.0 to 7.6) and molar ratios of [2,3-diphosphoglycerate]/[Hb4] (range 0.4 to 2.4). The d(log p50)/d(pH) coefficient varied from 0.39 at 19 degrees C to 0.35 at 43 degrees C. The relationship between log p50 and 1/T (T = degrees Kelvin) was linear under the experimental conditions used, and the d(log p50)/d(1/T) coefficient varied between -2138 at pH 7.0 and -2162 at pH 7.6, independent of the concentration of 2,3-diphosphoglycerate. Assuming that the effect of pco2 on the p50 value is the same at 19, 30, and 43 degrees C as at 37 degrees C, one can use the reported coefficients to calculate the p50 value for normal human blood under conditions of temperature, pH, pco2, and 2,3-diphosphoglycerate concentrations prevailing under physiological and pathological conditions. The p50 value calculated by empirical equations, taking into account the effect of temperature, correlated well with the values for p50 determined experimentally (y = 0.9774x + 0.453; r = 0.998; n = 60), with an SD of 52 Pa (0.39 mmHg).
Evaluation of a colorimetric method for determination of glycosylated hemoglobin.
Authors: Standefer J C , Eaton R P
Published: January 2020

We evaluated a colorimetric assay for glycosylated hemoglobin to determine the effects of several variables --oxalic acid concentration, extraneous glucose, hemoglobin concentration, hydrolysis interval, and 5-hydroxymethylfurfural destruction--and the precision. The interference seen when the blood glucose concentration exceeds 2.0 g/L (11 mmol/L) can be eliminated by washing the erythrocytes with 9 g/L saline. The accuracy of this assay is not influenced by hemoglobin concentrations from 80 to 150 g/L. The background nonspecific color, although substantial, is similar from sample to sample. After a 5-h hydrolysis at 100 degrees C, about 85% of the hexose is released, and the analytical recovery of 5-hydroxymethylfurfural is constant over a wide range of glycosylated hemoglobin concentrations. The 5th to 95th percentile reference interval for a population of 65 nondiabetic individuals was 4.6 to 6.1 mol per 100 mol of total hemoglobin. The range of values for a population of 85 diabetic patients was 6.9 to 20.4 mol per 100 mol.
Measurement of nifedipine in plasma by gas--liquid chromatography and electron-capture detection.
Authors: Hamann S R , McAllister R G
Published: January 2020

In this method for measuring nifedipine, a calcium-channel inhibitor now widely used in the treatment of cardiovascular disease, the drug is extracted from plasma under basic conditions into toluene, and an aliquot of the extract is injected directly into a gas chromatograph equipped with an OV-101 column and an electron-capture detector. The standard curve is linear between 1 and 100 micrograms/L; the assay measures not only nifedipine, but also a major metabolic product (found only after oral administration of the parent drug) and photodegradation products. The procedure is rapid and adequately sensitive for routine use in clinical monitoring of nifedipine in plasma.
Solid-phase radioimmunoassay for human placental lactogen in serum.
Authors: Samuel G , Sivaprasad N , Shah K B , Mani R S
Published: January 2020

We report a solid-phase radioimmunoassay for human placental lactogen, in which antibody-coated polystyrene tubes and polyurethane sponges are used. Incubation time and temperature, ionic strength of buffer, and nonspecific adsorption have been evaluated for optimum conditions required for routine clinical assays. The validity of the assay technique has been ascertained by its specificity, reproducibility, and recovery. The sensitivity of this method is 0.4 ng of placental lactogen per assay tube.
Cross reactivity of the EMIT digoxin assay with digoxin metabolites, and validation of the method for measurement of urinary digoxin.
Authors: Linday L , Drayer D E
Published: January 2020

