One-step chromogenic equivalent of activated partial thromboplastin time evaluated for clinical application

Abstract We evaluated the clinical usefulness of a recently developed semi-automated one-step chromogenic equivalent of activated partial thromboplastin time (APTT; Behring). This simple test is easily adaptable for automation. Generally, the results with this chromogenic one-step APTT were at least as precise as those obtained with comparative coagulometric methods. The chromogenic one-step APTT showed, both in vitro and in vivo, adequate sensitivity to congenital intrinsic factor deficiency but no sensitivity to Factor VII deficiency. Unlike a two-step coagulometric APTT (Dade), the one-step chromogenic APTT seemed sensitive to activation products of the contact system, which are present in immunoadsorbed factor-deficient plasma. The in vitro sensitivity of the chromogenic APTT to heparin was comparable with that of a coagulometric APTT, but the sensitivity to heparin in patients' samples differed slightly. The chromogenic APTT is relatively insensitive to anomalies in the fibrinogen-fibrin conversion. Finally, we observed discrepancies between the chromogenic and coagulometric APTT results for plasma of patients with disseminated intravascular coagulation. We conclude that this one-step chromogenic APTT warrants further evaluation for possible use as a routine test for the clinical laboratory.

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Thein vivocharacterization of electrospun heparin-bonded polycaprolactone in small-diameter vascular reconstruction

Vascular ◽

10.1177/1708538114550737 ◽

2014 ◽

Vol 23 (4) ◽

pp. 358-365 ◽

Cited By ~ 4

Author(s):

Hong-Yong Duan ◽

Lin Ye ◽

Xin Wu ◽

Qiang Guan ◽

Xiao-Fei Yang ◽

...

Keyword(s):

Partial Thromboplastin Time ◽

Small Diameter ◽

Gel Permeation Chromatography ◽

Vascular Reconstruction ◽

Activated Partial Thromboplastin Time ◽

Collagen Content ◽

Implantation Time ◽

Artery Reconstruction

ObjectiveTo evaluate the possibility of using heparin-bonded polycaprolactone grafts to replace small-diameter arteries.MethodsPolycaprolactone was bonded with heparin. The activated partial thromboplastin time of heparin-bonded polycaprolactone grafts was determined in vitro. Small-diameter grafts were electrospun with heparin-bonded polycaprolactone and polycaprolactone and were implanted in dogs to substitute part of the femoral artery. Angiography was used to investigate the patency and aneurysm of the grafts after transplantation. After angiography, the patent grafts were explanted for histology analysis. The degradation of the grafts and the collagen content of the grafts were measured.ResultsActivated partial thromboplastin time tests in vitro showed that heparin-bonded polycaprolactone grafts exhibit obvious anticoagulation. Arteriography showed that two heparin-bonded polycaprolactone and three polycaprolactone grafts were obstructed. Other grafts were patent, without aneurysm formation. Histological analysis showed that the tested grafts degraded evidently over the implantation time and that the luminal surface of the tested grafts had become covered by endothelial cells. Collagen deposition in heparin-bonded polycaprolactone increased with time. There were no calcifications in the grafts. Gel permeation chromatography showed the heparin-bonded polycaprolactone explants at 12 weeks lose about 32% for Mw and 24% for Mn. The collagen content on the heparin-bonded polycaprolactone grafts increased over time.ConclusionThis preliminary study demonstrates that heparin-bonded polycaprolactone is a suitable graft for small artery reconstruction. However, heparin-bonded polycaprolactone degrades more rapidly than polycaprolactone in vivo.

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Chemorheological Studies of the Effect of Heparin on the Course of Coagulation

Thrombosis and Haemostasis ◽

10.1055/s-0038-1647977 ◽

1976 ◽

Vol 35 (02) ◽

pp. 447-459

Author(s):

K. A Overholser ◽

C. B Baysinger ◽

T. R Harris ◽

T Deveau

Keyword(s):

Whole Blood ◽

Partial Thromboplastin Time ◽

Time Course ◽

Normal Subjects ◽

Activated Partial Thromboplastin Time ◽

Weissenberg Rheogoniometer ◽

Rheological Measurements ◽

Kinetics Of

SummaryThe influence of sodium heparin on viscoelastic change during coagulation was determined in vitro for whole blood samples from ten normal subjects at heparin concentrations ranging from 0 to 1.45 units/(ml whole blood). A four-parameter chemorheological model was used to describe the time course of coagulation as measured by the Weissenberg Rheogoniometer. One parameter compares closely with the whole blood activated partial thromboplastin time, while the other three may be related to the chemical kinetics of clotting.The chemorheological model and experimental techniques were then tested in a dog preparation. It was found that rheological measurements are more self-consistent than either thrombelastography or the activated partial thromboplastin time for the assay of in vivo heparin in two dogs.

