Trained study staff collected samples from volunteers at an outpatient clinic after obtaining an informed consent. The study was open to patients and hospital staff not taking antiplatelet or anticoagulant medications. Volunteers with diabetes, morbid obesity, renal disease, or liver disease were excluded. Blood samples were collected in tubes containing sodium citrate (3.5mL, 3.2% sodium citrate, Greiner Bio-One).
Sodium citrate
Manufactured by Greiner
Sourced in Austria, United States, United Kingdom
Sodium citrate is a chemical compound used as a laboratory reagent. It is a white, crystalline powder that serves as a buffer solution, maintaining a stable pH in various applications. Sodium citrate is a versatile compound that helps control acidity and promote specific chemical reactions in laboratory environments.
Automatically generated - may contain errors
Lab products found in correlation
Vacuette, by Greiner (2 mentions)
Vacuette tube, by Greiner (2 mentions)
Hemavet 950fs, by Drew Scientific (2 mentions)
L glutamine, by Thermo Fisher Scientific (2 mentions)
Culture flask, by Corning (2 mentions)
Low glucose dmem, by Thermo Fisher Scientific (2 mentions)
Acid citrate dextrose solution a, by Greiner (1 mentions)
Nunc cryotube, by Thermo Fisher Scientific (1 mentions)
Xn 9000, by Sysmex (1 mentions)
Lithium heparin, by Greiner (1 mentions)
Rbc lysis buffer, by Solarbio (1 mentions)
Polypropylene tube, by Thermo Fisher Scientific (1 mentions)
Dioc6, by Thermo Fisher Scientific (1 mentions)
K2edta, by Greiner (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
Microplate reader, by Agilent Technologies (1 mentions)
Human mmp1 elisa kit, by Abcam (1 mentions)
21 protocols using sodium citrate
1
Volunteer Outpatient Blood Collection
2
Plasma Thrombin Potential and Thromboelastography
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Samples for analysis of plasma thrombin potential by CAT were collected preoperatively from the cubital vein, without stasis, at arrival at the clinic and for thromboelastography by ROTEM before induction of anesthesia from the arterial line. Platelet-poor plasma for CAT analysis and fibrinogen concentration was prepared from 3.5-mL vacutainer tubes containing 3.2% sodium citrate (Greiner Bio-One GmbH, Kremsmünster, Austria), delivered to the laboratory within 2 hours, spun at 2000g for 20 minutes, and stored in 125-µL aliquots at –80°C until analysis. Fibrinogen plasma concentration was analyzed with Clauss method with STA Liquid Fib reagent (STAGO Diagnostica & Roche, Düsseldorf, Germany). The coefficient of variances for the method was 7%. Whole blood samples for ROTEM were collected in 3.5-mL vacutainer tubes containing 3.2% sodium citrate (Greiner Bio-One GmbH), maintained at 37°C, and analyzed within 30 minutes.
3
Allogeneic Stem Cell Donor Blood Sampling
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Venous blood samples from the allogeneic stem cell donors were collected (A) prior to G-CSF stimulation at the time of the pre-transplant evaluation (median 20.5 days before apheresis). For the three study groups undergoing apheresis, blood samples were also drawn (B) in the morning immediately before apheresis, (C) immediately after apheresis, and (D) approximately 24 h after start of apheresis. All venous blood samples from allotransplant recipients were collected between 07:00 and 09:00. Samples for plasma preparation were collected into Vacuette 9NC tubes and samples for cell preparation into acid-citrate-dextrose solution A (ACD-A) tubes with sodium citrate and acid-citrate-dextrose solution A as anticoagulants (Greiner Bio-One GmbH, Kremsmünster, Austria). Samples from stem cell allo- and autografts and platelet concentrates were transferred to plastic tubes without additives.
4
FXIII Deficiency Protocol: Blood Collection
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Peripheral blood was collected into vacutainers containing 3.2% sodium citrate (Greiner Bio-one) on 2 separate occasions from patients with FXIII deficiency (n = 2) with informed consent and ethical approval obtained from the Oxford Biobank (ORB Research Tissue Bank ethics approval 09/H0606/5+5).
Patient 1 was a female of Polish ethnicity aged 45 years on appointment (appt) 1 and 46 years on appt 2. Last FXIII treatment was given 25 days prior to appt 1 and 21 days prior to appt 2. FXIII was given in the form of 1250 units of fibrogammin. Phenotype was severe FXIII deficiency with multiple bleeding episodes, which was diagnosed in childhood. Baseline FXIII level was 2% (0.02 IU/mL). Platelet count (250 × 109/L) and mean platelet volume (9.6 fL) were within the normal ranges.
Patient 2 was a male of Pakistani ethnicity aged 21 years on appt 1 and 22 years on appt 2 who was not related to patient 1. Last FXIII treatment was given 22 days prior to appt 1 and 19 days prior to appt 2. FXIII was given in the form of 2000 units of fibrogammin. Phenotype was severe FXIII deficiency with multiple bleeding episodes, which was diagnosed in infancy. Baseline FXIII level was 4 % (0.04 IU/mL). Platelet count (244 × 109/L) and mean platelet volume (9.6 fL) were within the normal ranges.
Patient 1 was a female of Polish ethnicity aged 45 years on appointment (appt) 1 and 46 years on appt 2. Last FXIII treatment was given 25 days prior to appt 1 and 21 days prior to appt 2. FXIII was given in the form of 1250 units of fibrogammin. Phenotype was severe FXIII deficiency with multiple bleeding episodes, which was diagnosed in childhood. Baseline FXIII level was 2% (0.02 IU/mL). Platelet count (250 × 109/L) and mean platelet volume (9.6 fL) were within the normal ranges.