In evaluating the EMIT (Syva Co.) digoxin assay, we found no cross reactivity between dihydrodigoxin, the major digoxin metabolite, and the EMIT digoxin antibody from two different lots. However, the antibody does cross react, essentially completely, with the digoxin hydrolysis metabolites digoxigenin, digoxigenin mono-digitoxide, and digoxigenin bis-digitoxide. The EMIT method can be used to measure digoxin in urine diluted at least 50- to 100-fold with digoxin-free human plasma; the inter-assay coefficient of variation of this assay is 6%. In addition, we validated the manual EMIT serum digoxin assay, using external ("TRI-rac") quality controls.
Frequency of hypomagnesemia in hypokalemic states.
Authors: Boyd J C , Bruns D E , Wills M R
Published: January 2020

Although both hypokalemia and hypomagnesemia have been linked with life-threatening cardiac arrhythmias, published studies disagree regarding the frequency of simultaneously low serum potassium and magnesium concentrations. To investigate the possible associations between the concentrations of these ions in serum, we carried out a retrospective study of all paired magnesium-potassium measurements reported from our laboratory during two separate two-week periods. Of 963 paired measurements in 421 patients, 12% of the samples were hypokalemic and 26% were hypomagnesemic. The frequency of hypomagnesemia was significantly greater in hypokalemic samples (38%) compared with nonhypokalemic samples (25%). The potential clinical importance of this relationship requires further investigation.
Macro creatine kinase types 1 and 2: clinical significance in neonates and children as compared with adults.
Authors: Wu A H , Herson V C , Bowers G N
Published: January 2020

We examined the case histories of seven pediatric patients (ages four days to 12 years) whose sera showed significant activities of "macro" creatine kinase (CK; EC 2.7.3.2): two cases of macro CK type 1 (CK-BB isoenzyme bound to IgG) and five cases of macro CK type 2 (polymeric complexes of mitochondrial CK). The diagnoses of the two children with macro CK type 1 were similar to those in 55 adult patients with this form of CK found earlier during screening of a population of 5000. Macro CK type 1 has generally been noted only in older women; this is the first report of it in children. In four of the five cases of macro CK type 2 we found in children, myocardial muscle damage was clinically evident. In contrast, the 26 cases of adults with macro CK type 2 found during the same screening generally had neoplasms, most with metastases to the liver. We suggest that the finding of macro CK type 2 is an indicator of cellular necrosis; in pediatric cases with myocardial damage the associated conditions may be reversible but in adults with malignancies the finding is associated with an extremely poor prognosis.
Falsely high serum creatinine concentration associated with severe methanol intoxication.
Authors: WU A H , Stout R , McComb R B
Published: January 2020

A case of severe methanol intoxication (1300 mg/L) was associated with markedly increased serum creatinine (490 mg/L) despite normal urea values and the absence of any other signs of renal disease. These values declined progressively to normal, and the patient recovered with no visual impairment. Additional laboratory experimentation suggested that the high creatinine value was probably ascribable to some unknown foreign material(s) in the patient's blood that reacted with the alkaline picrate used in the measurement of creatinine. One of the presumed metabolites of methanol, formaldehyde, reacts with creatinine but the product does not react with picrate. We believe that the foreign material was derived from either commercial preparations of methanol or contaminants in the patient's drinking water.
On the calculation of a "reference change" for comparing two consecutive measurements.
Authors: Harris E K , Yasaka T
Published: January 2020

We describe a statistical method for calculating a "reference change," defined as that difference between two consecutive test results in an individual that is statistically significant in a given proportion of all similar persons. By allowing for variation in within-person variances, this procedure computes a reference change that is more specific (i.e., less prone to false positives) than that obtained directly from the distribution of observed differences between measurements. Moreover, the method may easily be extended to a test for trend in three successive measurements. The method has been applied to semi-annual measurements of serum calcium and alkaline phosphatase in 698 men and women enrolled in a large health-maintenance program. We believe that these ideas may also be usefully applied to successive laboratory tests in carefully defined patient populations--but this introduces special problems, which are discussed briefly.
A diazo-based dry film for determination of total bilirubin in serum.
Authors: Dappen G M , Sundberg M W , Wu T W , Babb B E , Schaeffer J R
Published: January 2020

