scholarly journals Use of lissamine rhodamine ceramide trihexoside as a functional assay for alpha-galactosidase A in intact cells

2010 ◽  
Vol 51 (9) ◽  
pp. 2808-2817 ◽  
Author(s):  
Christine R. Kaneski ◽  
Raphael Schiffmann ◽  
Roscoe O. Brady ◽  
Gary J. Murray
Keyword(s):  
Functional Assay ◽  
Intact Cells ◽  
Ceramide Trihexoside ◽  
Alpha Galactosidase

The Use of Urine Sediments in the Diagnosis of Fabry's Disease

1980 ◽  
Vol 38 ◽  
pp. 470-471
Author(s):  
J. T. McMahon ◽  
R. R. Tubbs ◽  
G. N. Gephardt
Keyword(s):  
Angina Pectoris ◽  
Metabolic Disorder ◽  
Correct Diagnosis ◽  
Skin Lesions ◽  
Galactosidase Activity ◽  
Fabry’S Disease ◽  
Fabry's Disease ◽  
Corneal Opacities ◽  
Ceramide Trihexoside ◽  
Alpha Galactosidase

Fabry's disease is a hereditary, metabolic disorder leading to tissue accumulation of ceramide trihexoside principally in the kidneys and arteriole walls due to a deficiency in catabolic alpha galactosidase activity. During the course of the disease, affected patients may present with angiokeratotic skin lesions, corneal opacities, complaints of extremity pain, and angina pectoris. Proteinuria usually appears between the second and third decade of life and may be the factor leading to correct diagnosis.

Oncornavirus assembly at the cell surface revealed in replicas of freeze-dried cells

1978 ◽  
Vol 36 (2) ◽  
pp. 354-355
Author(s):  
Anthony Demsey ◽  
Christopher W. Stackpole
Keyword(s):  
Cell Surface ◽  
Surface Membrane ◽  
Viral Origin ◽  
Intact Cells ◽  
Structural Formation ◽  
Virus Core ◽  
Freeze Dried ◽  
Cacodylate Buffer ◽  
Surface Replica ◽  
Leukemia Viruses

The murine leukemia viruses are type-C oncornaviruses, and their release from the host cell involves a “budding” process in which the newly-forming, RNA-containing virus core becomes enveloped by modified cell surface membrane. Previous studies revealed that the released virions possess a dense array of 10 nm globular projections (“knobs”) on this envelope surface, and that these knobs contain a 70, 000 MW glycoprotein (gp70) of viral origin. Taking advantage of this distinctive structural formation, we have developed a procedure for freeze-drying and replication of intact cells which reveals surface detail superior to other surface replica techniques, and sufficient to detect even early stages of virus budding by localized aggregation of these knobs on the cell surface.Briefly, cells growing in monolayer are seeded onto round glass coverslips 10-12 mm in diameter. After a period of growth, cells are fixed in situ for one hour, usually with 1% OsO4 in 0. 1 M cacodylate buffer, and rinsed in distilled water.

Detection and mapping of interactions between chromatin and the nuclear envelope in drosophila embryos

1995 ◽  
Vol 53 ◽  
pp. 764-765
Author(s):  
W.F. Marshall ◽  
A.F. Dernburg ◽  
B. Harmon ◽  
J.W. Sedat
Keyword(s):  
Nuclear Envelope ◽  
Chromatin Condensation ◽  
Three Dimensional ◽  
Physiological Role ◽  
Nuclear Surface ◽  
Intact Cells ◽  
Molecular Determinants ◽  
Surface Harmonic ◽  
Drosophila Embryos

Interactions between chromatin and nuclear envelope (NE) have been implicated in chromatin condensation, gene regulation, nuclear reassembly, and organization of chromosomes within the nucleus. To further investigate the physiological role played by such interactions, it will be necessary to determine which loci specifically interact with the nuclear envelope. This will not only facilitate identification of the molecular determinants of this interaction, but will also allow manipulation of the pattern of chromatin-NE interactions to probe possible functions. We have developed a microscopic approach to detect and map chromatin-NE interactions inside intact cells.Fluorescence in situ hybridization (FISH) is used to localize specific chromosomal regions within the nucleus of Drosophila embryos and anti-lamin immunofluorescence is used to detect the nuclear envelope. Widefield deconvolution microscopy is then used to obtain a three-dimensional image of the sample (Fig. 1). The nuclear surface is represented by a surface-harmonic expansion (Fig 2). A statistical test for association of the FISH spot with the surface is then performed.

