Application Note
USD2976(2)
Expansion and Characterization of Mesenchymal
Stem Cells on Pall SoloHill® Microcarriers
Table of Contents
1. Introduction ..................................................................................................................................................3
2. Materials and Methods................................................................................................................................4
2.1 Culturing of MSCs ..................................................................................................................................4
2.2 Attachment studies..................................................................................................................................4
2.3 Microcarrier spinner studies ....................................................................................................................4
2.4 Trypsinization from microcarriers ..............................................................................................................5
2.5 Stem cell marker visualization ..................................................................................................................5
2.6 Determining differentiation potential ........................................................................................................5
3. Results .........................................................................................................................................................6
3.1 Characterization on flatware ....................................................................................................................6
3.2 Attachment studies..................................................................................................................................6
3.3 Nuclei counts ..........................................................................................................................................8
3.4 Stem cell marker expression ....................................................................................................................9
3.5 Expansion on microcarriers....................................................................................................................10
3.6 Investigating differentiation ....................................................................................................................10
4. Conclusions................................................................................................................................................11
5. References .................................................................................................................................................12
2
1.
Introduction
Mesenchymal stem cells (MSCs) are self-renewing cells that differentiate into several terminally differentiated
cell types. These cells have been isolated from multiple sources such as bone marrow, adipose tissue,
peripheral blood, and other adult tissues(1-6). The interest in these cells is that they hold the potential to
cure disease and are being pursued in clinical trials. Three emerging fields of interest for stem cells are
cell therapy, regenerative medicine and screening of candidate drugs. In many cases, poor correlation
between efficacy of candidate drugs in animal models and humans is observed. This leads to high
attrition rates of candidate drugs from the developmental pipeline and also contributes to large losses
in revenue spent on animal model testing.
MesenchymalStemCellsseededatlowdensityandexpandedfor8daysonSoloHill’sPlasticMicrocarrier.Cellswerestainedwith
DAPI(blue)andFITC-labelledphalloidin(green)forvisualization
The ability to isolate, expand, and differentiate human stem cells invitro will streamline drug testing by
allowing candidate drug testing on human cells at early stages thereby better predicting how human
populations may react to new and developing drugs. It is hoped that the ability to reproducibly isolate
and expand these cell types will facilitate the identification of candidate drugs earlier in the development
process. In addition, the ability to differentiate stem cells into various cell lines should allow for more
relevant toxicity testing. These achievements should ultimately lead to overall cost savings and
decreased health risks in the future.
In addition to drug product testing, several clinical trials have been initiated using stem cells in cell
therapy treatments. Research has shown stem cell characteristics such as differentiation potential,
angiogenic potential, immunosuppression, or immune-privilege may be effective in the treatment of many
diseases. Clinical trials using stem cells for the treatment of osteoarthritis, spinal cord injuries, Parkinson’s
disease, ischemia due to stroke, cardiac arrests, or diabetes, are seeing promising results. However, for
toxicology screening and cell therapy applications, large numbers of cells are needed. Expansion of adult
stem cells is difficult since they have a finite life span and pluripotency can be lost. Two-dimensional (2D)
culture systems such as t-flasks, cell cubes/factories, and roller bottles are common production platforms for vaccine and biologics manufacturing as well as cell therapy. These systems are typically used
for expansion of cells to seed large bioreactors. Although well-established, these formats occupy a large
footprint, are labor intensive and are susceptible to contamination problems due to numerous open handling steps. Microcarriers offer a large surface area for growth of anchorage-dependent cell types, and
could thereby facilitate use of bioreactors for stem cell expansion in fewer passages.
In this application note we characterized MSC expansion on flatware and five SoloHill microcarriers
(Collagen, C102-1521; Plastic, P102-1521; Plastic Plus, PP102-1521; Pronectin◆ F, PF102-1521;
and Hillex® II, H112-170) in stirred vessels. Retention of multipotency of the MSCs expanded in stirred
culture was verified by immunostaining with stem cell specific antibodies and by assessing their ability
to differentiate into osteocytes and adipocytes.
www.pall.com/biopharm
3
2.