We have prepared a diazo-based dry film for use in determining total serum bilirubin. On a transparent support are a buffered gelatin layer containing a polymeric quaternary amine (the mordant) and a white, reflective spreading layer that contains all of the components necessary for detection of bilirubin. The method is based on the use of dyphylline and Triton X-100 surfactant to dissociate bilirubin from albumin and subsequent reaction of bilirubin with a diazonium salt [4-(N-carboxymethylsulfamyl)benzenediazonium hexafluorophosphate]. In the dry film, unconjugated, mono- and diconjugated, and strongly protein-linked (delta) bilirubin all react with the diazonium salt to produce azo dyes having absorption maxima at about 520 nm. With reflection densitometry and appropriate mathematical transformation, readings and bilirubin concentrations are linearly related to 260 mg/L. Results correlate well with those by the Jendrassik-Grof (Doumas modification) method (slope 0.994, intercept 1.1, correlation coefficient 0.993, Sy X x 4.0), and the method is precise (CV = 10.0% at Cav = 4.1 mg/L, 2.7% at Cav = 24.5 mg/L, 1.2% at Cav = 102 mg/L for patients' samples) and relatively free of interferences.
Time-resolved fluorometer for lanthanide chelates--a new generation of nonisotopic immunoassays.
Authors: Soini E , Kojola H
Published: January 2020

Pulsed-light time-resolved fluorometry of lanthanide chelates has proved to be very sensitive for use with nonisotopic immunoassays. We describe a manually operated fluorometer with a conventional xenon flashtube. Sensitivity for 1-s determinations is similar to that of radioisotopic methods.
Hormones in saliva: mode of entry and consequent implications for clinical interpretation.
Authors: Vining R F , McGinley R A , Symons R G
Published: January 2020

Assay of hormones in saliva would be more convenient than assay in blood, but there is no information on the route by which hormones enter saliva, information that would provide insight into the clinical value of such assays. We have examined the mode of entry of various hormones into saliva. The results suggest that unconjugated steroids enter saliva by diffusing through the cells of the salivary glands and that their concentration in saliva does not depend on the rate of saliva production. Conjugated steroids enter saliva via "ultrafiltration" through the tight junctions between the acinar cells, and their concentration in saliva is highly flow-rate dependent. Thyroxin and choriogonadotropin enter saliva via the ultrafiltration route or by contamination of the saliva by plasma or gingival fluid. We conclude that the salivary concentration of unconjugated steroids may usefully reflect the concentration of free (nonprotein-bound) steroids in plasma. Conversely, the concentration of conjugated steroids, thyroxin, and protein hormones such as choriogonadotropin in saliva probably does not reflect their concentration in plasma in any clinically useful way.
Liquid-chromatographic assay and identification of mono- and diester conjugates of bilirubin in normal serum.
Authors: Muraca M , Blanckaert N
Published: January 2020

This liquid-chromatographic procedure for determining bilirubin mono- and diester conjugates in normal serum is based on pre-analysis conversion of bilirubin monosugar and disugar conjugates to the corresponding methyl esters by alkaline methanolysis. Here, extracted unconjugated bilirubin, bilirubin monomethyl esters, bilirubin dimethyl ester, and internal standard are separated on a reversed-phase column within 15 min, detected in the effluent at 436 nm, and quantified from their peak areas. Carotenoids do not interfere. Within-day and day-to-day CVs range from 5 to 13%. The smallest concentrations of monoconjugated and diconjugated pigments that are detectable and measurable are about 10 and 20 nmol/L, respectively. Such data are given for sera from 43 healthy adults. Total bilirubin concentrations in serum tended to be lower in women than in men, but the relative amounts of the various bilirubin fractions in sera from men and women were comparable. Analysis of ethyl anthranilate azoderivatives from serum permitted identification of the bilirubin ester conjugates in normal serum as bilirubin 1-O-acyl glucuronides.
Free thyroxin by radioimmunoassay: evaluation of a new direct method involving a radiolabeled thyroxin analog.
Authors: Kubasik N P , Lundberg P A , Brodows R G , Hallauer G D , Same D G , Lindstedt G , Bengtsson C , Nyström E
Published: January 2020

