Filter Buffy Coats (FBC): a source of peripheral blood leukocytes recovered from leukocyte depletion filters.
In compliance with federal regulations, blood banks routinely use leukocyte depletion filters to eliminate contaminating leukocytes from blood products such as red blood cell and platelet concentrates. We developed and optimized conditions to elute leukocytes adsorbed to these filters; resulting in leukocyte suspensions which we termed Filter Buffy Coats (FBCs). These Filter Buffy Coats can replace standard buffy coats for various research applications. After optimizing both the filter elution medium as well as elution protocols, we compared commonly used leukocyte depletion filters from four different manufacturers. Relative fractions as well as total recoveries of leukocyte subsets, such as lymphocytes, monocytes and granulocytes, found in Filter Buffy Coats were identified and compared among the filters as well as to standard buffy coats and whole blood. Flow cytometric analysis of Filter Buffy Coats confirmed the presence of T- and B-lymphocytes, NK cells and monocytes. Furthermore, a significant quantity of CD34(+) hematopoietic stem or progenitor cells (HSC/HPC) was detected in Filter Buffy Coats prepared from different filters, thus making FBCs a valuable source for research on HSC/HPC. Colony assays revealed that most of these CD34(+) cells are functional. Using immunomagnetic cell sorting (MACS), we isolated a variety of leukocyte populations from FBC mononuclear cells (Filter-PBMCs) including T lymphocytes (CD4(+), CD8(+), CD3(+)), B lymphocytes (CD19(+)), NK cells (CD56(+)), HSC/HPC (CD34(+), CD133(+)) or dendritic cells (BDCA-4(+)). Functional properties of Filter-PBMCs, as well as of some of these isolated leukocyte populations, were confirmed using standard assays. In summary, Filter Buffy Coats are a valuable and convenient source of different peripheral leukocyte populations and can replace standard buffy coat preparations for research applications.
Meyer
T P H
TP
Blood Donor Service, Bavarian Red Cross (BRK Blutspendedienst), Munich, Germany. t.meyer@blutspendedienst.com
Zehnter
I
I
Hofmann
B
B
Zaisserer
J
J
Burkhart
J
J
Rapp
S
S
Weinauer
F
F
Schmitz
J
J
Illert
W E
WE
D015703:Antigens, CD / Q000032:analysis; D001402:B-Lymphocytes / Q000737:chemistry / Q000166:cytology; D002021:Buffers; D002452:Cell Count; D002469:Cell Separation / Q000295:instrumentation / Q000379:methods; D003114:Colony-Forming Units Assay; D003713:Dendritic Cells / Q000166:cytology / Q000276:immunology; D005434:Flow Cytometry; D006098:Granulocytes / Q000166:cytology; D006412:Hematopoietic Stem Cells / Q000737:chemistry / Q000166:cytology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D018189:Immunomagnetic Separation; D007694:Killer Cells, Natural / Q000737:chemistry / Q000166:cytology; D047589:Leukocyte Reduction Procedures / Q000295:instrumentation*; D007963:Leukocytes, Mononuclear / Q000737:chemistry / Q000166:cytology*; D008213:Lymphocyte Activation / Q000276:immunology; D007959:Lymphocyte Culture Test, Mixed; D009000:Monocytes / Q000737:chemistry / Q000166:cytology; D016176:T-Lymphocyte Subsets / Q000737:chemistry / Q000166:cytology / Q000276:immunology
D015703:Antigens, CD; D002021:Buffers