Cobe 2991

Manufactured by Terumo BCT

Sourced in United States

The COBE 2991 is a laboratory equipment product from Terumo BCT. It is designed for the separation and collection of blood components. The COBE 2991 utilizes centrifugation technology to fractionate whole blood into its constituent parts, such as red blood cells, platelets, and plasma. The device is capable of processing a variety of blood volumes and sample types. The COBE 2991 is intended for use in research and clinical laboratory settings.

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Lab products found in correlation

Dithizone, by Merck Group (2 mentions) Biocoll, by Biochrome (1 mentions) Rpmi 1640 solution, by Thermo Fisher Scientific (1 mentions) Liberase mtf, by Roche (1 mentions) Thermolysin, by Roche (1 mentions) Fluorescein diacetate propidium iodide fda pi, by Merck Group (1 mentions) Glutamine, by Lonza (1 mentions) Pen strep, by Lonza (1 mentions)

Spelling variants of this product

COBE 2991 Cell Processor (6 citations)

6 protocols using cobe 2991

Islet Purification Using Iodixanol Gradient

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Twenty-five milliliters or less of digested tissue were layered over an iodixanol-based continuous density gradient (20 ), ranging from 1.060 to 1.100 ± 0.01 g/mL, and were centrifuged at 1,800–2,000 rpm in a COBE 2991 cell processor (Terumo BCT, Lakewood, CO) at 4–8°C. Twelve 25-mL gradient fractions were collected in 250 mL conical tubes prefilled with 225 mL CMRL 1066, Supplemented, CIT Modification Solution (Mediatech, Manassas, VA) (38 ). Islet purity from each fraction was estimated by DTZ staining. Purified fractions were classified as high (≥70%), middle (40–69%), and low (30–39%) purity, and those with the same purity were pooled. If >50,000 islets were present in fractions with purity <30%, supplementary purification was performed using Biocoll (Biochrome AG, Berlin, Germany) (25 ), polysucrose discontinuous gradients (Mediatech) (24 ), or OptiPrep (Nycomed/Takeda, Osaka, Japan) (26 ). Purity of the fractions obtained by supplementary purification was assessed by DTZ staining, and each fraction was combined with fractions of equivalent purity to those obtained by using the iodixanol-based gradient. From this point forward, the high-, medium-, and low-purity fractions were processed separately and combined only after culture and immediately before filling the infusion bags.

Ricordi C., Goldstein J.S., Balamurugan A.N., Szot G.L., Kin T., Liu C., Czarniecki C.W., Barbaro B., Bridges N.D., Cano J., Clarke W.R., Eggerman T.L., Hunsicker L.G., Kaufman D.B., Khan A., Lafontant D.E., Linetsky E., Luo X., Markmann J.F., Naji A., Korsgren O., Oberholzer J., Turgeon N.A., Brandhorst D., Chen X., Friberg A.S., Lei J., Wang L.J., Wilhelm J.J., Willits J., Zhang X., Hering B.J., Posselt A.M., Stock P.G, & Shapiro A.M. (2016). National Institutes of Health–Sponsored Clinical Islet Transplantation Consortium Phase 3 Trial: Manufacture of a Complex Cellular Product at Eight Processing Facilities. Diabetes, 65(11), 3418-3428.

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Pancreatic Islet Transplantation Protocol

Cited 2 times

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TP-IAT was performed according to previously published protocols.^{8 (link),12 (link)} Briefly, the decision to proceed with TP-IAT was made on clinical grounds by a multidisciplinary team. On surgical excision of the pancreas, the blood supply to the pancreas was maintained until pancreatic mobilization was completed to minimize warm ischemia time and maximize islet preservation. After recovery, the pancreas was immediately transported to the University of Minnesota Molecular and Cellular Therapeutics good manufacturing practice facility. Islet isolation was performed using enzymatic digestion and mechanical dispersion as previously described.^{26 (link)} After digestion, the islets were purified using density gradients in a COBE 2991 cell processor (Terumo BCT, Lakewood, CO) if needed to reduce transplanted tissue volume (generally performed for a postdigest tissue volume >0.25 mL/kg).^{27 (link)} The final islet tissue preparation was transplanted by intraportal infusion during a 15- to 60-min period. In all cases, all or a majority of islets were transplanted intraportally, and a small proportion of islets were transplanted elsewhere (peritoneum, omentum) if portal pressures were elevated (typically above ~25 cm water).

Nanno Y., Hodges J.S., Freeman M.L., Trikudanathan G., Schwarzenberg S.J., Downs E.M., Ramanathan K., Pruett T.L., Beilman G.J., Chinnakotla S., Hering B.J, & Bellin M.D. (2023). Early Metabolic Measures Predict Long-term Insulin Independence in Recipients of Total Pancreatectomy and Islet Autotransplantation. Transplantation Direct, 10(1), e1561.