We present here the first performance evaluation of a new direct method for free thyroxin (T4) in serum by radioimmunoassay, with use of coated tubes and a radiolabeled T4 analog (Diagnostic Products Corp.). The assay is precise and robust: within-run imprecision (CV), 3.1-6.6%; between-run imprecision, 4.0-7.9%; no demonstrable variation between technologists irrespective of experience with the method. No outliers were observed when we compared the free T4 results with serum total T4. Reference values are reported for a total of 1243 euthyroid subjects; there was no significant age effect on serum free T4 in women 26 to 72 years old. The biological variation was about +/- 35% of the mean (2 SD). Free T4 results are the same for serum and plasma. The assay performs well in hypothyroidism and hyperthyroidism, and distinguishes individuals with thyroid disease from normal individuals. Free T4 values in women taking oral contraceptives are normal. Depressed results were often observed in acute nonthyroidal illness and continuing pregnancy. These results were directly comparable with those of another commercial direct radiolabeled-T4 analog kit for free T4.
An equilibrium model of the oxygen association curve of normal human erythrocytes under standardized conditions.
Authors: Ryall R G , Story C J
Published: January 2020

Hemoglobin-oxygen association curves of human erythrocytes were measured in metabolically stable cells under the standardized conditions of extracellular pH 7.400, carbon dioxide tension 38 mmHg (approximately 5 kPa) and temperature 37 degrees C. A model of the oxygen association curve of normal erythrocytes was subsequently developed, based on assumed equilibrium reactions between hemoglobins (Hb), oxygen, and 2,3-diphosphoglycerate (DPG). Conditional equilibrium constants were determined for reactions leading to the formation of species designated Hb-O2 (K = 0.14 mmHg-1), Hb-(O2)4 (K = 0.44 X 10(-4) mmHg-4), Hb-DPG (K = 0.30 X 10(4) L mol-1) and Hb-O2-DPG (K = 0.61 X 10(2) L mol-1 mmHg-1) for unmodified hemoglobin, and Hb-O2 (K = 0.14 mmHg-1) and Hb-(O2)4 (K = 0.44 X 10(-4) mmHg-4) for glycosylated hemoglobin. This model provides a practical means of calculating the expected oxygen association curve for a sample of erythrocytes under standardized conditions, given the intracellular concentrations of hemoglobins and 2,3-diphosphoglycerate and the assumption that the hemoglobins in those cells function normally.
Temperature correction of blood-gas and pH measurements.
Authors: Ashwood E R , Kost G , Kenny M
Published: January 2020

We critically review formulas for temperature correction of pH, pCO2, and pO2 measurements in whole blood and the clinical usefulness of these formulas. We discuss both the theoretical derivation and experimental verification of temperature-induced changes. We recommend when to use and when not to use these formulas, based upon the clinical interpretation of these assays.
Monoclonal antibodies to human plasma low-density lipoproteins. I. Enhanced binding of 125I-labeled low-density lipoproteins by combined use of two monoclonal antibodies.
Authors: Mao S J , Patton J G , Badimon J J , Kottke B A , Alley M C , Cardin A D
Published: January 2020

Four monoclonal antibodies (IgG2b) to human plasma low-density lipoproteins (LDL) have been characterized. The binding affinities of each monoclonal antibody to 125I-labeled LDL were moderately high, ranging from 10(8) to 10(10) L/mol at 4 degrees C, but were reduced by at least 50-70% at 37 degrees C. The maximum binding of each monoclonal antibody was unique, ranging from 20 to 95% of total 125I-labeled LDL, suggesting that LDL particles were immunochemically heterogeneous. One antibody, LP-34, had both high and low binding affinities to LDL. Another, LP-47, exhibited high affinity for isolated LDL, yet reacted poorly with native LDL in plasma, indicating that the conformation of isolated LDL differs from that of native LDL in plasma. Unlike polyclonal serum antibodies, a mixture of four monoclonal antibodies failed to precipitate LDL, but did show a drastic increase in binding to LDL. We found that only two of our monoclonal antibodies were necessary for such synergistic enhancement. We propose that one of the monoclonal antibodies may serve as a catalytic reagent, and discuss the clinical significance of this finding.
Evaluation of accuracy of 20 different test kits for the enzymic determination of cholesterol.
Authors: Demacker P N , Boerma G J , Baadenhuijsen H , van Strik R , Leijnse B , Jansen A P
Published: January 2020