Materials and Methods
2.1
Culturing of MSCs
Human bone marrow-derived MSCs (Passage 1) were purchased from EMD Millipore (SCR108)
and expanded in spinners or on flatware in DMEM (GIBCO◆ 11054) supplemented with 10%
fetal bovine serum (FBS) (HyClone◆ SH30071.03), 2 mM L-glutamine (HyClone SH30034.02),
penicillin/streptomycin (ATCC 30-2300), and basic Fibroblast Growth Factor (bFGF) (EMD
Millipore◆ GF003). Unless otherwise noted, medium refers to this complete formulation.
For growth experiments on flatware, MSCs were cultured on Corning◆ T-flasks (430825,
430639, and 430641). To subculture cells, medium was decanted and cells were rinsed once
with Dulbecco’s phosphate buffered saline (DPBS; HyClone SH30028.03). The DPBS was
immediately decanted and 1-3 mL of TrypLE◆ Select (Life Technologies 12563) was added
(depending on T-flask size). Flasks were incubated at 37 °C until cells detached (5-8 minutes).
The cells were re-suspended with medium and then centrifuged at 300 g for 5 minutes to pellet
the cells. Medium and trypsin were decanted. Cells were re-suspended in 2-5 mL plus 10% FBS
but without bFGF (volume depends on T-flask size and number) and counted using trypan blue
stain and a Nexcelom Cellometer◆ with associated software. Fresh T-flasks were seeded
at 3x103 cells/cm2. Fresh bFGF was added (8 ng/mL) to seeded T-flasks, which were then
incubated at 37 ºC ± 0.5 with 5% CO2 in complete medium. 100% media exchanges (with
fresh bFGF) were performed every other day beginning on the second day of culture .
4
2.2
Attachment studies
For initial microcarrier attachment studies 200,000 cells were seeded onto the equivalent of
seven cm2 of each microcarrier type. Although this seeding density of ~3x104 cells/cm2 was
higher than anything used subsequently, this density was used to provide enough cells for
counting and visualization on the microcarriers. Cells were incubated with microcarriers in 1mL
of medium (either +/- FBS) in 1.5 mL Eppendorf◆ tubes at 37 ºC ± 0.5 with 5% CO2. At various
time points, tubes were removed from the incubator and microcarriers were allowed to settle.
20 µL samples of the supernatant were taken for counting on the Nexcelom counter. Time
courses for percent cells attached versus unattached were determined for each condition
and plotted (Figure 2).
2.3
Microcarrier spinner cultures
For growth experiments on the various SoloHill microcarriers, 0.5 g of microcarriers were used
per 50 mL of medium in each Corning brand 125 mL spinner vessel (Fisher Scientific 10-203B).
All microcarriers were prepared according to manufacturer’s instructions by autoclaving at 121 ºC
in deionized water. Spinner cultures were essentially performed as described in a previous
microcarrier protocol(7). Briefly, spinners were seeded in complete medium low protein
concentration (less than 0.25% FBS and no bFGF) for 30 minutes for initial attachment. After
30 minutes > 85% of cells had attached to the microcarriers. Final protein concentrations of
10% FBS and 8ng/μL bFGF were added slowly to prevent any osmotic shock from the serum.
The spinners were incubated at 37 ºC ± 0.5 ºC with 5% CO2. Cell counts were performed using
standard assays to quantify cell numbers and determine viability. Spinners containing cells on
Hillex II microcarriers were grown at 60 rpm, whereas, all other microcarrier spinners were kept
at 40 rpm. Media exchanges of 25 mL (50% volume) were performed every other day beginning
on the second day of culture. Samples were retrieved daily for nuclei counts using the citric
acid/crystal violet method. Nuclei were counted using the Nexcelom counter. The number of
nuclei per cm2 surface area was calculated for each sample.
