Fetal Hemoglobin: Optimum Conditions for its Estimation by Alkali Denaturation

Molden, Daniel P.; Alexander, Natalie; Neeley, William E.

doi:10.1093/ajcp/77.5.568

Cited by 21 publications

(9 citation statements)

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“…1) (Table I). Percentage hemoglobin S was measured using a Paragon hemoglobin electrophoresis kit at pH 8.6 (Beckman Instruments Inc., Fullerton, CA) and fetal hemoglobin by alkali denaturation (19). Oxygen saturation curves on erythrocytes in HBS (PCV -0.35; 35%) were determined as association curves at 370C using an Aminco oxygen dissociation analyzer (Hem-O-Scan; American Instrument Co., Silver Spring, MD).…”

Section: Methodsmentioning

confidence: 99%

Polymerization of sickle cell hemoglobin at arterial oxygen saturation impairs erythrocyte deformability.

Green¹,

Noguchi²,

Keidan³

et al. 1988

J. Clin. Invest.

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We have examined the filterability of sickle erythrocytes, using an initial-flow-rate method, to determine whether sufficient hemoglobin S polymer forms at arterial oxygen saturation to adversely affect erythrocyte deformability. The amount of intracellular polymer was calculated as a function of oxygen saturation to estimate the polymerization tendency for each of eight patients with sickle cell anemia (SCA). Progressive reduction of oxygen tension within the arterial range caused a sudden loss of filterability of SCA erythrocytes through 5-,nmdiam pores at a critical P02 between 110 and 190 mmHg. This loss of filterability occurred at a higher P02 than did morphological sickling, and the critical Po2 correlated significantly (r = 0.844-0.881, P < 0.01) with the polymerization tendency for each patient. Study of density-gradient fractionated cells from four SCA patients indicated that the critical P02 of dense cells was reached when only a small amount of polymer had formed, indicating the influence of this subpopulation on the results obtained for unfractionated cells. Impairment of erythrocyte filterability at high oxygen saturation (> 90%) suggests that small changes in oxygen saturation within the arterial circulation cause Theological impairment of sickle cells.

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Section: Methodsmentioning

confidence: 99%

Polymerization of sickle cell hemoglobin at arterial oxygen saturation impairs erythrocyte deformability.

Green¹,

Noguchi²,

Keidan³

et al. 1988

J. Clin. Invest.

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“…A 1-min alkali denaturation test was used to determine the percentage of peripheral HbF in hemolysates prepared from SS patients' red cells as described [42]. Normal controls had HbF levels of less than 1%.…”

Section: Hbf Determinationmentioning

confidence: 99%

Circulating cytokines response and the level of erythropoiesis in sickle cell anemia

Croizat

Nagel

1999

Am. J. Hematol.

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A hemoglobin F (HbF) level between eight and nine percent divides sickle cell anemia (SS) patients into two populations, according to the kinetics of circulating burst forming units-erythroid (BFU-E), long term culture-initialing cells (LTC-IC), and cytokine plasma concentrations. The SS patients with HbF levels lower than 8-9% are more anemic (LFSS patients) than those with HbF levels higher than 8-9% who have less severe anemia (HFSS patients). We report here that the level of erythropoiesis [evaluated by the levels of soluble transferin receptors (sTfR)] is not identical in these two patient populations, supporting the idea that a different set of regulatory mechanisms might be required to maintain the two levels of increased hematopoiesis. The plasma sTfR concentration was increased in all SS samples compared with controls (P < 0.002) and sTfR levels were negatively correlated with peripheral HbF%. (r = -0.574, P < 0.002). Furthermore, sTfR levels were higher in LFSS than in HFSS patients. Erythropoietin (Epo) levels were increased in the plasma of LFSS individuals (range = 34-215 ml U/ml), while the values in HFSS patients were in the normal range (3-20 ml U/ml). Furthermore, we identify here stem cell factor (SCF) and transforming growth factor-beta (TGF-beta) as regulatory factors specifically affected by the presence of SS genotype and its level of severity. The plasma concentrations of SCF and TGF-beta were increased compared with normal controls and high levels of SCF (up to 7,000 pg/ml) were detected in LFSS patients. The latter also showed increased proportion of SCF+ CD34 enriched circulating cells (49%). Low SCF in HFSS patients is associated with elevated TGF-beta, suggesting a regulatory role of the latter on either SCF release or c-kit expression in progenitor cells. Occasional elevation of granulocyte macrophage-colony stimulating factor (G-CSF), interleukin (IL)-7, and macrophage inflammatory protein (MIP)-1alpha in plasma of SS patients is not specific because no relation to HbF could be demonstrated. All plasma tested for leukemia inhibitory factor (LIF) were negative. Data presented here, complementing previously published information, supports a model in which HFSS patients achieve a balance between inhibitory (TGF-beta) and stimulatory (SCF, IL-3) factors, resulting in moderate erythropoietic response. In contrast, in LFSS patients, low levels of TGF-beta and the increased release of GM-CSF and SCF maintain the intense erythropoiesis in response to higher erythropoietic stress, in these more severe patients.