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Effect Of A Non-Activated Prothrombin Complex Concentrate (Prothrombinex) On Platelets: In Vitro And In Vivo Studies

10.1055/s-0038-1653035 ◽

1981 ◽

Author(s):

H Ekert ◽

F L Dean ◽

J L Lane

Keyword(s):

Partial Thromboplastin Time ◽

Prothrombin Complex Concentrate ◽

Platelet Rich Plasma ◽

Inhibitory Effect ◽

Activated Partial Thromboplastin Time ◽

In Vivo Studies ◽

Chromogenic Substrates ◽

Activated Prothrombin Complex Concentrate

Chromatography on Sephadex G-200 of the prothrombin complex concentrate, prothrombinex (Px) showed it to have inhibitor and potentiator fractions when tested by the non-activated partial thromboplastin time (NAPTT). The inhibitory effect was related to the heparin content of Px, as it was removed on ECTEOLA-cellulose. The potentiator fraction clotted fibrinogen and this could be inhibited by hirudin. Thrombin-like activity of this fraction was shown using chromogenic substrates. The effect of the potentiator fraction in the NAPTT was markedly enhanced by platelets. Diluted Px and the potentiator fractions caused aggregation of washed platelets which could be inhibited by hirudin. Aggregation was independent of the prostaglandin pathway, as it occurred in washed aspirin-treated platelets. Neither diluted Px nor potentiator fractions aggregated platelets in platelet rich plasma. Infusion of Px was followed by a rise in β-thromboglobulin (β-TG) in 2 of 3 patients two minutes after infusion. This rise could not be ascribed to the presence of β-TG in Px or to the heparin present in Px. These findings suggest that Px has a thrombin-like activity and that its mode of action in patients with factor VIII antibodies may result from its effect on platelets and their interaction with coagulation enzymes.

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106 THE WARMING PROCEDURE: A FIRST STEP FOR IMPROVING THE NONSURGICAL DEEP INTRAUTERINE TRANSFER OF SOPS-VITRIFIED PORCINE EMBRYOS

Reproduction Fertility and Development ◽

10.1071/rdv24n1ab106 ◽

2012 ◽

Vol 24 (1) ◽

pp. 165

Author(s):

J. Gomis ◽

C. Cuello ◽

J. Sanchez-Osorio ◽

M. A. Gil ◽

I. Parrilla ◽

...

Keyword(s):

Reproductive Performance ◽

Petri Dish ◽

Control Group ◽

Hatching Rate ◽

Embryo Viability ◽

Step Method ◽

One Step ◽

The One

We previously reported successful nonsurgical deep intrauterine embryo transfer (ET) of fresh in vivo–derived porcine embryos. However, several trials from our laboratory demonstrated that when this procedure was used in combination with vitrified/warmed (VW) embryos, its efficiency was very low. Recently, we have shown that the one-step warming method in syringe, which was used in the earlier trials, compromises the in vitro embryo viability. The aim of this study was to confirm the negative effect of the direct warming in syringe and to evaluate the effect of alternative warming procedures on the in vivo development of VW embryos after nonsurgical ET. In Experiment 1, morulae and early blastocysts were collected on Days 5 to 6 (Day 0: onset of oestrus) and assigned to one of the following groups: 1) syringe group: vitrified embryos (n = 88) were warmed by the one-step method directly in a 1-mL syringe containing 300 μL of warming medium, which was connected to the ET catheter and then transferred to recipients (n = 6); 2) dish group: vitrified embryos (n = 194) were warmed with one-step warming method in a Petri dish containing 1 mL of warming medium, loaded into a Tom Cat catheter and transferred to recipients (n = 13); and 3) control group: fresh embryos (n = 129) were loaded in a 1-mL syringe in 100 μL of transfer medium and transferred to 9 recipients. An average of 15 embryos were transferred to each recipient on Day 4 or 5. Embryos were surgically recovered 24 h after ET. Data were analysed by ANOVA. The embryo recovery rate was similar among groups (range: 70.7 ± 4.8% to 77.2 ± 6.5%). The embryo survival (ES) and the hatching rate (HR) from the control group (94.0 ± 2.1% and 33.4 ± 7.6%, respectively) were higher (P ≤ 0.05) than those from the dish group (80.4 ± 4.6% and 14.5 ± 4.1%, respectively). All embryos from the syringe group were degenerated. Some viable recovered embryos (n = 135) were cultured for 48 h to evaluate their subsequent in vitro development. No differences were observed in ES between the control and the dish group (100.0 ± 0.0% vs 98.9 ± 1.0%). The HR in the control group (71.5 ± 2.1%) was higher (P ≤ 0.01) than that of the dish group (42.7 ± 6.1%). In experiment 2 we evaluated the reproductive performance of naturally cycling recipients after nonsurgical ET of vitrified embryos warmed with the one-step method in a Petri dish. An average of 35 VW morulae and blastocysts were transferred to each recipient (n = 10) on Days 4 to 6. Four recipients returned to oestrus at Days 21 to 22. The remaining 6 recipients were diagnosed pregnant by ultrasonography on Day 26. At Days 50 to 55, 5 recipients remained pregnant. In conclusion, the one-step in syringe warming method for vitrified porcine embryos had a completely adverse effect on the vivo viability, whereas nonsurgical ET of embryos warmed in a Petri dish allowed us to obtain promising reproductive performance. Supported by MICINN (AGL2009-12091) and SENECA (GERM 04543/07).

