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Cfu hill medium

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CFU-Hill medium is a culture medium used for the enumeration of colony-forming units (CFU) of hematopoietic progenitor cells. It is designed to support the growth and proliferation of various types of hematopoietic progenitor cells, including granulocyte-macrophage progenitors (CFU-GM) and burst-forming units-erythroid (BFU-E).

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Lab products found in correlation

Fibronectin coated dishes, by BD (2 mentions) Ficoll paque, by BD (2 mentions) Fibronectin, by BD (1 mentions)

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CFU-Hill liquid medium

6 protocols using cfu hill medium

1

Quantification of EPC in Sternal Bone Marrow

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The EPC compartment in the sternal bone marrow was analyzed by the colony-forming unit (CFU) method [20 (link)] and by flow cytometry (below). For the CFU assay, MNCs were resuspended in CFU-Hill medium (StemCell Technologies, Vancouver, Canada), plated on 6-well plates coated with fibronectin (BD-Brazil, SP, Brazil) at a concentration of 5 × 105 cells/cm2, and cultured for 48 hours. Nonadherent cells were then collected, resuspended in the same medium, plated in duplicate samples onto fibronectin-coated, 24-wells plates at 5.2 × 105 cells/cm2, and cultured for 3 days. After staining with Giemsa, CFU-Hill units, characterized by a central cluster surrounded by elongated cells, were blindly counted and shown as median and interquartile intervals per 106 MNCs.
Ghem C., Dias L.D., Sant'Anna R.T., Kalil R.A., Markoski M, & Nardi N.B. (2017). Combined Analysis of Endothelial, Hematopoietic, and Mesenchymal Stem Cell Compartments Shows Simultaneous but Independent Effects of Age and Heart Disease. Stem Cells International, 2017, 5237634.
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2

Quantifying Early Endothelial Progenitor Cells

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From 45 mL of intravenous blood, measures of EPC-CFU were obtained by isolating early EPCs using Ficoll Paque and seeding 5 million cells on 6-well Fibronectin-coated dishes (BD Biosciences) in CFU-Hill medium (Stem Cell Technologies, cat#05900). Non-adherent cells were collected 48 hours later, and 1 million cells seeded on 24 well Fibronectin-coated dishes. On day 5, EPC-CFUs were counted in 5 wells. The mean EPC-CFU was used in analyses as previously described [14 (link), 39 (link)].
Martinez C.A., Rikhi R., Pester M.S., Parker M., Gonzalez A., Larson M., Chavez J., Mendez A., Raines J.K., Kolber M.A., Schulman I.H., Alcaide M.L, & Hurwitz B.E. (2022). Abacavir antiretroviral therapy and indices of subclinical vascular disease in persons with HIV. PLoS ONE, 17(3), e0264445.
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3

Isolation and Quantification of Endothelial Progenitor Cells

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Peripheral mononuclear cells (PMNCs) were fractionated using Ficoll density-gradient centrifugation. Isolated PMNCs were re-suspended in CFU-Hill Medium (Stem Cell Technologies, Vancouver, Canada) and plated on 6-well plates, coated with human fibronectin at a concentration of 5×106 cells per well. After 48 hours, the non-adherent cells were collected and replated onto fibronectin-coated 24-well plates. EPC colonies were counted using an inverted microscope 7 days after plating. An EPC colony was defined as a cluster of at least 100 flat cells surrounding a cluster of rounded cells, as previously described [11 (link)] . A central cluster alone without associated emerging cells was not counted as a colony. Results are expressed as the mean number of colony-forming units (CFUs) per well.
Peterson S.J., Shapiro J.I., Thompson E., Singh S., Liu L., Weingarten J.A., O’Hanlon K., Bialczak A., Bhesania S.R, & Abraham N.G. (2019). Oxidized HDL, Adipokines and Endothelial Dysfunction: A Potential Biomarker Profile for Cardiovascular Risk in Women with Obesity. Obesity (Silver Spring, Md.), 27(1), 87-93.
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4

Isolation and Quantification of EPCs

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Peripheral blood samples were obtained from patients before and three months after MSC injection. EPCs were isolated from samples using Ficoll-Paque and five million cells were seeded on 6-well fibronectin-coated dishes (BD biosciences) in CFU-Hill medium (stem cell technologies, cat#05900) (Hill et al., 2003 (link), Solomon et al., 2012 (link)). The non-adherent cells were collected 48 h later and one million cells were seeded on 24-well fibronectin-coated dishes. On day five, EPC-CFUs were counted in five wells and the average was obtained.
Premer C., Blum A., Bellio M.A., Schulman I.H., Hurwitz B.E., Parker M., Dermarkarian C.R., DiFede D.L., Balkan W., Khan A, & Hare J.M. (2015). Allogeneic Mesenchymal Stem Cells Restore Endothelial Function in Heart Failure by Stimulating Endothelial Progenitor Cells. EBioMedicine, 2(5), 467-475.
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5

Quantifying Endothelial Progenitor Cells

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Peripheral mononuclear cells (PMNCs) were fractionated using Ficoll density-gradient centrifugation. Isolated PMNCs were resuspended in CFU-Hill Medium (Stem Cell Technologies, Vancouver, Canada) and plated on 6-well plates, coated with human fibronectin at a concentration of 5 × 106 cells per well. After 48 hours, the nonadherent cells were collected and replated onto fibronectin-coated 24-well plates. EPC colonies were counted using an inverted microscope 7 days after plating. An EPC colony was defined as a cluster of at least 100 flat cells surrounding a cluster of rounded cells, as previously described [12 (link)]. Results are expressed as the mean number of colony-forming units (CFUs) per well.
Fakhouri E.W., Weingarten J.A., Singh S.P., Shah P, & Peterson S.J. (2021). The Association of Nephroblastoma Overexpressed (NOV) and Endothelial Progenitor Cells with Oxidative Stress in Obstructive Sleep Apnea. Oxidative Medicine and Cellular Longevity, 2021, 7138800.
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6

Evaluating Endothelial Progenitor Cells

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Endothelial progenitor cell-colony forming units were used to assess endothelial function and assays were performed as previously described (Premer et al., 2015 (link)). Briefly, EPC-CFUs were determined by isolating EPCs from peripheral blood samples, plating them on 6-well fibronectin-coated dishes followed by 24-well fibronectin-coated dishes in CFU-Hill medium (Stem Cell Technologies, cat #05900), and counting EPC-CFU formation on day five.
Premer C., Wanschel A., Porras V., Balkan W., Legendre-Hyldig T., Saltzman R.G., Dong C., Schulman I.H, & Hare J.M. (2019). Mesenchymal Stem Cell Secretion of SDF-1α Modulates Endothelial Function in Dilated Cardiomyopathy. Frontiers in Physiology, 10, 1182.
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