Whole blood samples were collected by Virginia Department of Agriculture and Consumer Services (VDACS) staff beginning in August 2018 with five sites (three Northern, one Central, one Southwest) and scaled up, in January 2019, with seven additional sites (one Northern, three Central, three Southwest). These included 11 of the 12 regularly scheduled, weekly Virginia Livestock Auction sites as well as one additional weekly auction. To gain wide geographic coverage with continuity over time, approximately five adult animals from different lots were sampled during each visit. Fewer samples were collected if there were less than five lots of cattle presented and when staff travel or auction sales were interrupted, mainly due to pandemic COVID restrictions. Whole blood was collected in purple-top BD Vacutainer® blood collection tubes containing EDTA anticoagulant (Becton, Dickinson and Company, Franklin Lakes, NJ, USA). Sex, breed, and radio frequency identification (RFID) tag were recorded. Breeds were also coded as groups (beef, dairy or unknown). Origin of each animal was approximated based on address of the owner, abstracted from market sales records, and geocoded using the Google Maps geocoder API (Available online: https://developers.google.com/maps/documentation/geocoding/overview , accessed on 5 August 2021).
Edta anticoagulant
Manufactured by BD
Sourced in United States
EDTA (Ethylenediaminetetraacetic acid) is an anticoagulant commonly used in laboratory settings. Its primary function is to prevent blood clotting by binding to calcium ions, essential for the coagulation process. EDTA is widely used in various blood collection and laboratory analysis procedures to ensure the integrity of the sample.
Automatically generated - may contain errors
Lab products found in correlation
Gentle macs mechanical dissociator, by Miltenyi Biotec (2 mentions)
Tumor dissociation kit, by Miltenyi Biotec (2 mentions)
Vacutainer, by BD (1 mentions)
Qubit 4 fluorometer, by Thermo Fisher Scientific (1 mentions)
Minicap, by Sebia (1 mentions)
Xn 1000, by Sysmex (1 mentions)
Vacutainer blood collection tube, by BD (1 mentions)
Vacutainer collection tube, by BD (1 mentions)
Macs tissue storage solution, by Miltenyi Biotec (1 mentions)
Vacutainer blood tubes, by BD (1 mentions)
Sodium citrate, by BD (1 mentions)
Cell lysis buffer, by Thermo Fisher Scientific (1 mentions)
Conical falcon tube, by Corning (1 mentions)
Vacutainer tube, by BD (1 mentions)
Leucosep system, by Greiner (1 mentions)
Sca 1, by Thermo Fisher Scientific (1 mentions)
Cd117, by Thermo Fisher Scientific (1 mentions)
13 protocols using edta anticoagulant
1
Longitudinal Cattle Sampling for Genetic Analysis
2
Genetic Screening for β-Thalassemia
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Patients underwent hematological testing, including CBC, Hb electrophoresis, and genetic testing, to screen for β-thalassemia. Vacutainer blood samples (4 ml) containing ethylenediaminetetraacetic acid K3 (EDTA) anticoagulant (Becton Dickinson, Franklin Lakes, NJ, USA) were drawn from the patients for routine hematological investigations, including complete blood count (CBC) using Sysmex XN-1000 (Sysmex Corporation, Kobe, Japan) and Hb electrophoresis using MINICAP (Sebia, Lisses, France). DNA was subsequently extracted from the blood samples using an automated nucleic acid extraction instrument, EZ1 (Qiagen, Hilden, Germany). A dsDNA high-sensitivity assay kit was used to measure the DNA concentration in ng/μl using the QuBit 4 fluorometer, following the manufacturer’s instructions (Thermo Fisher Scientific, USA).
The isolated DNA was amplified by multiplex PCR (Veriti Thermal 96-Well Thermal Cycler Applied Biosystems, USA) using commercial β-globin strip assay, allowing the multiplication of 22 targets of β-globin and detection using reverse hybridization (Vienna Lab Labor diagnostika GmbH, Austria). The extrapolation of the results from the membrane strip was performed using StripAssay Online Calculator v2.17. Samples were processed at the hematology and genetics departments of Biolab Diagnostic Laboratories, Amman, Jordan.
The isolated DNA was amplified by multiplex PCR (Veriti Thermal 96-Well Thermal Cycler Applied Biosystems, USA) using commercial β-globin strip assay, allowing the multiplication of 22 targets of β-globin and detection using reverse hybridization (Vienna Lab Labor diagnostika GmbH, Austria). The extrapolation of the results from the membrane strip was performed using StripAssay Online Calculator v2.17. Samples were processed at the hematology and genetics departments of Biolab Diagnostic Laboratories, Amman, Jordan.
3
Pilot Study of Smear-Positive TB in Uganda
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After obtaining permission and informed consent (see ethical considerations) a pilot study in newly diagnosed smear positive TB patients and healthy controls was conducted between the months of April and June 2013 at the Mulago National Referral and Teaching Hospital located in North of Kampala, Uganda. By consecutive sampling adult patients who presented with persistent cough for more than 3 weeks had a positive Ziehl-Nielsen smear test for the first time and signed a written consent were considered eligible. Health workers and medical trainees who worked at the TB out patient’s clinic and other wards exposed to TB were matched for sex and age with the control group. All subjects were screened for human immunodeficiency virus (HIV) and data and consent forms were filled. Blood was then collected into tubes with ethylenediaminetetraacetic acid (EDTA) anticoagulant (Becton, Dickinson and company New Jersey USA) and whole blood samples were stored at 2–8°C before DNA column extraction.