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Monitoring Heparin Therapy: Relationships between the Activated Partial Thromboplastin Time and Heparin Assays Based on Ex-Vivo Heparin Samples

Thrombosis and Haemostasis ◽

10.1055/s-0038-1645678 ◽

1990 ◽

Vol 63 (01) ◽

pp. 016-023 ◽

Cited By ~ 41

Author(s):

A M H P van den Bessekaar ◽

J Meeuwisse-Braun ◽

R M Bertina

Keyword(s):

Partial Thromboplastin Time ◽

Plasma Sample ◽

Ex Vivo ◽

Correlation Coefficients ◽

Heparin Therapy ◽

Thromboembolic Disease ◽

Activated Partial Thromboplastin Time ◽

Venous Thromboembolic Disease ◽

Venous Thromboembolic

SummaryFive different APTT reagents, two amidolytic anti-ITa assays, one amidoiytic anti-Xa assay, and one coagulometric anti-Xa/ anti-IIa assay were used to assess the effect of heparin in patients treated for venous thromboembolic disease. Good correlations were observed between lug-transformed APYE> determined with the various reagents (correlation coefficients: 0.92-0.96).Nevertheless there were important differences in the slopes of the lines of relationship between the APTT reagents.Good correlations were observed between the anti-Xa and anti-IIa assay results (correlation coefficients: 0.92-0.97). However, the amidolytic anti-Xa activity was significantly higher (p <0.001) than the two amidolytic anti-IIa activities. Less good correlations were observed between the log-transformed APTTs and the anti-Xa or anti-IIa activities (correlation coefficients: 0.64-0.78). The correlations were improved by transforming the APTT into APTT-ratio, i.e. the ratio of the patient’s APTT to the same patient’s APTT after removal of heparin from the plasma sample by means of ECTEOLA-cellulose treatment. The correlation coefficients of log (AFTT-ratio) with anti-Xa or anti-IIa ranged from 0.76 to 0.87.For both APTT and amidolytic heparin assay, the response to in vitro heparin was different from the response to ex vivo heparin.Therefore, equivalent therapeutic ranges should be assessed by using ex vivo samples rather than in vitro heparin. Because of the response differences between the APTT reagents, it is not adequate to define a therapeutic range for heparin therapy without specification of the reagent.

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Monitoring Anticoagulant Therapy by Activated Partial Thromboplastin Time: Hirudin Assessment

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648943 ◽

1994 ◽

Vol 72 (05) ◽

pp. 685-692 ◽

Cited By ~ 43

Author(s):

Michael T Nurmohamed ◽

René J Berckmans ◽

Willy M Morriën-Salomons ◽

Fenny Berends ◽

Daan W Hommes ◽

...