Patient 2 was a male of Pakistani ethnicity aged 21 years on appt 1 and 22 years on appt 2 who was not related to patient 1. Last FXIII treatment was given 22 days prior to appt 1 and 19 days prior to appt 2. FXIII was given in the form of 2000 units of fibrogammin. Phenotype was severe FXIII deficiency with multiple bleeding episodes, which was diagnosed in infancy. Baseline FXIII level was 4 % (0.04 IU/mL). Platelet count (244 × 109/L) and mean platelet volume (9.6 fL) were within the normal ranges.
5
Arterial Blood Sampling Protocol
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Arterial blood samples were collected from each patient at admission and 2, 4, 6, and 8 hours thereafter. If the patient did not have an arterial cannula, venous samples were obtained.22 (link) After a discard tube, blood was drawn into citrated tubes (Vacuette, 3.5 mL; sodium citrate, 0.109 mol/L; Greiner Bio-One, Kremsmünster, Austria) and centrifuged (2,500g for 15 minutes in room temperature [RT]) within 15 minutes. The supernatant was immediately transferred to sterile polypropylene tubes (NUNC CryoTubes; Thermo Fisher Scientific, Waltham, MA) and stored at −80°C. Venous blood from 20 healthy volunteers was obtained in 2014 and processed according to the same protocol.
6
Clotting and Fibrinolysis Kinetics of Leukemic Cells
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Venous blood was drawn by using antecubital venipuncture with a 21-gauge butterfly needle in a Vacuette tube coated with 3.2% sodium citrate (Greiner Bio-One). Platelet-poor plasma (PPP) was obtained by gradient centrifugation (200g for 15 minutes, 1500g for 20 minutes). Clotting and fibrinolysis were performed as previously described.22, (link)23 (link) PPP was mixed with leukemic cells and incubated with CaCl2 (25 mM) and thrombin to initiate clotting with/without tissue-type plasminogen activator (0.5 μg/mL). The final reaction volume was 100 μL in a 96-well plate. Clot formation was monitored by turbidity at 405 nm at room temperature for 2 hours (Infinite Pro 20 plate reader; Tecan Group Ltd., Männedorf, Switzerland). The time to peak was defined as the time to obtain the turbidity plateau. Lysis time was the gap between time to peak and the curve back to the baseline. Anti-CD44 antibodies were used to perform an inhibition assay.
7
Comparative Hematology: Citrate vs. EDTA
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Whole blood was collected on EDTA and citrated tubes. The anticoagulant in the used tubes were sodium citrate: 3.2%, 0.109 M (Greiner). Complete blood count was performed using XN‐9000 (Sysmex, Villepinte, France). The platelet measurement, with impedancemetry and fluorimetry was performed. The immature platelet fraction (IPF) and absolute immature platelet count (A‐IPC) was determined with a fluorescent method. This method improves the gating of platelets, using side fluorescence (reflecting RNA content), side scatter (intracellular structure), and forward scatter (cell sizer). For thrombocytopenic patients, a blood smear was performed to evaluate EDTA clumping.
The citrate sample was diluted with anticoagulant at a ratio of 1:10 and assessed using the following ratio: analysis with citrate sample/analysis with EDTA sample.
Each sample was kept at room temperature and analyzed at 30 min, 1, 2, 4, 6 and 24 h.
To evaluate no technical error with dilution effect of citrate versus EDTA, we performed haematocrit citrate/haematocrit EDTA ratio.
The citrate sample was diluted with anticoagulant at a ratio of 1:10 and assessed using the following ratio: analysis with citrate sample/analysis with EDTA sample.
Each sample was kept at room temperature and analyzed at 30 min, 1, 2, 4, 6 and 24 h.
To evaluate no technical error with dilution effect of citrate versus EDTA, we performed haematocrit citrate/haematocrit EDTA ratio.
8
Blood Sample Collection Protocol
9
Preparation of Platelet-Derived Microparticles
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For preparation of platelets blood from 4 female healthy donors (age 26–33 years) was obtained. Venous blood was collected in Vacuette tubes containing 3.8% sodium citrate (Greiner Bio-one GmbH, Austria) using a 21-gauge needle (Becton Dickinson Austria GmbH, Austria) without applying venostasis. The first 3 ml of blood were discarded. Platelet-poor plasma was prepared by centrifugation at 1,500xg for 10 min at 20°C. Aspirated supernatants were centrifuged again to obtain PFP (1,500xg, 10 min, 20°C) containing MPs. This PFP from individual donors was stored in aliquots (250 μl each) at -70°C [46 (link)].
10
Platelet Isolation and Characterization
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Whole blood was collected from subjects via venipuncture of the antecubital vein into vacutainers containing sodium citrate (3.2%; Greiner Bio‐One). Platelet‐rich plasma (PRP) was obtained through centrifugation at 200 g for 10 min with no brake. The PRP was then treated with acid citrate dextrose (2.5% sodium citrate tribasic, 1.5% citric acid, and 2.0% D‐glucose) and apyrase (0.02 U/ml) and centrifuged at 2000 g for 10 min with no brake, thereby pelleting the platelets. The platelets were resuspended in Tyrode's buffer (10 mM N‐2‐hydroxyethylpiperazine‐N9‐2‐ethanesulfonic acid, 12 mM sodium bicarbonate, 127 mM sodium chloride, 5 mM potassium chloride, 0.5 mM monosodium phosphate, 1 mM magnesium chloride, and 5 mM glucose), and brought to a concentration of 3 × 108 platelets/ml, which was confirmed using a complete blood cell counter (Hemavet 950FS; Drew Scientific).
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