4
Cattle Blood Surveillance Across Virginia
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Whole blood samples were submitted from client-owned cattle across the state of Virginia. Animals represented clinical submissions by private veterinarians, as well as herds owned and maintained by the Virginia Department of Corrections (VADOC), and as part of an ongoing, random sampling surveillance effort of animals sent to auction by the Virginia Department of Agriculture and Consumer Services (VDACS). Whole blood was collected in purple-top BD Vacutainer® blood collection tubes containing EDTA anticoagulant (Becton, Dickinson and Company). Except for animals submitted by referring veterinarians as part of a routine clinical diagnostic workup, all animals were randomly sampled, and the presence and degree of clinical signs were unknown. In total, 1,359 blood samples were available for evaluation.
5
Pharmacokinetic Study of Doxorubicin-Loaded Nanoparticles
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Female nude mice (7–8 weeks old) were intravenously administered DOX or PLGA-PEG@DOX/anti-EGFR (5 mg/kg for DOX, n = 3/group). Samples of blood were collected retro-orbitally at 3 min, 20 min, 1 h, 2 h, 4 h, 8 h, 12 h, and 24 h post-injection in tubes containing EDTA anticoagulant (BD, Franklin Lakes, NJ, USA). Samples were spun for 10 min at 4500 rpm, after which supernatants were combined with an equal volume of acetonitrile to facilitate plasma protein precipitation. Samples were again centrifuged, after which supernatants were collected for UV-vis spectroscopy analyses.
6
Blood and Tumor Processing for Flow Cytometry
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Blood was obtained by venipuncture and collected in Vacutainer collection tubes containing EDTA anticoagulant (BD Biosciences). Blood was rotated on shaker until flow cytometry analysis (10 min-4 h post collection).
Surgically resected tumors were collected and stored in MACS tissue storage solution at 4°C (Miltenyi). Storage time was 2–16 h post-tumor collection. Tumor cells and tumor infiltrating immune cells were analyzed after tumors were processed using the Tumor Dissociation Kit and Gentle MACS mechanical dissociator (Miltenyi) following the manufacturer's recommendations. Single cell suspension was immediately analyzed by flow cytometry.
Surgically resected tumors were collected and stored in MACS tissue storage solution at 4°C (Miltenyi). Storage time was 2–16 h post-tumor collection. Tumor cells and tumor infiltrating immune cells were analyzed after tumors were processed using the Tumor Dissociation Kit and Gentle MACS mechanical dissociator (Miltenyi) following the manufacturer's recommendations. Single cell suspension was immediately analyzed by flow cytometry.
7
Multimodal Analysis of Tumor Samples
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Blood was collected on the day of surgery prior to surgical incision. Blood for flow cytometry was obtained by venipuncture and collected in Vacutainer collection tubes containing Heparin or EDTA anticoagulant (BD Biosciences). Blood was rotated on a shaker at room temperature until flow cytometry analysis (10 minutes-4 hours post collection). Blood for serum assays was collected in tubes without anticoagulant (BD Biosciences). Surgically resected tumors were collected and stored in MACS tissue storage solution at 4°C (Miltenyi). Storage time was 1–16 hours post-tumor collection. Tumor tissue was mechanically disrupted with scissors until tumor pieces were 1-2 mm. PDAC was digested using the Tumor Dissociation Kit and Gentle MACS mechanical dissociator (Miltenyi) following manufacturer’s recommendations. IPMN tissue was digested with a separate protocol which is detailed below. Tissue and digestion media (RPMI with 10% FBS, 10mM Hepes, 5mM CaCl2 1x protease inhibitor cocktail, 1x trypsin inhibitor and 100 U/ml DNase) was heated to 37°C for 20-40 minutes until homogenization was achieved. Both PDAC and IPMN tissues were filtered 3 times using a 70 μM cell strainer to remove undigested tissue and dead cell debris and resuspended in 1 ml of PBS. The resulting single cell suspension was immediately analyzed by flow cytometry.
8
Whole Blood Collection from Healthy Volunteers
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Whole blood samples from 51 healthy volunteers were collected in Vacutainer blood tubes containing EDTA anti-coagulant (BD Bio- Sciences) at the Army Medical Center. All subjects in this study provided written informed consent. The study was approved by the Medical Ethics committee of Army Medical Center in accordance with relevant guidelines and regulations. Generally, 2–3 ml of whole blood was withdrawn from each healthy volunteer within an hour of the start of each experiment.
9
Plasma Collection Tube Comparison
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Fresh blood samples were obtained from healthy individuals using three different plasma collection tubes sourced from distinct manufacturers. The collection tubes included those containing EDTA anticoagulant (BD, Franklin Lakes, NJ, USA), sodium citrate (BD, Franklin Lakes, NJ, USA), and Lakebio cell-free RNA storage tubes (Lakebio, Hefei, China). Following blood collection, each type of collection tube was individually incubated at 25 °C for designated time intervals of 0, 4, 8, 16, 24, 48, 72, and 120 h. At each specified time point, two-step centrifugation was performed to separate the plasma, which was subsequently stored at −80 °C for subsequent experiments.
10
Isolation of Bovine PBMCs
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Peripheral blood from the jugular vein of M. bovis-infected and control animals was collected in 10 ml Vacutainer® tubes containing EDTA anticoagulant (BD Diagnostics, Oxford, UK). For cell separation, whole blood was diluted 1:1 with phosphate buffered saline (PBS) in 50-ml conical Falcon tubes (Corning, Inc., Kaiserslautern, Germany). Peripheral blood mononuclear cells (PBMCs) were isolated from buffy coat fractions using Histopaque-1077 (Sigma-Aldrich Ireland Ltd., Wicklow, Ireland) and the Leucosep system (Greiner Bio-One Ltd., Stonehouse, UK) with gradient centrifugation at 1034 × g for 25 min. PBMCs were collected and washed in PBS to remove platelets. Contaminating red blood cells were removed using cell lysis buffer (Thermo Fisher Scientific, Waltham, MA, USA).
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