2.4
Trypsinization from microcarriers
For trypsinization of cells from microcarriers, cells/microcarriers were allowed to settle and
medium was removed. Cells and microcarriers were washed with DPBS for five minutes at room
temperature with occasional rocking back and forth by hand to re-suspend microcarriers. After
five minutes, the microcarriers were allowed to settle and the DPBS was removed. Five (5) mL
of TrypLE Select was added. The spinners were gently pipetted once or twice to thoroughly
mix and then incubated at 37 ºC for 10-15 minutes (with occasional rocking by hand). Cells and
microcarriers were pipetted after five minutes and again after ten minutes to achieve a single-cell
suspension that could be used to reseed fresh microcarriers.
2.5
Stem cell marker visualization
To visualize the expression of several stem cell markers on MSCs expanded on SoloHill microcarriers, samples were transferred from the spinners into 15 mL tubes. Once the microcarriers
settled, medium was removed and cells/microcarriers were carefully washed with DPBS for five
minutes at room temperature. Once cells and microcarriers settled, DPBS was removed and
cells and microcarriers were fixed in 4% paraformaldehyde for ten minutes at room temperature.
The paraformaldehyde was removed and cells were washed in DPBS and stored at 4 °C until
use. To visualize stem cell markers, 250 µL of each sample was transferred to a 1.5 mL tube,
microcarriers settled and DPBS removed. Non-specific binding was blocked by incubation
with 5% FBS in DPBS for one hour at room temperature. Samples were washed in 500 μL
DPBS three times for five minutes at room temperature. Samples were then incubated in 250 µL
of the dye/antibody solutions. All antibodies were used at 1:1000 except for Stro-1 (1:500). Dyes
and antibodies used were: DAPI (Life Technologies, D3571), phalloidin-FITC (Life Technologies,
A12379), FITC anti-human CD44 (BioLegend 338803), APC anti-human CD90 (BioLegend
328113), Alexa Fluor 647 anti-human Stro-1 (BioLegend 340103), FITC anti-human CD18
(BioLegend 302105), FITC anti-human CD19 (BioLegend 302205), Alexa Fluor◆ 647 anti-human
CD14 (BioLegend 325611), and Alexa Fluor 647 anti-human CD146 (BioLegend 342005).
2.6
Determining differentiation potential
To determine differentiation potential of MSCs expanded on SoloHill microcarriers, spinners were
seeded at 3x103 cells/cm2 and grown for eight days. Spinners were subsequently passaged into
new spinners or flatware (24 well plate) at 3x103 cells/cm2. Spinners at passage 2 on microcarriers
were allowed to expand until near-confluency (2-3x104 cells/cm2). Samples from the spinners
were transferred to a 24 well plate to determine differentiation capabilities on microcarriers
compared to cells grown on flatware.
Growth/expansion medium was removed and 1 mL of either osteogenesis induction medium
(EMD Millipore SCR028) or adipogenesis induction medium (EMD Millipore SCR020) was added.
Induction and maintenance media were changed according to EMD Millipore’s protocol (as
recommended by supplier). Osteocyte differentiation was determined by Alizarin Red S
staining and adipocyte differentiation was determined by Oil Red O staining (protocols with
EMD Millipore kits).
www.pall.com/biopharm
5
3.
Results
3.1
Characterization on flatware
To characterize MSCs on flatware, T-25s were seeded at 3x103 cells/cm2 and incubated
for up to ten days to generate growth curves (Figure 1). As shown in Figure 1, MSCs seeded
at 3x103 cells/cm2 MSCs reached a maximum confluent density of ~4x104 cells/cm2. Over the
course of the ten day growth curve, cells had an average doubling time of about 48 hours.