We evaluated the accuracy of 20 different test kits for the enzymic determination of cholesterol. Using a selected set of standards, we found mean values differing by -9.9 to + 10.7% from values obtained by the Abell-Kendall procedure. Nine kits gave values with a relative bias of less than 2.5%. In the case of the other 11 kits, recalculation with secondary serum standards resulted in values within 5% of the reference values. Depending on the kit used, two different commercially available standard solutions produced results differing by, at most, 8.7%. We obtained indirect evidence that use of esterase of bacterial origin is associated with low values. We ascribe the high values obtained with some kits to positive interference by stabilizers or unspecified compounds in the reagents.
Evaluation of the Coomassie Brilliant Blue G-250 method for urinary protein.
Authors: Lott J A , Stephan V A , Pritchard K A
Published: January 2020

The Coomassie Brilliant Blue G-250 method for protein in urine has been evaluated for analytical accuracy and clinical applicability. Extremely simple to perform, the test exhibits good precision and sensitivity. The color developed per gram of protein is protein-type dependent, so no single protein standard is completely satisfactory. Color intensity is linearly related to concentration up to 1500 mg/L when used with a manual bichromatic method. Accuracy was clinically acceptable for patients with a variety of protein-losing diseases, and for patients having received renal transplants; however, the method underestimates urinary light-chain proteins. In athletes and premature neonates, we observed increased protein excretion during periods of stress. The upper reference limit for protein excretion in healthy adults is about 120 mg/24 h.
Cystic fibrosis--its biochemical detection.
Authors: Heeley A F , Watson D
Published: January 2020

We examine critically the biochemical methods capable of detecting and monitoring the end-organ disease processes in patients with cystic fibrosis. Although the diagnosis of cystic fibrosis is never justified on the basis of the sweat salt test alone, the original filter-paper technic (Gibson-Cooke, Pediatrics 23:545-549, 1959) for determining Na+ and Cl- concentrations in sweat remains the most discriminating method. We discuss the contributions for neonatal screening of the so-called cystic fibrosis protein, associated decreased enzymic activities in the homo- and heterozygous state, and immunoreactive trypsin. Because evidence of either intestinal malabsorption or a pancreatic lesion must be sought, we review the use and interpretation of some tests of pancreatic dysfunction (meconium albumin, duodenal juice components, serum pancreatic isoamylase, and trypsinogen), both in establishing and in confirming the diagnosis of cystic fibrosis.
Quantification of high-density-lipoprotein cholesterol by precipitation with phosphotungstic acid/MgCl2.
Authors: Assmann G , Schriewer H , Schmitz G , Hägele E O
Published: January 2020

We evaluated the use of a modified phosphotungstic acid/MgCl2 precipitation procedure for the precipitation of apolipoprotein B-containing lipoproteins. Precipitation of these lipoproteins [very-low- and low-density lipoproteins, and lipoprotein (a)] is complete, with negligible coprecipitation of high-density lipoprotein subfractions (HDL1, HDL2, HDL3), even in hypertriglyceridemic sera. In comparison with ultracentrifugation, the precipitation method yields, on the average, values that are 0.17 mmol/L lower for cholesterol values but almost identical for apolipoprotein A-I and phosphatidylcholine. Looking for delta 3,5-cholestadiene formed from cholesterol in the precipitation residue, we used "high-performance" liquid chromatography and "high-performance" thin-layer chromatography and found none.
Liquid-chromatographic determination of 5-S-L-cysteinyl-L-dopa with electrochemical detection in urine prepurified with a phenylboronate affinity gel.
Authors: Kågedal B , Pettersson A
Published: January 2020