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Isolation and Purification of Pancreatic Islets

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A collagenase solution containing liberase MTF (0.5 g per 1 vial) and thermolysin (15 mg per 1 vial) (Cat#05339880001; Roche CustomBiotech, Penzberg, Germany) was instilled into the disinfected pancreas via the catheter placed in the pancreatic duct. The distended pancreas was cut into several pieces and then placed into a Ricordi Chamber. The digestion was started by commencing the gentle shaking of the Ricordi chamber, while warmed collagenase solution was circulated. When the digestion was stopped, the digested tissue was diluted in RPMI 1640 solution (Cat#11875085; Gibco^™) containing 10% inactivated plasma (Fetal Bovine Serum, qualified, United States, Cat#26140079; Gibco^™) and ulinastatin and then collected in Belzer UW^® Cold Storage Solution. The purification process was performed using IBM 2991 (COBE 2991; Terumo BCT, Tokyo, Japan) by centrifugation with a continuous density gradient between 1.077 g/cm³ and 1.100 g/cm³ created using Optiprep (Cat#ST-07820; Veritas Co., Tokyo, Japan). After centrifugation, the gradient density solutions containing highly-purified islets (≥ 70%) were collected. The purity was determined using the percentage of the total number of cell clusters staining positive for dithizone (Cat#D5130; Sigma-Aldrich, St. Louis, MO, USA).

Sakata N., Yoshimatsu G., Kawakami R., Aoyagi C, & Kodama S. (2023). Optimal temperature for the long-term culture of adult porcine islets for xenotransplantation. Frontiers in Immunology, 14, 1280668.

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Isolation and Purification of Human Islet-Derived Extracellular Vesicles

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The use of human specimens (supernatant of cultured islets or islets discarded from clinical use) was approved by Institutional Review Board. Islet-free supernatants were obtained from freshly purified human islet preparations (n = 10) from Diabetes Research Institute (HSR-DRI), San Raffaele Scientific Institute, Milan, Italy. Islets were isolated and purified according to the automated method described by Ricordi, with local modifications [18] (link). Briefly, the pancreatic duct was cannulated with 14–20 G catheter and distended by intraductal infusion of a cold collagenase solution (Collagenase NB1 Premium Grade, Serva, Heidelberg, Germany). After digestion at 37°C in a modified Ricordi chamber, islets were purified on a Cobe 2991 (TerumoBCT, Lakewood, CO, USA) using a continuous HBSS-Ficoll (Biochrom, Berlin, Germany) gradient. Purified islet fractions were cultured in Final Wash (Mediatech Cellgro, VA, USA) plus 1% Pen/Strep, 1% Glutamine (Lonza, Basel, Switzerland), counted and their numbers expressed as number of islets normalized to a 150-µm diameter (IEQ). Final islet preparations were cultured at a density of 1000±150 IEQ/ml.
Human islet preparations were selected for purity (>70%) and viability (>70%) for EV isolation by Dithizone (Sigma-Aldrich, St. Louis, MO) and Fluorescein Diacetate/Propidium Iodide (FDA/PI; (Sigma-Aldrich, St. Louis, MO) dye analysis.

Figliolini F., Cantaluppi V., De Lena M., Beltramo S., Romagnoli R., Salizzoni M., Melzi R., Nano R., Piemonti L., Tetta C., Biancone L, & Camussi G. (2014). Isolation, Characterization and Potential Role in Beta Cell-Endothelium Cross-Talk of Extracellular Vesicles Released from Human Pancreatic Islets. PLoS ONE, 9(7), e102521.

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Automated Washing of Transfused RBCs

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For transfusions deemed necessary by the clinical team, units were handed to the study team from the 4 red cell units typically provided in a cooler, under ice, for all cardiac cases. These were washed on-demand using a continuous automated transfusion system (CATS, Fresenius Kabi USA LLC, Lake Zurich, IL) with the FDA-approved 0.9% saline wash solution, following a 4:1 dilution with 0.9% saline in the cell-saver reservoir, as previously described.^{12 (link)} Pre-dilution was included to reduce the hematocrit of packed RBCs to better approximate that of the salvaged blood mixed with heparin anticoagulant that cell-savers were designed to process. This automated washing procedure has been shown to more efficiently remove contaminants from the supernatant.^{12 (link),13 (link)} This cell-saver system was chosen as the g-force applied to the cells is less for this apheresis belt system (~800G) than for the Latham bowl design favored by most cell-saver devices (~2000 G),^{14 (link)} and has been reported to induce less hemolysis when compared to a standard blood bank based cell washing device (Cobe 2991, Terumo BCT, Lakewood, CO).^{15 (link)} Staff trained to use this cell-saver included research personnel, anesthesia technicians and perfusionists (Duke), and transfusion medicine personnel with specific expertise in cellular washing techniques (Mayo).

Welsby I.J., Norris P.J., Mauermann W.J., Podgoreanu M.V., Conn C.M., Meade L., Cannon T., Keating S.M., Silliman C.C., Kehler M., Schulte P.J, & Kor D.J. (2021). Bedside allogeneic red blood cell washing with a cell saver to remove cytokines, chemokines and cell-derived microvesicles, a clinical feasibility study.. Anesthesiology, 134(3), 395-404.

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Human Islet Isolation and Characterization

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Human islet preparations were isolated from deceased donor pancreata, as previously described using a collagenase/thermolysin enzyme mixture (Roche Diagnostics, Laval, QC, Canada) and the resulting digest suspension was purified using a modified COBE 2991 cell processor (Terumo BCT, Inc., Lakewood, CO, United States) with continuous density gradients 2,10-12 . The purified islet fraction(s) were assessed for clinical suitability and cultured in CMRL 1066-based medium at 22°C (5% CO2) for up to 72 hours prior to transplantation.

Gala-Lopez B.L., Neiman D., Kin T., O'Gorman D., Pepper A.R., Malcolm A.J., Pianzin S., Senior P.A., Campbell P., Glaser B., Dor Y., Shemer R, & Shapiro A.M.J. (2018). Beta Cell Death by Cell-free DNA and Outcome After Clinical Islet Transplantation. Transplantation, 102(6).

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