The uncalibrated prothrombinase-induced clotting time test

2010 ◽  
Vol 30 (04) ◽  
pp. 212-216 ◽  
Author(s):  
R. Jovic ◽  
M. Hollenstein ◽  
P. Degiacomi ◽  
M. Gautschi ◽  
A. Ferrández ◽  
...  
Keyword(s):  
Factor Xa ◽  
Heparin Therapy ◽  
Thrombin Inhibitors ◽  
Coagulation Cascade ◽  
Diagnostic Tools ◽  
Direct Thrombin Inhibitors ◽  
Functional Assay ◽  
Clotting Time ◽  
Coagulation Time ◽  
Time Test

SummaryThe activated partial thromboplastin time test (aPTT) represents one of the most commonly used diagnostic tools in order to monitor patients undergoing heparin therapy. Expression of aPTT coagulation time in seconds represents common practice in order to evaluate the integrity of the coagulation cascade. The prolongation of the aPTT thus can indicate whether or not the heparin level is likely to be within therapeutic range. Unfortunately aPTT results are highly variable depending on patient properties, manufacturer, different reagents and instruments among others but most importantly aPTT’s dose response curve to heparin often lacks linearity. Furthermore, aPTT assays are insensitive to drugs such as, for example, low molecular weight heparin (LMWH) and direct factor Xa (FXa) inhibitors among others. On the other hand, the protrombinase-induced clotting time assay (PiCT®) has been show to be a reliable functional assay sensitive to all heparinoids as well as direct thrombin inhibitors (DTIs). So far, the commercially available PiCT assay (Pefakit®-PiCT®, DSM Nutritional Products Ltd. Branch Pentapharm, Basel, Switzerland) is designed to express results in terms of units with the help of specific calibrators, while aPTT results are most commonly expressed as coagulation time in seconds. In this report, we describe the results of a pilot study indicating that the Pefakit PiCT UC assay is superior to the aPTT for the efficient monitoring of patients undergoing UFH therapy; it is also suitable to determine and quantitate the effect of LMWH therapy. This indicates a distinct benefit when using this new approach over the use of aPPT for heparin monitoring.

Clinical evaluation of a new thyroglobulin immunoradiometric assay in the follow-up of differentiated thyroid carcinoma

2003 ◽  
Vol 42 (02) ◽  
pp. 71-77 ◽  
Author(s):  
I. Schreivogel ◽  
C. Angerstein ◽  
U. Siefker ◽  
K. Lehmann ◽  
G. Altenvoerde ◽  
...  
Keyword(s):  
Thyroid Carcinoma ◽  
Residual Disease ◽  
False Negative ◽  
Differentiated Thyroid Carcinoma ◽  
Threshold Concentration ◽  
High Rate ◽  
Functional Assay ◽  
Hypothyroid Patient ◽  
Assay Sensitivity ◽  
Significant Difference

SummaryAim: Formal and clinical comparison of a new 3rd-gene-ration-Tg-IRMA (3-G-IRMA; Dynotest®Tg-plus) with a conventional Tg-IRMA (3-G-IRMA; SELco®Tg-assay) for patients with differentiated thyroid carcinoma. In addition we evaluated, if thyroglobulin (Tg) levels above a specific threshold concentration indicate the need for further investigations for residual disease. Patients, methods: Tg concentration of 105 sera of 93 consecutive patients with a differentiated thyroid cancer was determined with both assays and compared at different cut-off values (Dynotest®Tg-plus: 0.2, 1, 2 ng/ml; SELco®Tg-assay: 0.5, 1, 2 ng/ml) with the clinical results in respect to the corresponding TSH concentration. Results: Tg concentration did not show any significant difference (SELco®Tg-assay 0.5 ng/ml, Dynotest® Tg-plus 0.2 ng/ml). The Tg-values of both assays correlated with 97%. However, correlation of recovery in both assays was small (40%). The sensitivities and specificities of both assays at different cut-offs and TSH values did not reveal significant differences. In patients with TSH concentration >30 µU/ml the functional assay sensitivity was superior to arbitrary cut-offs in the decision to start further evaluations. Conclusions: In our study neither formal nor clinical significant differences between two Tg-assays were found. In a hypothyroid patient (TSH >30 µU/ml, Tg concentration exceeding the functional assay sensitivity) further investigations for residual disease are warranted. Higher thresholds are of limited value, due to a inacceptable high rate of false negative results.