Figure 1
MSCGrowthCurveonFlatware
4.5
Cell Density (x 104 cells/cm2)
4
3.5
3
2.5
2
1.5
1
0.5
0
0
2
1
3
4
5
6
7
8
Cellsseededat3x10 cells/cm weregrownfor10days.Dataispresentedasmeans±SEM(n=3).
3
2
Figure 2
MSCAttachmentStudies:RemovingFBSincreasedattachmentratetoallSoloHillmicrocarriers
insemi-staticconditions.
B.
80
Collagen
60
Plastic
40
Plastic Plus
Pronectin F
20
100
Percent Attachment
100
Percent Attachment
A.
80
Collagen
60
Plastic
40
Plastic Plus
Pronectin F
20
Hillex II
0
0 30 60 90 120 150 180 210 240
Attachment Time
Time (Minutes)
3.2
6
Hillex II
0
0
5 10 15 20 25 30 35 40
Attachment Time
Time (Minutes)
Attachment studies
To determine initial attachment conditions for MSCs to the SoloHill microcarriers, attachment
studies in which FBS was removed from the attachment media were performed as described
earlier. As shown in Figure 2A and 2B, MSCs were 70-80% bound to all microcarriers after
15 minute incubations at 37 °C. However, with 10% FBS present in the medium, attachment
ranged from 30-80% after two hours and from 50-90% after four hours. Observations under
light microscopy after 30 minutes of incubation at 37 °C supported these cell counts (Figure 3).
Since these experiments were performed under semi-static conditions, the faster attachment
rates in the conditions with low FBS were chosen for future spinner cultures.
To determine the growth capabilities on SoloHill microcarriers, spinner cultures were seeded
at 3x103 cells/cm2. The attachment was done in low serum concentration conditions for
30 minutes. As shown in Figure 4, after this 30 minute attachment period in the spinner flasks,
very few cells remain unbound. The low seeding density of 3x103 cells/cm2 is approximately
2-3 cells/bead. Some microcarriers were observed to have more than three cells and some
had no cells attached.
Figure 3
MSCAttachmentStudyImage
After30minutesofattachmentat37°C,highpercentagesofcellsremainedunboundtomicrocarriers(leftcolumn).
DecreasingFBSconcentrationsinmediumincreasedattachmentrateandalmostnocellswerevisibleinmedium
after30minutes(rightcolumn).
MSC attachment to microcarriers in spinner cultures: When seeded at low densities, low serum
attachment conditions led to quick attachment, although not completely uniform attachment.
Figure 4
MSCattachmenttomicrocarriersinspinnercultures.
At this low seeding density, uniform attachment was not possible in medium with 10% FBS.
However, cells attached to approximately 70% of the microcarriers allowing good expansion
over the 10 day growth period.
www.pall.com/biopharm
7
3.3
Nuclei counts
Figure 5 shows the nuclei counts for spinner samples over the ten day growth periods. Nuclei
density reached between 6-10 x104 nuclei/cm2. MSC density on microcarriers appeared to
reach a higher maximal confluent density than what was seen in T-flask growth.
Figure 5
NucleiCountsforMSCSpinnerCultures
14
Cell Density (x 104 cells/cm2)
12
10
Collagen
8
Plastic
6
Plastic Plus
Pronectin F
4
Hillex II
2
0
0
2
4
6
8
10
Days
Nucleicountsshowmaximalconfluentdensitiesbetween6and10x104 nuclei/cm2.Datapresentedismeans±SEM
(n=3).
As shown in Figure 6, cells grown on all microcarriers tended to stretch across two or three
microcarriers (with Hillex II microcarriers as the exception). By stretching between multiple
microcarriers, cells could effectively have a larger available three-dimensional volume in which
to grow, which is not possible on 2D surfaces.
Figure 6
MSCExpansioninSpinnerCulture
MSCstendedtoclumponcehigherdensitieswerereachedinmicrocarrierspinnercultures(Days5-10).