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“…2 are consistent with the use of Fe͑CN͒ 6 3− as a mild oxidant that can denature some proteins. 19,20 However, other reports have suggested that Fe͑CN͒ 6 3−/4− interactions with proteins are either more complex 21 or in some cases benign. 22 Fe͑CN͒ 6 3−/4− has also been reported to disrupt the formation of Authiol self-assembled monolayers ͑SAM͒ through mild etching of Au.…”

Section: Resultsmentioning

confidence: 99%

Electrochemical Impedance Biosensor for Glucose Detection Utilizing a Periplasmic E. coli Receptor Protein

Wang¹,

Carmon

Luck

et al. 2005

Electrochem. Solid-State Lett.

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We report the development of a reagentless electrochemical impedance biosensor for glucose that employs the D-glucose/galactose receptor from E. coli for direct glucose detection. The biological platform for this sensor is an Au surface to which the protein is immobilized through formation of an Au-S bond to a genetically engineered cysteine residue at the N-terminus. The impedance signal detects the extensive ligand-induced domain motion within the protein upon glucose binding. We show the applicability of impedance spectroscopy in conjunction with periplasmic binding proteins as a general method for detecting small molecules.For control of diabetes mellitus, maintenance of blood glucose concentrations within the normal physiological range is critical to minimizing diabetic complications. As a result, accurate in vivo and in vitro measurement of glucose concentration in physiological fluids has long been a central goal of biosensor research. Traditionally, glucose biosensors employ indirect detection of the products of the glucose oxidase or glucose dehydrogenase reactions, although direct detection has also been reported. In general, glucose oxidase has been the favored biological recognition element due to the high binding specificity, high turnover rate, and high stability of this enzyme. However, indirect detection of hydrogen peroxide, or other products, often suffers from low selectivity arising from chemical interference by other easily reducible substances, such as ascorbate and urate. Direct detection suffers from two general drawbacks that limit the efficiency with which electrons can be collected: proteins are usually electrically insulating, and the reaction site may be located far from the collection electrode. 1 In addition, electron transfer proteins are also required. To circumvent these problems, the current report focuses on direct glucose detection by electrochemical impedance using the D-glucose/galactose receptor ͑GGR͒ from E. coli.The potential use of the bacterial periplasmic binding protein superfamily ͑bPBP͒ in biosensors has arisen due to their solubility, stability and ability to reversibly bind a large variety of small ligands including sugars, amino acids, and inorganic ions. 2 These proteins comprise a large family of functionally similar receptors with a two-domain structure that exhibit a large hinge-twist conformational change upon ligand binding. Whereas the proteins are open and flexible when the ligand is absent, they are more structurally compact and closed when the ligand is bound. This large amplitude motion can be exploited for biosensor development of small molecules. [3][4][5][6] The GGR protein is specific for glucose or galactose and binds these ligands in the micromolar range, which is more sensitive than that needed for blood glucose. A number of groups have detected sugars by labeling this protein with fluorescent tags in sites where the chemical environment is altered during ligand binding. 7-10 Two recent studies have used surface plasmon resonance ͑SPR͒ methods w...

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Fetal Hemoglobin: Optimum Conditions for its Estimation by Alkali Denaturation

Cited by 21 publications

References 0 publications

Polymerization of sickle cell hemoglobin at arterial oxygen saturation impairs erythrocyte deformability.

Polymerization of sickle cell hemoglobin at arterial oxygen saturation impairs erythrocyte deformability.

Circulating cytokines response and the level of erythropoiesis in sickle cell anemia

Electrochemical Impedance Biosensor for Glucose Detection Utilizing a Periplasmic E. coli Receptor Protein

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