A phenylboronate affinity gel has been investigated for use in the prepurification of urine before "high-performance" liquid chromatography and electrochemical determination of 5-S-L-cysteinyl-L-dopa. At pH 5.6 this naturally occurring 5-S-cysteinyldopa was adsorbed on a phenylboronate column and was quantitatively eluted with trichloroacetate, pH 3.0. This pretreatment of urine before "high-performance" liquid chromatography produced satisfactory chromatographic separations, and the results were further improved when the purification procedure also included treatment on a cation exchanger. By using 5-S-D-cysteinyl-L-dopa--a diastereomer to 5-S-L-cysteinyl-L-dopa--as an internal standard, we have developed a practical routine method for the quantitative determination of urinary 5-S-L-cysteinyl-L-dopa. The precision (CV = 2.4%) and analytical recovery (96.9%, SD 5.3%) were satisfactory and the results obtained correlated well with a previously described method.
Mechanism of cefoxitin and cephalothin interference with the Jaffé method for creatinine.
Authors: Kroll M H , Elin R J
Published: January 2020

Some cepha antibiotics interfere positively in the Jaffé method for determining creatinine. The extent of this interference differs with the modification used, and its mechanism is unknown. We studied the spectrophotometric and thermodynamic properties of the reaction of picrate with creatinine and two cepha antibiotics, cefoxitin and cephalothin. The absorbance spectra of the chromophore produced by each of the three compounds when reacted with picrate were essentially identical, with an absorbance maximum at 485 nm. We determined the molar absorptivity, rate constant, and equilibrium constant for the reaction of the three compounds with picrate. The molar absorptivity was similar for creatinine and cefoxitin but about fourfold smaller for cephalothin. The rate constant and equilibrium constant for the reaction with picrate were respectively 20- and 10-fold greater for creatinine than for either of the two cepha antibiotics. Thus, these differences indicate the mechanism by which these two cepha antibiotics interfere with the Jaffé procedure for creatinine, and explain why the degree of interference depends on details of the procedure.
The "HemoQuant" test: a specific and quantitative determination of heme (hemoglobin) in feces and other materials.
Authors: Schwartz S , Dahl J , Ellefson M , Ahlquist D
Published: January 2020

We describe a new, specific, quantitative method for fecal blood, based on conversion of nonfluorescing heme to fluorescing porphyrins, that obviates serious deficiencies inherent in currently used tests. A two-reagent system is used to determine the two hemoglobin-related fractions that are found in feces. The hot citric acid extract includes only the variable fraction of porphyrins that have been preformed from heme in the intestinal tract; this often is the major fraction. Total hemoglobin is indirectly determined by reaction with heated oxalic acid:FeSO4 reagent, which converts the remaining heme to porphyrin without loss of the preformed porphyrins. A three-step purification procedure eliminates interfering materials. Analytical recovery of added hemoglobin is linearly related to concentration over a several-thousand-fold range. The assay is equally applicable to whole blood or to sub-microgram amounts of hemoglobin in the 8-mg (wet weight) fecal sample tested. Quality control by liquid chromatographic and fluorometric analysis documents fluorescence specificity of the heme-derived porphyrins.
Gas-liquid chromatographic determination of propylene glycol in plasma and urine.
Authors: Yu D K , Sawchuk R J
Published: January 2020

We describe a gas-chromatographic assay for propylene glycol in human plasma and urine. The method is sensitive enough to allow quantification of the compound at concentrations observed clinically. It requires no derivative preparation and takes only 13 min of chromatographic time.
Meaning of serum free-thyroxin values in nonthyroidal illnesses: seven methods compared.
Authors: Chan D W , Waud J M , Taylor E , Stem J , Drew H , Oropeza M , Sucupira M S
Published: January 2020