To increase initial attachment, bFGF was removed from the medium during attachment
(complete medium with low FBS and without bFGF). After 30 minutes all cells appeared
attached to greater than 90 percent of microcarriers. Expansion for seven days verified
uniform attachment and excellent growth, shown in Figure 7.
8
To verify the stem cell-like character of MSCs grown on SoloHill microcarriers, the expression
of MSC-associated cell surface markers were determined as well as the ability of these cells to
differentiate after being expanded on microcarriers. To check for the expression of cell surface
markers, samples were incubated with fluorophore-conjugated antibodies. Shown in Figure 8,
imaging on a Nikon (Ti65) determined these cells to be CD44+, CD90+, Stro1+ and CD146+
when grown on all five SoloHill microcarriers. The hematopoetic cell markers CD14 and CD19
were not expressed in MSCs (data not shown).
Figure 7
UniformAttachmenttoMicrocarriers
UsinglowFBSconcentrationandnobFGFmediumduringattachmentincreasedattachmentefficiencyfrom
~75%to>95%.
3.4
Stem cell marker expression
Figure 8
StemCellMarkerExpressionofMicrocarrier-ExpandedMSCs
A)MSCsexpandedonSoloHillmicrocarriers(collagenshown)wereincubatedwithanti-CD44(green),anti-CD90(red)
andDAPI(blue).PopulationsofCD44expressingcellsareindicatedbygreenarrows.PopulationsofCD90expressing
cellsareindicatedbyredarrows.Populationsofcellsexpressingbothmarkersareindicatedbyyellowarrows.B)MSCs
wereincubatedwithanti-CD44(green),anti-Stro1(red),andDAPI(blue).PopulationsofCD44expressingcellsare
indicatedbygreenarrows.PopulationsofStro-1expressingcellsareindicatedbyredarrows.Populationsofcells
expressingbothmarkersareindicatedbyyellowarrows.
www.pall.com/biopharm
9
3.5
Expansion on microcarriers
To determine the potentiality of MSCs when expanded on microcarriers, cells were grown
on Plastic microcarriers for multiple passages. MSCs were seeded at 3 x 103 cells/cm2 and
expanded in spinner flasks for 8 days. Cells were trypsinized from microcarriers and seeded
onto flatware for differentiation into adipocytes and osteocytes. After 21 days cells grown
on plastic microcarriers were able to differentiate into adipocytes and osteocytes at a level
comparable to cells grown on T-flasks alone (Figures 9A and 9B).
Figure 9
PotentialityofMicrocarrier-ExpandedMSCs
A)MSCsgrownonT-FlasksandPlasticmicrocarriersremaininanundifferentiatedstate.B)MSCsgrownonT–flasks
andplasticmicrocarriersunderwentadipogenesisandosteogenesis.Cellsexpandedonmicrocarriersappearedto
havesimilardifferentiationcapabilitiescomparedtocellsexpandedonT-flasksonly.
3.6
10
Investigating differentiation
To determine if several passages on microcarriers affected the differentiation ability of MSCs,
cells were passaged multiple times on plastic microcarriers. After six passages on microcarriers,
cells were seeded onto flatware for differentiation. Table 1 shows that over the six passages on
microcarriers, the harvesting density was consistently above 3 x 104 cells/cm2 with a doubling
time of 48-51 hours when seeded at 3 x 103 cells/cm2, showing that multiple passages on
microcarriers did not decrease maximal confluent density or doubling rates. The ability of cells
grown on microcarriers for six passages is shown in Figure 10. The cells grown on plastic
microcarriers appear undifferentiated (top panels). Additionally, these cells were able to
differentiate into both adipocytes and osteocytes (bottom panels) similar to levels seen
previously on earlier passages and on flatware (Figure 9).
Figure 10
Differentiation
MSCsgrownonplasticmicrocarriersformultiplepassagesareshownundifferentiatedinthelefttwopanels,and
differentiatedintoadipocytesandosteocytesinthetworightpanels.
