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B T Doumas

Publications and source records attributed to B T Doumas.

At least 37 records · Page 2Linked to original sources
PubMed1986

Measurement of total bilirubin by use of bilirubin oxidase.

We used an enzymatic method for measuring total bilirubin in serum. Results by this method varied linearly with bilirubin concentrations to at least 300 mg/L. The day-to-day precision (CV) of the method ranged from less than 1% to about 11% at bilirubin concentrations of 183 and 12 mg/L, respectively. Commonly used anticoagulants, serum preparation materials, and selected drugs had no effect on the apparent bilirubin concentration, but turbidity caused a slight increase and hemoglobin concentrations of 2 g/L resulted in lower values, by as much as 17 mg/L at a bilirubin concentration of 95 mg/L. Patients' results obtained with this enzymatic method were slightly lower than those obtained with methods based on the Jendrassik-Grof principle. The largest differences, seen in samples with high "direct" bilirubin concentrations, can be decreased by measuring the absorbance at 425 nm instead of at 465 nm as recommended by the supplier of the bilirubin oxidase method.

Adult↗
PubMed1985

Chemical nature of a synthetic bilirubin conjugate and its reactivities in the total and direct reactions by the Jendrassik-Gróf method.

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

Adsorption↗
PubMed1985

Candidate reference method for determination of total bilirubin in serum: development and validation.

This candidate Reference Method for measuring total bilirubin in serum is based on the Jendrassik-Gróf principle (Clin Chem 29: 297-301, 1983). Standard Reference Material no. 916 bilirubin (National Bureau of Standards) is used as the standard. Bilirubin standard solutions may be prepared either in human serum or in 40 g/L albumin solution (human or bovine), because we found the molar absorptivity of the azopigment at 598 nm to be identical in these media. The absorptivities of the unconjugated and conjugated azopigments appear to be identical, but the conjugated azopigment is completely hydrolyzed in the final reaction mixture. Bilirubin added to serum from adults or neonates was quantitatively accounted for. Interference by hemoglobin (up to 2 g/L), ascorbic acid (up to 20 mg/L), or zinc (at physiological concentrations) is negligible. Of the therapeutic drugs we tested, only L-dopa and alpha-methyldopa interfere. We established normal adult reference values for total bilirubin and examined the intraindividual variation in 19 subjects.

Adult↗
PubMed1982

Pitfalls in the American Monitor kit methods for determination of total and "direct" bilirubin.

We evaluated the American Monitor Corporation kit for total and direct-reacting bilirubin and found that it suffers serious deficiencies, which lead to inaccurate and imprecise results. The main problem with the total-bilirubin procedure is that the short reaction time (2 min) is inadequate for completion of the reaction. The poor precision of the direct-bilirubin method is due to the short reaction time and the inability of the "stabilizer" (hydroxylamine sulfate) to completely destroy the diazo reagent. Depending on when Fehling's reagent is added, the reaction time may vary from 2 min to 7 min. Values for direct bilirubin at 7 min exceed those obtained at 2 min by 17 to 29%. The short reaction time makes color development temperature dependent, an additional source of imprecision. The suboptimal concentration of the diazo reagent results in underestimation of direct-reacting bilirubin. We recommend changes that improve both precision and accuracy of the kit procedures.

Ascorbic Acid↗
PubMed1981

A candidate reference method for determination of total protein in serum. I. Development and validation.

We developed a candidate Reference Method for measuring total serum protein by use of the biuret reaction. The method involves a previously described biuret reagent (Clin. Chem. 21: 1159, 1975) and Standard Reference Material (SRM) 927 bovine albumin (National Bureau of Standards) as the standard. At 25 degrees C, color development for 30 or 60 min provides identical serum protein values. Glucose (up to 10 g/L) and bilirubin (up to 300 mg/L) do not interfere. Hemoglobin, at 3 g/L, increases apparent serum protein by 0.4 g/L. The presence of dextran in serum causes easily detected turbidity, but this interference can be eliminated by centrifuging the reaction mixture. Therapeutic concentrations of ampicillin, carbenicillin, penicillin, oxacillin, nafcillin, chloramphenicol, cephalothin, and methicillin in blood do not interfere, nor do triglycerides up to 10 g/L. Within-run and day-to-day standard deviations of the method are 0.1 and 0.4 g/L, respectively.

Bilirubin↗
PubMed1981

A candidate reference method for determination of total protein in serum. II. Test for transferability.

The transferability of the candidate Reference Method for total serum protein was tested in eight laboratories in the United States and Europe. National Bureau of Standards SRM 927 (bovine serum albumin) was used in each analytical run as the calibration standard. The mean absorptivity value obtained for this material was 0.2983 L g-1 cm-1. Four serum pools prepared at the Centers for Disease Control were analyzed on each of 15 days. Within-run variation of the protein values (expressed as CV) in the eight laboratories ranged from 0.1 to 2.5% and day-to-day (total) variation in six of the laboratories ranged from 0.4 to 1%.

Biuret Reaction↗
PubMed1980

Plasma dopamine-beta-hydroxylase in uremia--increase in enzyme activity after renal transplantation.

