Garvan Flow Cytometry Instrument talk. Given by Dr. Katharina Winnemöller, Research and Clinical Sales Manager (NSW, ACT) at the Garvan Instituteof Medical Research on the 15th October 2010.
AutoMacs Pro magnetic Cell Seapartion - "the faster way to ultra-pure, viable cells"
Glomerular Filtration rate and its determinants.pptx
Automacs pro talk @ Garvan Flow
1. MACS cell separation - the faster way to ultrapure, viable cells
+
Dr. Katharina Winnemoeller
katharinaw@miltenyibiotec.com.au
2. About us
Germany's largest independent, privately owned biotech
company
1300 employees in 19 countries
Cell biologists, biochemists, biophysicists, clinicians, and
engineers
all acting as a community
20% of staff in R&D, engineering & clinical development
Manufacturing on site
minimal reliance on external suppliers
3. About us
Founded in
Germany in 1989
World market leader
in magnetic cell
separation (MACS)
Cologne
(Bergisch Gladbach)
corporate headquarters
18 000+ publications
with products from
Miltenyi Biotec
12. Next generation MicroBeads:
CD4+CD25+ T reg Isolation Kit, human
CD4+CD25+CD127dim/– Regulatory T Cell Isolation Kits
CD4+ T Cell Isolation Kit II, human, mouse
CD8+ T Cell Isolation Kit II, human, mouse
Pan T Cell Isolation Kit II, human, mouse
Plasmacytoid Dendritic Cell Isolation Kit II, human
New Pan Monocyte Isolation Kit, human
NK Cell Isolation Kit II, mouse
Naive Pan T Cell Isolation Kit, human (optional TCR γ/δ T cell depletion)
13. Whole Blood MicroBead Technology
Convenience standardized
Positive cell isolation directly
from peripheral blood
Automated with the autoMACS Pro
Complete walkaway automation
Higher recovery
No RBC lysis
No density centrifugation
14. A peek into the future (mouse cells):
New Kits
Pan-DC Enrichment Kit, mouse
Pan-DC Isolation Kit, mouse
PDC Isolation Kit II, mouse
Classical DC Isolation Kit, mouse
Monocyte Isolation Kit (BM), mouse
Monocyte Isolation Kit (WB), mouse
17. Automated multisample cell sorting
Designed and manufactured in Germany
Up to 6 samples
Option for automated labeling
Sample cooling racks
Volumes: 0.25-50 mL (for 5, 15, 50 mL sample tubes)
Samples stay at 4 degrees
Choice of 12 separation programs
18. Re-usable columns
For 14 days, or up to 100 separations
High capacity – cell sorting within a few minutes
105 up to 4 x 109 total nucleated cells
up to 2 x 108 magnetically labeled cells
up to 15 mL of whole blood
21. autoMACS Pro programs
Positive selection
Possel
positive selection in standard mode for isolation of cells with normal
antigen expression and frequencies higher than 5%
4 mL/min
Possel_s
positive selection in sensitive mode for isolation of cells with low
antigen expression and frequencies higher than 5%;
1 mL/min
Posseld
positive selection in standard mode I, double-column program for
isolation of rare cells in low elution volume
4 mL/min
Posselds
positive selection in sensitive mode, double-column program for
isolation of rare cells with low antigen expression
1 mL/min
Posseld2
positive selection in standard mode II, double-column program for
isolation of rare cells
4 mL/min
Posselwb
special positive selection in double-column program for isolation
of cell subsets from whole blood
4 mL/min
22. autoMACS Pro programs
Depletion
Deplete
Depletion in standard mode for removal of cells with normal to high
antigen expression and results in better target cell yield.
4 mL/min
Depletes
depletion in sensitive mode I for removal of cells with low antigen
expression and results in better target cell purity.
1 mL/min
Depl05
depletion in sensitive mode II for removal of cells with low antigen
expression and results in stringent depletion of cells
0.5
mL/min)
Depl025
depletion in sensitive mode III for removal of cells with low antigen
expression and results in stringent depletion of cells
0.25
mL/min)
A_depl7
special depletion in standard mode for removal of cells with normal
antigen expression; used for large sample sizes where magnetically
labeled cells exceed autoMACS Column
binding capacity (2x108).
depends
A_deplS7
special depletion in sensitive mode for removal of cells with low
antigen expression; used for large sample sizes
4 mL/min
24. CD4 MicroBeads, human
autoMACS™
program
Start
percentage
CD4
CD4
Purity
% CD4
cells in
negative
fraction
POSSEL
39,3%
97,9%
16,7%
2.5 min
POSSEL_S
39,3%
94,7%
3,2%
5 min
POSSELD
39,3%
99,6%
17,3%
7 min
POSSELDS
39,3%
97,2%
2,21%
8.5 min
CD4-Cy5 stained sample
Results analysed by flow cytometry
By Courtesy of A. Sheffold, A. Thiel - Berlin, Germany
time
25. Whole Blood CD14 MicroBeads, human
Anticoagulated whole blood; Purity: > 98%
program Posselwb /Rinse
26. CD34 MicroBead Kit, human
Original
sample
Human cord blood,
Purity > 95%
Program POSSELD / Rinse
Count
7-AAD
0.37%
CD34+
cells
Count
7-AAD
95.38%
CD34+
cells
CD34-PE
CD34-PE
CD34+ cells
autoMACS Separator,
by Courtesy of
Dr. Haberstock,
Dr. Allam, Hautklinik
Bonn, Germany
28. Daily Maintenance
Priming: Program Rinse (automated before first separation)
Shut down: Program Sleep for overnight storage (fill with Ethanol)
Periodic Maintenance
Exchange Columns: Every 2 weeks or after 100 separations, whatever comes first
Clean pump syringe: Every 1-2 month
Long term storage; (substitute columns)
Service – Service engineer and spare parts located in
Sydney
29. autoMACS Pro - the best way for automated magnetic
cell separation
Close to 400 Products for cell separation
Most flexibility
positive or negative selection
true Whole Blood separation, PBMC, other sources
Prettiest instrument!
30. autoMACS Pro demo – experience the difference
From November 1st you can try out the autoMACS Pro by yourself
Until November 15th
I will be around regularly to help you with your separation
Let me know what you would like to separate
please let me know by tomorrow morning
I will order the respective samples