Table 1
ContinuousPassagesonMicrocarriersinSpinnerCulture
Plastic Spinners
P1
P2
P3
P4
P5
P6
Seeding Density
(x104 cells/cm2)
Harvest Density
(x104 cells/cm2)
Days of Growth
Number of Doublings
0.3
2
1
0.3
0.3
0.3
4.6
3.3
2.7
3.6
4.3
4.5
8
4
5
<1
5
1.3
7
3.6
8
3.8
8
3.9
SeveralpassagesonmicrocarriersdemonstratetheabilitytoexpandMSCscontinuouslyonmicrocarrierswithout
decreasingmaximalconfluentdensityordoublingtimewhenseededat3x103 cells/cm2.
4.
Conclusions
Current obstacles limiting the use of stem cells for therapeutic benefits include a limited number of cell
divisions and the potential loss of pluripotency. Due to the restricted number of population doublings,
achieving maximal possible expansion in the fewest passages is vital. We have shown here that MSCs
can be expanded on various types of SoloHill microcarriers. The benefit of MSC expansion on microcarriers is two-fold. First, expansion on microcarriers allows growth on large surface areas within single
containers, and second microcarrier expansion increases the ratio of apparent surface area to medium
volume due to the fact that MSC growth on microcarriers outpaces growth on flatware.
This is particularly important with stem cells grown in medium that contains expensive supplements.
Therefore, the use of microcarriers allows minimal passages for expansion of cells while decreasing
the overall cost required to grow enough cells for a therapeutic dose in clinical trial treatments. We have
shown that multiple passages on microcarriers do not affect the ability of MSCs to differentiate into
adipocytes and osteocytes. The ability to maintain pluripotency while expanding MSCs on microcarriers
for five or six passages allows for the isolation of cells from bone marrow onto a T-150 flask. Cells
expanded in this fashion can subsequently seed a small scale spinner culture which could be used
to seed a small bioreactor. For example, the maximal confluent cell density in a T-150 results in 2.5-3 x
106 cells and 3.5-4 x 104 cells/cm2 on microcarriers. To seed a 200 mL spinner volume requires 3.1x106
cells using 5150 cm2/L. The maximal densities on spinner cultures achieved here would result in enough
cells for a minimum 10-fold expansion into a 2 L bioreactor, by the third passage after isolation and the
second passage on microcarriers.
Considering the recoverable cell numbers presented here, a 6.7 L bioreactor volume (at 5150 cm2/L)
would result in enough cells for one therapeutic dose (~1 x 109 cells). Assuming similar growth between
small scale spinners and bioreactors, a 2 L bioreactor could be used to seed a 20L bioreactor, which
would result in enough cells for three doses from a single T-150 and multiple passages on microcarriers.
Additionally, increasing the microcarrier concentration beyond 5150 cm2/L would decrease bioreactor
volume required for a large number of recoverable MSCs. Table 2 extrapolates the expected number of
cells if the Plastic microcarrier concentrations used were 10 g/L and 20 g/L. The resulting liters needed
for therapeutic doses of stems cells would be 7-8 liters and 3-4 liters, for concentrations of 10 g/L and
20 g/L respectively.
www.pall.com/biopharm
11
Table 2
TherapeuticDoses
Microcarrier
Concentration
(Plastic)
Surface Area
Max Confluent
Density (cells/cm2)
Total Cells
Per Liter
Liters Per
Therapeutic Dose
10 g/L
20 g/L
3600cm2
7200cm2
3.5-4 x 104
3.5-4 x 104
13-14 x 107
25-29 x 107
7.1-7.8
3.4-4
ExtrapolatedlitersneededfortherapeuticdosesforvariousconcentrationsofPlasticmicrocarriercultures.