Serum free thyroxin (FT4) was determined in 40 patients with various nonthyroidal illnesses. We studied seven methods: (1) a free thyroxin index calculated from total T4 and triiodothyronine resin uptake; (2) a free T4 index determined by enzyme inhibitor assays (Abbott's "Tetrazyme" and "Thyrozyme"); (3) a free T4 index calculated from total T4 and thyroxin-binding globulin; (4) free T4 by equilibrium dialysis; (5) Amersham's free T4 RIA; (6) Clinical Assays' one-step free T4 RIA; and (7) Clinical Assays' two-step free T4 RIA. Approximately half of the free T4 results were in the euthyroid range and the other half in the hypothyroid range by methods 1, 2, 5, and 6. Results for free T4 by methods 3 and 7 were similar to those by equilibrium dialysis (method 4), the percentages of patients with results in the euthyroid range being 68%, 65%, and 76%, respectively.
Isotope dilution--mass spectrometry of thyroxin proposed as a reference method.
Authors: Möller B , Falk O , Björkhem I
Published: January 2020

We describe a highly accurate and specific method based on isotope dilution--mass spectrometry for assay of thyroxin in serum. A fixed amount of 2H2-labeled thyroxin is added to a fixed amount of serum. Thyroxin is isolated from the serum by an extraction--solvent-distribution method, converted into the N,O-bis(trifluoroacetyl)methyl ester, and subjected to combined gas chromatography--mass spectrometry. The amount of thyroxin is determined from recordings at m/e 799 and m/e 801. These two ions correspond to the M-184 peak in the mass spectrum of the derivative of unlabeled and 2H2-labeled thyroxin, respectively. With this derivative and with the specific gas-chromatographic conditions used, the "memory" effects observed in previous work were absent. The accuracy of the method was checked by analyses of serum samples without addition of internal standard and by different recovery experiments. The intra-assay variation (CV) was less than 2%, the interassay variation (CV) less than 3.5%. We suggest that this method may be used as a Reference Method. Results by the present method were compared with those by a commercial radioimmunoassay. To our knowledge this represents the first time that radioimmunoassay of thyroxin has been evaluated with a more nearly accurate method. There was excellent agreement between results by the two methods (r = 0.997). We conclude that radioimmunoassay of serum thyroxin is sufficiently accurate for use in clinical work.
Creatinine determined by "high-performance" liquid chromatography.
Authors: Ambrose R T , Ketchum D F , Smith J W
Published: January 2020

We describe a "high-performance" liquid-chromatographic procedure for determination of creatinine in serum. Creatinine is separated from other species by cation-exchange chromatography. Absorbance of the creatinine in the eluate is monitored at 234 nm. Results from this procedure on sera correspond closely to those of a manual Jaffé-reaction procedure involving use of a protein-free filtrate with additional cleanup of the sample with Lloyd's reagent. None of the 21 endogenous and exogenous substances that we screened interfered. The overall CV at 138 mumol of creatinine per liter is 1.35% and at 539 mumol/L is 0.72%.
Value of the anion gap in clinical diagnosis and laboratory evaluation.
Authors: Lolekha P H , Lolekha S
Published: January 2020

We report the incidence of normal (50.4%), increased (46.7%), and decreased (2.9%) anion gap among hospitalized patients in a retrospective study. The mean and range of increased anion gaps were 25 and 19-28 mmol/L. Values exceeding 30 mmol/L were uncommon and may indicate either acidosis or laboratory error. The most common causes of the increased anion gap among patients were chronic renal failure, congestive heart failure, malignant neoplasm, and diabetes mellitus. Increased anion gap in this study may be due to excess acids along with decreases in sodium, chloride, and carbon dioxide. The mean and range of decreased anion gap were 6 and 3-8 mmol/L. Anion-gap values less than 3 mmol/L were uncommon (one of 500 cases), and a high incidence of such values may indicate laboratory error. Nephrotic syndrome, liver cirrhosis, intestinal obstruction, and severe hemorrhage were the common disorders associated with decreased anion gap, which resulted from hypoalbuminemia and hyponatremia. Although most patients with decreased anion gap had hypoalbuminemia, hypoalbuminemic patients did not necessarily have decreased anion gap.