Keyword(s):

Whole Blood ◽

Partial Thromboplastin Time ◽

Ex Vivo ◽

Fold Increase ◽

Heparin Therapy ◽

Activated Partial Thromboplastin Time ◽

Recombinant Hirudin ◽

Interindividual Differences ◽

The Relationship

SummaryBackground. Recombinant hirudin (RH) is a new anticoagulant for prophylaxis and treatment of venous and arterial thrombosis. To which extent the activated partial thromboplastin time (APTT) is suitable for monitoring of RH has not been properly evaluated. Recently, a capillary whole blood device was developed for bed-side monitoring of the APTT and it was demonstrated that this device was suitable to monitor heparin therapy. However, monitoring of RH was not evaluated.Study Objectives. To evaluate in vitro and ex vivo the responsiveness and reproducibility for hirudin monitoring of the whole blood monitor and of plasma APTT assays, which were performed with several reagents and two conventional coagulometers.Results. Large interindividual differences in hirudin responsiveness were noted in both the in vitro and the ex vivo experiments. The relationship between the APTT, expressed as clotting time or ratio of initial and prolonged APTT, and the hirudin concentration was nonlinear. A 1.5-fold increase of the clotting times was obtained at 150-200 ng/ml plasma. However, only a 2-fold increase was obtained at hirudin levels varying from 300 ng to more than 750 ng RH/ml plasma regardless of the assays. The relationship linearized upon logarithmic conversion of the ratio and the hirudin concentration. Disregarding the interindividual differences, and presuming full linearity of the relationship, all combinations were equally responsive to hirudin.Conclusions. All assays were equally responsive to hirudin. Levels up to 300 ng/ml plasma can be reliably estimated with each assay. The manual device may be preferable in situations where rapid availability of test results is necessary.

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In Vitro Thrombogenicity Tests of Factor IX Concentrates

Thrombosis and Haemostasis ◽

10.1055/s-0038-1657035 ◽

1979 ◽

Vol 42 (05) ◽

pp. 1355-1367 ◽

Cited By ~ 8

Author(s):

C V Prowse ◽

A Chirnside ◽

R A Elton

Keyword(s):

Factor Viii ◽

Partial Thromboplastin Time ◽

Generation Time ◽

Activated Partial Thromboplastin Time ◽

Factor Ix ◽

In Vitro Tests ◽

Factor Viii Inhibitor ◽

Factor Ixa ◽

Viii Inhibitor

SummaryVarious factor IX concentrates have been examined in a number of in vitro tests of thrombogenicity. The results suggest that some tests are superfluous as in concentrates with activity in any of these tests activation is revealed by a combination of the non-activated partial thromboplastin time, the thrombin (or Xa) generation time and factor VIII inhibitor bypassing activity tests. Assay of individual coagulant enzymes revealed that most concentrates contained more factor IXa than Xa. However only a small number of concentrates, chiefly those that had been purposefully activated, contained appreciable amounts of either enzyme.

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Efficient Cryopreservation of Populus tremula by In Vitro-Grown Axillary Buds and Genetic Stability of Recovered Plants

Plants ◽

10.3390/plants10010077 ◽

2021 ◽

Vol 10 (1) ◽

pp. 77

Author(s):

Elena O. Vidyagina ◽

Nikolay N. Kharchenko ◽

Konstantin A. Shestibratov

Keyword(s):

Survival Rate ◽

Axillary Buds ◽

Long Term Storage ◽

Average Survival ◽

Transgenic Lines ◽

Ssr Loci ◽

One Step ◽

The One

Axillary buds of in vitro microshoots were successfully frozen at –196 °C by the one-step freezing method using the protective vitrification solution 2 (PVS2). Microshoots were taken from 11 transgenic lines and three wild type lines. Influence of different explant pretreatments were analyzed from the point of their influence towards recovery after cryopreservation. It was found out that the use of axillary buds as explants after removal of the apical one increases recovery on average by 8%. The cultivation on growth medium of higher density insignificantly raises the regenerants survival rate. Pretreatment of the osmotic fluid (OF) shows the greatest influence on the survival rate. It leads to the increase in survival rate by 20%. The cryopreservation technology providing regenerants average survival rate of 83% was developed. It was based on the experimental results obtained with explant pretreatment. Incubation time in liquid nitrogen did not affect the explants survival rate after thawing. After six months cryostorage of samples their genetic variability was analyzed. Six variable simple sequence repeat (SSR) loci were used to analyze genotype variability after the freezing-thawing procedure. The microsatellite analysis showed the genetic status identity of plants after cryopreservation and of the original genotypes. The presence of the recombinant gene in the transgenic lines after cryostorage were confirmed so as the interclonal variation in the growth rate under greenhouse conditions. The developed technique is recommended for long-term storage of various breeding and genetically modified lines of aspen plants, as it provides a high percentage of explants survival with no changes in genotype.

