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Magnetic isolation
of cells & molecules
with MACS® Technology
Thuesday:
MACS® Technology for magnetic isolation of cells
and molecules
• Introduction
• Features of paramagnetic MicroBeads
• General procedure of magnetic cell isolation
• Overview of applications in molecular biology
µMACS epitope tagged protein isolation
• Expression of tagged/fusion proteins, e.g. GFP fusion
proteins
• Magnetic protein isolation
Wednesday:
• Detection of proteins, results and trouble-shooting
• Optimizing protein expression analysis by transfected cell
selection (MACSelect)
Cell separation with MACS® Technology
MD0347.01
Miltenyi Biotec GmbH (headquarter)
MACS® Magnetic Cell Sorting
Magnetic
labeling with
MACS
MicroBeads
Elution of
positively
selected
cells
Flow through with
unlabeled cells
MD0651.01
MACS® Technology
Equipment and reagents
» MACS MicroBeads
» MACS Columns
» MACS Magnet
MD0197.02
MiniMACS™ separator
MD0343.02
Magnetic field generated in an MS Column
MD0061.02
Major advantages of MACS® MicroBeads
• Extremely small beads (50 nm Ø)
=> Colloidal suspension, non-sedimenting
=> fast reaction kinetics => short incubation times
• Super-paramagnetic
• Biodegradable & non - toxic
=> Straight to experiment or cell culture
=> Cell function preserved
• Flow cytometry compatible
=> No bead detachment required
=> Only 20-30% of binding sites occupied
=> Decreased flow sorting time +MACS® separation
CD8+ T cells isolated by MACS® Technology
MD0058.03
CD8+ T cells isolated by MACS® Technology
MD0057.03
1/1 MD0xxx.01
MACS® isolated dendritic cell
MD0159.02
Positive selection strategy
Magnetic
labeling with
MACS®
MicroBeads
Elution of
positively
selected
cells
Flow through with
unlabeled cells
MD016102
Isolation of human NK cells
from PBMC using CD56 MicroBeads
CD56-FI TC
2.73%
CD56-FI TC
Positive fraction
Relative cell number
17.86%
Negative fraction
Relative cell number
Relative cell number
Before separation
89.51%
CD56-FI TC
MD0158.02
Depletion strategy
Magnetic labeling
Isolation of
untouched cells
MD0160.02
Isolation of untouched B cells
from PBMC using B Cell Isolation Kit
Before separation
Untouched B cells
95.86%
CD19-PE
CD19-PE
11.4%
CD45-FITC
CD45-FITC
MD0147.02
Indirect MicroBeads
Anti-Fluorochrome MicroBeads
Anti-FITC, Anti-PE, Anti-APC
Streptavidin MicroBeads
Anti-Biotin MicroBeads
Anti-Immunoglobulin
MicroBeads
MD0xxx.01
autoMACS™ Separator
For automated cell separation including automated elution
and system cleaning
• Touch screen
• Integrated computer
• Uptake port
• Elution ports
• Solution bottles
• Sample rack
MD0423.02
The CliniMACSTM System
• CD34+ cell selection:
hematopoetic precursor cell
• CD133+ cell selection:
(hematopoetic) stem cell
Depletion of T cells
(allogenic, GvHD)
or tumor cells
(autologous)
1/1 MD0xxx.01
MACS® separation strategies: NO LIMITS!
• Positive selection and depletion
• Isolation of cell subsets
• Isolation using any antibody
• Isolation and detection of secreting cells
• Isolation of molecules
MD0044.02
MACSmolecular
Specific protein isolation
• µMACS Protein A and Protein G MicroBeads for
immunoprecipitation
• µMACS Epitope Tag Protein Isolation Kits
• µMACS Streptavidin Kit
mRNA isolation & cDNA synthesis
• µMACS mRNA Isolation Kits
• µMACS One-step cDNA Kit
Enrichment of transfected cells
• MACSelect
PIQOR™ gene expression profiling technology
• Microarray & Bioinformatics Service
cDNA Synthesis from small sample material
500 cells, µMACS One-step cDNA
500 cells mRNA + RT in tube*
5 cells, µMACS One-step cDNA
5 cells, mRNA + RT in tube*
Negative control
Jurkat cells
*mRNA + RT in tube:
competitor (magnetic) mRNA
isolation kit and conventional
in-tube synthesis
LightCycler quantitative PCR
with intron-spanning primers
for 87 bp GAPDH fragment, 10
and 20 % of each cDNA was
used.