Plasma dopamine-beta-hydroxylase in uremia. Plasma dopamine-beta-hydroxylase (DBH) activity was found to be low in 26 uremic patients when compared with 56 normal individuals (p less than 0.001). Hemodialysis caused only a slight increase in plasma DBH levels in the uremic group. In contrast, a group of kidney transplanted patients with a return of good renal function had DBH values similar to the normal group (p greater than 0.1). The mean plasma DBH activity in eight patients measured pre- and post-transplantation increased from 4.5 to 28 International Units/l (p less than 0.01). No evidence was found to indicate that the depressed levels of plasma DBH in uremia were secondary to genetic or enzyme inhibiting factors. It is suggested that low levels of DBH activity in patients with renal failure may be a consequence of altered sympathetic nervous activity which is known to occur in the uremic state.

Adolescent↗
PubMed1979

Performance of the Du Pont aca ammonia method.

We evaluated the performance of the Du Pont aca ammonia procedure with regard to (a) linearity, (b) precision, (c) interferences, and (d) effect of anticoagulants. Linearity extends to 2,000 mumol/L. The SD of the method was essentially constant (2 to 3 mumol/L) and independent of the NH3 concentration. Hemoglobin, bilirubin, and lipemia do not interfere. Either EDTA or heparin is suitable as anticoagulant. Recovery of NH3 added to plasma samples averaged 102% (range: 97--107%). We established normal values by measuring NH3 in 188 plasma samples from apparently healthy individuals. The 95% confidence range is from 10 to 35 mumol/L. The aca ammonia method compares very well with that of Kingsley and Tager but correlates less strongly with that of Reinhold and Chung. We describe a protein-based solution with stable NH3 concentration that is suitable as a control material.

Ammonia↗
PubMed1976

An improved freeze-dried bilirubin preparation.

We describe the conditions for successful freeze-drying of bilirubin solutions in the presence of dimethylsulfoxide and human serum albumin. Freeze-dried bilirubin preparations rehydrate readily, rapidly resulting in clear solutions. The preparations exhibit excellent homogeneity and stability at -20 degrees C. Accelerated storage tests indicate that the freeze-dried product would also be stable at higher temperatures but our results do not permit, at the present time, to make accurate predictions regarding the stability at 4 degrees C or 0 degrees C. We believe that our approach will permit manufacturers of bilirubin calibrators for clinical use to improve the quality of their products.

Animals↗
PubMed1976

Lactate dehydrogenase activities in serum and plasma.

We investigated lactate dehydrogenase activity in serum and plasma because of the conflicting data found in the literature. We assayed serum, platelet-rich, and platelet-poor plasma by two colorimetric endpoint methods and by an ultraviolet kinetic procedure. Platelet-poor plasma and serum had essentially the same activities by all three methods, whereas the activity in platelet-rich plasma plasma averaged fourfold that in platelet-poor plasma or serum when the assay was performed under conditions that result in lysis of platelets and release of their lactate dehydrogenase. When measurements were performed in platelet-rich plasma under conditions that prevented lysis of platelets, all three types of specimens gave the same results. This occurred when the osmolality of the reaction mixture was about 240 mOsm/kg of water. At an osmolality of about 120 mOsm, the activity of platelet-rich plasma was substantially lower than that of platelet-poor plasma or serum. If plasma must be used, the sample must first be centrifuged (3000 X g, 15 min) to provide a platelet-free plasma.

Blood Cell Count↗
PubMed1975

Standards for total serum protein assays--a collaborative study.

We have studied the standardization of total serum protein assay with the biuret reaction. Standard solutions were prepared from lyophilized preparations of human serum albumin and bovine serum albumin, with corrections made for volatile material and ash contents. These solutions and a solution of crystalline albumin standard were analyzed with a new stable biuret reagent, to establish absorptivity values (values for the absorbance of a 1 g/liter final reaction mixture). The mean values obtained were 0.302, 0.292, and 0.290 for human serum albumin, bovine serum albumin, and the crystalline albumin, respectively. We believe that the established absorptivity value will improve the accuracy of serum protein determinations. We studied the linearity of the relation between color produced and protein concentration, with use of the solutions described above and a serum pool. The color adheres to Beer's law up to the highest concentration tested: 3 g/liter for HSA and BSA, and 2.8 g/liter for serum in the final reaction mixture. The new biuret reagent has been stable for one year at room temperature. We recommend the use of bovine serum albumin as a primary standard for serum protein assays. It is inexpensive, easily available, and exhibits the best linearity in the biuret reaction.

Animals↗
PubMed1975

The effect of heparin and EDTA on the NBT test.

Heparin precipitates nitroblue tetrazolium (NBT) in the absence of protein (plasma). This NBT-heparin precipitate, when presented to neutrophils, results in NBT scores higher than those obtained with NBT in solution. It is postulated that the high NBT scores obtained with increasing heparin concentrations are due to increased NBT precipitation and hence increased phagocytosis. It is proposed that the variability of NBT scores obtained when heparin is used as the anticoagulant is related to the amount of precipitate formed and to inhibition of phagocytosis by excess heparin. EDTA, in the presence of plasma and NBT, forms a precipitate consisting of protein and little, if any, NBT. In the presence of EDTA, NBT scores are lower than those obtained with heparin. EDTA, at concentrations used to prevent coagulation, inhibits phagocytosis and this could explain the low NBT scores. These observations provide a mechanism for entry of NBT dye into neutrophils and may help explain the inconsistent results found in both normal and disease states.

Bacterial Infections↗