Use of Colloidal Silica (Sepracell-MN) for Enrichment of Dendritic Cells From Human Peripheral Blood: Comparison With Other Methods
IR@IICB: CSIR-Indian Institute of Chemical Biology, Kolkata
View Archive Info| Field | Value | |
| Title |
Use of Colloidal Silica (Sepracell-MN) for Enrichment of
Dendritic Cells From Human Peripheral Blood: Comparison With Other Methods
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| Creator |
Chehimi, Jihed
Kawashima, Hisashi Starr, Stuart E Hassan, Nassef F Douglas, Steven D Bandyopadhyay, Santu |
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| Subject |
Infectious Diseases and Immunology
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| Description |
Three methods are described for enrichment of dendritic cells from human peripheral
blood. In method 1 , mononuclear cells were incubated in plastic tissue culture flasks for
two h. Nonadherent cells were removed. Adherent cells were washed to remove floating
cells and incubated for 14 h at 37#{176in}C 5% CO2. Carbonyb iron was added, and the flasks
were incubated for another 2 h. Nonadherent cells were subjected to centrifugation over
metrizamide gradient. Phagocytic cells containing ingested carbonyl iron, small lymphocytes,
and free carbonyl iron particles passed through the metrizamide, while the interface
cell population was enriched for dendritic cells. The purity and yield of enriched
dendritic cells were 52.8% and 0.05%, respectively. In method 2, adherent mononuclear
cells were cultured overnight, and the released cells (released adherent cells) were centrifuged
over metrizamide to separate low-density cells. Monocytes from this low-density
cell population were removed by panning over human gamma globulin-coated plastic
Petri dishes. In this method the average purity and yield of DC were 63.8% and 0.1%,
respectively. In method 3, released adherent cells were treated with anti-CD5 and anti-
CDI4 monoclonal antibodies plus baby rabbit complement for 15 mm, washed, and centrifuged
with colloidal silica (Sepracell-MN). Centrifugation with Sepracell-MN was
repeated three times. Low-density cells were panned twice over human gamma globulincoated
plastic Petri dishes. Nonadherent cells were highly enriched for DC. Contamination
of T cells, B cells, and NK cells was undetectable by flow cytofluorometry. Contamination
of monocytes was < 2%. This method provided > 85.0% purity and 0.4% yield.
This method (method 3) combines adherence, complement-dependent lysis, centrifugation
with colloidal silica, and panning and provides the best yield and purity; it is therefore
recommended for optimal purification of DC.
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| Date |
1990
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| Type |
Article
PeerReviewed |
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| Format |
application/pdf
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| Identifier |
http://www.eprints.iicb.res.in/1918/1/J._Leukocyte_Biol%2C48%2C74%2D80_1990.pdf
Chehimi, Jihed and Kawashima, Hisashi and Starr, Stuart E and Hassan, Nassef F and Douglas, Steven D and Bandyopadhyay, Santu (1990) Use of Colloidal Silica (Sepracell-MN) for Enrichment of Dendritic Cells From Human Peripheral Blood: Comparison With Other Methods. Journal of Leukocyte Biology , 48. pp. 74-80. |
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| Relation |
http://www.eprints.iicb.res.in/1918/
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