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Monocyte tissue factor induction by lipopolysaccharide (LPS): dependence on LPS-binding protein and CD14, and inhibition by a recombinant fragment of bactericidal/permeability-increasing protein

Blood ◽

10.1182/blood.v83.9.2516.2516 ◽

1994 ◽

Vol 83 (9) ◽

pp. 2516-2525 ◽

Cited By ~ 13

Author(s):

K Meszaros ◽

S Aberle ◽

R Dedrick ◽

R Machovich ◽

A Horwitz ◽

...

Keyword(s):

Tissue Factor ◽

Mononuclear Cells ◽

Human Peripheral Blood ◽

Binding Protein ◽

Mononuclear Phagocytes ◽

Factor Vii ◽

Peripheral Blood Mononuclear ◽

Recombinant Fragment

Abstract Mononuclear phagocytes, stimulated by bacterial lipopolysaccharide (LPS), have been implicated in the activation of coagulation in sepsis and endotoxemia. In monocytes LPS induces the synthesis of tissue factor (TF) which, assembled with factor VII, initiates the blood coagulation cascades. In this study we investigated the mechanism of LPS recognition by monocytes, and the consequent expression of TF mRNA and TF activity. We also studied the inhibition of these effects of LPS by rBPI23, a 23-kD recombinant fragment of bactericidal/permeability increasing protein, which has been shown to antagonize LPS in vitro and in vivo. Human peripheral blood mononuclear cells, or monocytes isolated by adherence, were stimulated with Escherichia coli O113 LPS at physiologically relevant concentrations (> or = 10 pg/mL). The effect of LPS was dependent on the presence of the serum protein LBP (lipopolysaccharide-binding protein), as shown by the potentiating effect of human recombinant LBP or serum. Furthermore, recognition of low amounts of LPS by monocytes was also dependent on CD14 receptors, because monoclonal antibodies against CD14 greatly reduced the LPS sensitivity of monocytes in the presence of serum or rLBP. Induction of TF activity and mRNA expression by LPS were inhibited by rBPI23. The expression of tumor necrosis factor showed qualitatively similar changes. Considering the involvement of LPS-induced TF in the potentially lethal intravascular coagulation in sepsis, inhibition of TF induction by rBPI23 may be of therapeutic benefit.

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Effect of microbial isolates on microbial yield estimation in RUSITEC system

BSAP Occasional Publication ◽

10.1017/s0263967x0003295x ◽

1998 ◽

Vol 22 ◽

pp. 306-308

Author(s):

M. D. Carro ◽

E. L. Miller

Keyword(s):

Protein Synthesis ◽

Liquid Phase ◽

Solid Phase ◽

Liquid Fraction ◽

Microbial Protein ◽

Microbial Protein Synthesis ◽

Continuous Culture System ◽

The One

The estimation of rumen microbial protein synthesis is one of the main points in the nitrogen (N)-rationing systems for ruminants, as microbial protein provides proportionately 0.4 to 0.9 of amino acids entering the small intestine in ruminants receiving conventional diets (Russell et al., 1992). Methods of estimating microbial protein synthesis rely on marker techniques in which a particular microbial constituent is related to the microbial N content. Marker : N values have generally been established in mixed bacteria isolated from the liquid fraction of rumen digesta and it has been assumed that the same relationship holds in the total population leaving the rumen (Merry and McAllan, 1983). However, several studies have demonstrated differences in composition between solid-associated (SAB) and fluid-associated bacteria in vivo (Legay-Carmier and Bauchart, 1989) and in vitro (Molina Alcaide et al, 1996), as well in marker : N values (Pérez et al., 1996). This problem could be more pronounced in the in vitro semi-continuous culture system RUSITEC, in which there are three well defined components (a free liquid phase, a liquid phase associated with the solid phase and a solid phase), each one having associated microbial populations.The objective of this experiment was to investigate the effect of using different bacterial isolates (BI) on the estimation of microbial production of four different diets in RUSITEC (Czerkawski and Breckenridge, 1977), using (15NH4)2 SO4 as microbial marker, and to assess what effects any differences would have on the comparison of microbial protein synthesis between diets.This study was conducted in conjunction with an in vitro experiment described by Carro and Miller (1997). Two 14-day incubation trials were carried out with the rumen simulation technique RUSITEC (Czerkawski and Breckenridge, 1977). The general incubation procedure was the one described by Czerkawski and Breckenridge (1977) and more details about the procedures of this experiment are given elsewhere (Carro and Miller, 1997).

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