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Ficoll plaque plus

Manufactured by Merck Group
Sourced in Austria, United States

Ficoll-Plaque Plus is a laboratory product manufactured by Merck Group. It is a density gradient medium used for the separation and isolation of cells and other biological particles based on their density. The product provides a consistent and reliable method for the purification of a variety of cell types, including mononuclear cells, lymphocytes, and stem cells, from complex biological samples.

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3 protocols using ficoll plaque plus

1

Immunomodulatory Effects of Canine ASC-EVs

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RAW 264.7 cells, a murine macrophage-like cell line, and DH82 cells, a canine macrophage-like cell line, were purchased from the Korean Cell Line Bank (Seoul, Korea). RAW 264.7 and DH82 cells were seeded in 6-well plates (1 × 106 cells/well) in triplicate and incubated for 24 h. After adherence to the plates was confirmed, the RAW 264.7 and DH82 cells were treated with LPS (200 ng/mL; Sigma-Aldrich) or control for 24 h. Similarly, with the consent of the owners, 10 mL of blood was obtained from each of 3 dogs and canine peripheral blood mononuclear cells (cPBMCs) were obtained using ficoll-plaque PLUS (Sigma-Aldrich) according to the manufacturer’s instructions. cPBMCs were seeded in 6-well plates (1 × 106 cells/well) in triplicate, incubated for 24 h, and exposed to Con A (5 μg/mL) or control for 24 h. Next, the medium was removed and replaced with media containing EVs (50 μg/well) derived from naive (unstimulated) or primed (stimulated) cASCs. Next, the cells were incubated for 48 h and then harvested for RNA extraction and immunofluorescence analysis.
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2

Isolation and Expansion of T Cells from Dry AMD Patients

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This research followed the tenets of the Declaration of Helsinki, and protocols for this study were approved by the by the Internal Research Board of Taipei Veterans General Hospital (Board No. 2013-11-012B). All samples were obtained after patients had given informed consent. PBMC was isolated from five patients with dry AMD and two unaffected control using Ficoll-Plaque Plus (Amersham Biosciences) according to the manufacturer's protocol. The characteristics of 5 patients with AMD were summarized in Figure 1C. In brief, one ratio of blood sample was layered on one ratio of Ficoll-Plaque Plus, pellet (400 × g, 30 min at 20°C) and the buffy coat was collected, washed twice with PBS and cultured in DMEM (Sigma) (100 IU/mL of penicillin, 100 μ g/mL of streptomycin [Flowlab] and 10% v/v Fetal Bovine Serum [FBS] [PAA, Austria]). T cells were expanded in freshly prepared AIM-V Medium (Invitrogen) supplemented with pen/strep/glutamine (Invitrogen) plus 300 IU/ml rhIL2 (Peprotech) and 10 ng/ml soluble anti-CD3 antibody (eBioscience, OKT3 clone) (Berger et al., 2003 (link)).
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3

PBMC Isolation and Culture Protocol

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This study was carried out in compliance with the Helsinki Declaration. Peripheral blood mono-nuclear cells (PBMCs) were isolated from six donors whose written informed consents were obtained in accordance to the guidelines of the Institutional Review Board (Taipei Veterans General Hospital). This study was approved by our Institutional Review Board (VGHIRB 2014-05-002C). The patients were all documented as deceased prior to our data review. In brief, one ratio of blood sample was layered on one ratio of Ficoll-Plaque Plus, and the pellet (400g, 30 minutes at 20°C) and buffy coat were collected, washed twice with phosphate buffered saline (PBS), and cultured in Dulbecco's modified Eagle's medium (DMEM; Sigma-Aldrich Corporation, St. Louis, MO, USA) with 100 IU/mL of penicillin, 100 μg/mL of streptomycin and 10% v/v fetal bovine serum (PAA, Pasching, Austria).
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