Work will continue to further characterize stem cells grown on SoloHill microcarriers, such as defining
various populations within bone marrow-derived MSCs, any fluctuations in these populations when
grown on microcarriers for multiple passages, and additional stem characteristics such as the ability of
microcarrier-expanded cells to produce angiogenesis signals such as VEGF. Future work will also expand
to include other stem cells such as induced-pluripotent stem cells, mouse embryonic stem cells, and
mesenchymal stem cells from other sources such as placental-derived or adipose-derived. Additionally,
work will continue to define growth and expansion conditions under animal component free environments,
as well as direct expansion on microcarriers of isolated stem cells, thereby bypassing flatware tissue
culture and increasing the number of available passages before senescence.
5.
References
1. Friedenstein AJ, Gorskaja JF, Kulagina NN. Fibroblast precursors in normal and irradiated mouse
hematopoietic organs. Exp Hematol 1976, 4: 267-274.
2.Fraser etal. Fat tissue: an underappreciated source of stem cells for biotechnology. Trends Biotechnol
2006, 24: 150-154.
3.Cao C, Dong Y. Study on culture and in vitro osteogenesis of blood-derived human mesenchymal
stem cells. Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2005. 19: 642-647.
4. Griffiths Mu, Bonnet D, Janes SM. Stem Cells of the Aveolar epithelium. Lancet. 2005, 366: 249-260.
5. Beltrami etal. Adult cardiac stem cells are multipotent and support myocardial regeneration. Cell.
2003, 114: 763-776.
6. Pittenger etal. Multilineage potential of adult human mesenchymal stem cells. Science. 1999, 284:
143-147.
Visit us on the Web at www.pall.com/biopharm
E-mail us at microcarriers@pall.com
Customer Ordering
Pall Life Sciences
4370 Varsity Drive, Suite B
Ann Arbor, MI 48108
microcarriers@pall.com
734-973-2956 phone
Corporate Headquarters
Port Washington, NY, USA
+1.800.717.7255 toll free (USA)
+1.516.484.5400 phone
biopharm@pall.com e-mail
European Headquarters
Fribourg, Switzerland
+41 (0)26 350 53 00 phone
LifeSciences.EU@pall.com e-mail
Asia-Pacific Headquarters
Singapore
+65 6389 6500 phone
sgcustomerservice@pall.com e-mail
International Offices
Pall Corporation has offices and plants throughout the world in locations such as: Argentina,
Australia, Austria, Belgium, Brazil, Canada, China, France, Germany, India, Indonesia, Ireland,
Italy, Japan, Korea, Malaysia, Mexico, the Netherlands, New Zealand, Norway, Poland, Puerto
Rico, Russia, ingapore, South Africa, Spain, Sweden, Switzerland, Taiwan, Thailand, the United
Kingdom, the United States, and Venezuela. Distributors in all major industrial areas of the
world. To locate the Pall office or distributor nearest you, visit www.pall.com/contact
The information provided in this literature was reviewed for accuracy at the time of publication.
Product data may be subject to change without notice. For current information consult your
local Pall distributor or contact Pall directly.
© 2015, Pall Corporation. Pall,
, Hillex and SoloHill are trademarks of Pall Corporation.
® indicates a trademark registered in the USA and TM indicates a common law trademark.
Filtration.Separation.Solution. is a service mark of Pall Corporation. ◆ ProNectin is a
trademark of Sanyo Chemical Industries, Ltd.; EMD Millipore is a trademark of KGaA;
GIBCO is a registered trademark of Life Technologies Corporation; HyClone is a trademark
of Hyclone Laboratories, Inc.; TrypLE is a trademark of Life Technologies Corporation;
Cellometer is a trademark of Nexcelom Bioscience, LLC; Corning is a trademark for Corning
Corporation; Eppendorf is a trademark of Eppendorf AG; Alexa Fluor is a trademark of
Molecular Probes, Inc.
2/15, PDF, GN15.6111
USD2976(2)