The Uptake of Adenine by Isolated Human Platelet Membranes

1974 ◽  
Vol 32 (02/03) ◽  
pp. 457-464
Author(s):  
Paul C. French ◽  
Jan J. Sixma ◽  
Holm Holmsen
Keyword(s):  
Human Platelet ◽  
Facilitated Diffusion ◽  
Adenine Phosphoribosyltransferase ◽  
Ph Optimum ◽  
Intact Cells ◽  
Platelet Membranes ◽  
Group Translocation ◽  
Triton X

SummaryAdenine uptake into isolated platelet membranes had about the same Km (151 ± 21 • 9 nM) as uptake into intact cells (159 ± 21 nM) and was also competitively inhibited by papaverine and hypoxanthine. No uptake occurred at 0° and accumulated adenine was converted to AMP. AMP was not firmly bound to protein as judged by chromatography of triton X-100 solubilized membranes on Sephadex G25. The pH optimum for adenine uptake was at pH 5-5. Exogenous 5-phosphoribosyl-l-pyrophos- phate strongly stimulated uptake. These data may be explained by uptake of adenine by facilitated diffusion followed by conversion to AMP by adenine phosphoribosyltransferase but group translocation cannot be entirely excluded.

A New Functional Assay for Human Protein S Activity Using Activated Factor Vas Substrate

1989 ◽  
Vol 62 (04) ◽  
pp. 1144-1145 ◽  
Author(s):  
Martine Wolf ◽  
Catherine Boyer-Neumann ◽  
Jean-Luc Martinoli ◽  
Catherine Leroy-Matheron ◽  
Amiral Jean ◽  
...  
Keyword(s):  
Protein S ◽  
Human Protein ◽  
Functional Assay ◽  
Protein S Activity

A Novel Functional Assay of Protein C in Human Plasma and fts Comparison with Amidolytic and Anticoagulant Assays

1992 ◽  
Vol 67 (01) ◽  
pp. 046-049 ◽  
Author(s):  
H A Guglielmone ◽  
M A Vides
Keyword(s):  
Oral Anticoagulant ◽  
Protein C ◽  
Activated Protein C ◽  
Protein S ◽  
Normal Subjects ◽  
Test Sample ◽  
Functional Assay ◽  
Fast Method ◽  
Factor Va ◽  
Protein C Activity

SummaryA simple and fast method for the quantitative determination of protein C activity in plasma is here described. The first step consists in the conversion of protein C in the test sample into activated protein C by means of an activator isolated from Southern Copperhead venom. Subsequently, the degradation of factor Va, in presence of protein C-deficient plasma, is measured by the prolongation of the prothrombin time which is proportional to the amount of protein C in the sample. The dose-response curve showed a linear relationship from 6 to 150% protein C activity and the inter- and intra-assay reproducibility was 3.5% and 5.6% respectively. In normal subjects, a mean of protein C level of 98 ± 15% of normal pooled plasma was found. Comparison with the anticoagulant assay in samples of patients with oral anticoagulant, liver cirrhosis, disseminated intravascular coagulation and severe preeclampsia revealed an excellent correlation (r = 0.94, p <0.001). Also, a similar correlation (r = 0.93, p <0.001) existed between amidolytic assay and the method here proposed for all the samples studied without including the oral anticoagulant group. These results allowed us to infer that this method evaluates the ability of protein C to interact with protein S, phospholipids, calcium ions and factor Va.

Behaviour and Quantitation of Extrinsic (Tissue-Type) Plasminogen Activator in Human Blood

1983 ◽  
Vol 50 (02) ◽  
pp. 518-523 ◽  
Author(s):  
C Kluft ◽  
A F H Jie ◽  
R A Allen
Keyword(s):  
Plasminogen Activator ◽  
Venous Occlusion ◽  
Tissue Type ◽  
Functional Assay ◽  
Uterine Tissue ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator ◽  
Baseline Values

SummaryFunctional assay of extrinsic (tissue-type) plasminogen activator (EPA) in plasma on fibrin plates was evaluated. Using specific quenching antibodies, we demonstrated the method to be specific for EPA under all conditions tested. Contributions of urokinases and intrinsic activators were excluded. The quantity of EPA in blood samples, as compared with purified uterine tissue activator, shows 1 blood activator unit (BAU) to be comparable to 0.93 ng.The median values for EPA activity for healthy volunteers were: baseline, 1.9 BAU/ml (n = 123); diurnal, 5.5 BAU/ml (n = 12); DDAVP administration, 11.7 BAU/ml (n = 39); exhaustive exercise, 25 BAU/ml (n = 24); venous occlusion (15 min), 35 BAU/ml (n = 61). A large inter-individual variation in EPA activity was found, while individual baseline values tended to be constant for periods of weeks.In vitro in blood EPA activity shows a disappearance of 50% in about 90 min at 37° C; EPA activity in euglobulin fractions is stable for ≤2 hr at 37° C.A rapid decrease in EPA activity occurs in vivo, as noted after extracorporal circulation and exercise stimulation (t½ decay, 2-5 min).

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