BERG | BioEngineering Research Group
2008
Annual Report
Contents
4
Executive Summary
5
The Bioengineering Research Group
6
Bioprocess Engineering and Biocatalysis Laboratory
8
Nucleic Acid Bioengineering Laboratory
10
Stem Cell Bioengineering Laboratory
12
Research Highlights
20
Publications
26
Oral Presentations
30
Poster Presentations
34
Prizes
35
Staff
Annual Report
2008
3
Executive Summary
This annual report is the first of the “new” BioEngineering Research Group (BERG) within the recently created Institute for Biotechnology and Bioengineering (IBB), an Associated Laboratory approved by the Ministry of Science, Technology and
Higher Education in Portugal.
During 2008 major achievements were obtained on
Bioprocess Engineering and Biocatalysis, namely:
the design of surfactant-stable cutinase mutants by
direct evolution; an enzyme technology platform for
esters biosyntheses by cutinase in nonconventional media; the use of microtiter plates as
platforms for multi-step bioconversions process
development; and the purification of human antibodies from CHO cell cultures with high yields and
purity using aqueous two-phase systems. The Nucleic Acid Bioengineering achievements include
the development of a plasmid DNA purification
chromatographic process based on DNA-amino
acid interactions and a bead-based hybridisation
assay for detection of traces of E. coli genomic
DNA present in purified plasmid DNA samples;
and the construction of DNA vaccine prototypes by
cloning parasitic antigenic proteins associated with
sleeping sickness and tested in mice models. Also
with impact on the Healthcare sector, the major
achievements on Stem Cell Bioengineering include
a high-throughput cell based screening device for
fast identification of small molecules to selectively
control mouse Embryonic Stem Cell (mESC) fate;
the development of serum-free culture systems for
the ex-vivo expansion of human Hematopoietic
Stem Cells (hHSC) in co-culture with human Mesebchymal Stem Cells (hMSC) and an integrated
culture system for mESC expansion and neural
differentiation; and the isolation and ex-vivo expan-
sion of human Mesenchymal Stem Cells (hMSC)
under GMP conditions for the treatment of graft
versus host disease, as well as adjuvant in hHSC
transplantation. The clinical trials, performed in
collaboration with Instituto Português de Oncologia
Francisco Gentil de Lisboa and Centro de Histocompatibilidade do Sul, are part of the European
Blood and Marrow Transplantation group activities
and represent a pioneer initiative in Portugal.
With the recent creation of IBB, a scientific strategy
was implemented to integrate complementary expertise in the group by hiring new Faculty members and Postdoctoral researchers in emergent
scientific areas, through contracts with Instituto
Superior Técnico, the MIT-Portugal Program and
Programa Ciência 2007. These researchers will
contribute to reach our ambitious goals and
strengthen our research at international level on
bioengineering science. I would like also to express my confidence in BERG to promote excellent
quality research and advanced education programmes, ensuring national and international competitiveness in the areas of Industrial and Health
Biotechnology.
Joaquim M.S. Cabral
BERG Head and
Director of IBB
Bioengineering Research Group | BERG
BERG
BEBL
NABL
SCBL
The BioEngineering Research Group (BERG) is a research unit in engineering and
life sciences at the Centre for Biological and Chemical Engineering (CEBQ). CEBQ is
the leading Centre of the Associated Laboratory Institute for Biotechnology and Bioengineering (IBB), a network of research centres across Portugal. IBB has been
identified by the Portuguese Ministry of Science, Technology and Higher Education
as a strategic infrastructure for the development of the Portuguese R&D and innovation policies in the areas of Biotechnology, Bioengineering, Biomaterials and Life,
Biomedical and Agricultural Sciences. BERG activities within the Associated Laboratory IBB are focused on the Thematic Areas of Industrial Biotechnology and Health
Biotechnology.
BERG aims at excellence in research and advanced education in biotechnology and
bioengineering. The overall goal is to contribute for a better understanding of the
mechanisms that occur at the molecular and cellular levels, in order to translate them
into rational applications of biological systems relevant to the Industrial and Health
care sectors. BERG research priorities have special emphasis on Bioprocessing and
Biomolecular Engineering, Gene/Nucleic Acid Bioengineering, Nanobiotechnology
and Stem Cell Engineering, featuring an integrated cross-disciplinary approach
through three laboratories:
Bioprocess Engineering and Biocatalysis Laboratory (BEBL)
Nucleic Acid Bioengineering Laboratory (NABL)
Stem Cell Bioengineering Laboratory (SCBL)
Annual Report
2008
5
Bioprocess Engineering and Biocatalysis
Objectives
Bioprocess Engineering and Biocatalysis
aims to design and develop value-added bioproducts with potential application in key areas, such as food and feed, aroma, pharmaceutical industry and biofuels. Research is
focused on the production, purification and
stabilisation of proteins/enzymes and on the
design of improved bioconversion processes.
Research Topics
Research in BEBL is currently focused on the
development of technological platforms for
biocatalysis and biomolecules purification.
The projects under study are centred in three
major areas: i) Protein Stabilisation; ii) Biocatalysis; and iii) Production and purification
of proteins and biopharmaceuticals.
1. Protein Stabilisation - Approaches to enhance the stability of proteins/enzymes, including synthetic mimetic affinity ligands and
encapsulation in biocompatible hydrogels, are
investigated in order to improve performance
and develop specific industrial and diagnostic
applications.
2. Biocatalysis - Ester biosyntheses (flavours,
biodiesel and macrocyclic esters) by cutinase
and engineered mutants are addressed in an
enzymatic platform. Nano/micro-biocatalysts
(biocomposites) are being developed based
on hydrogels, protein/cell assemblies and
nano-magnetic particles. The biocomposites
are used as nano/micro-bioreactors and their
performance is evaluated by on-line coupling
with analytical techniques (FIA/SIA) and microfluidic systems. New protein-ionicconducting-based biocompatible materials
(Ion Jelly) with tailor-made properties have
been designed to build new planar amperometric biosensors.
Mini-scale devices are used to speed optimisation of biotransformation/fermentation systems.
3. Production and Purification of Biopharmaceuticals - Alternative processes for the purification of proteins and biopharmaceuticals
(antibodies) integrating aqueous two-phase
extraction with chromatographic steps are
being developed. Extraction separation units
using affinity materials and nanomagnetic
particles are being evaluated, characterised
and validated by modelling.
Stimuli-responsive beads based on PNIPAM
are also being developed as a strategy to
produce novel bio-inspired affinity polymer
systems for antibody recognition.
Validation of microtiter plates as suitable
platforms for the characterisation of multistep bioconversions in conventional and
non-conventional media, using sitosterol
side-chain cleavage as model system.
Implementation of an effective system for
the production of inverted sugar syrup
using inulinase immobilised in PVA capsules.
Human antibodies from CHO cell cultures
were purified using aqueous two-phase
system (ATPS) comprising a temperature
responsive polymer composed of ethylene
oxide and propylene oxide (EOPO), with
high yield and purity. The use of this
smart polymer has considerably simplified
the re-extraction step of antibodies.
Selected Publications
Major Achievements
The activity and stability of cutinase and
mutants obtained by direct evolution were
evaluated and compared. The S54D mutant was much more resistant to AOT
denaturation than the native enzyme.
A new approach for protein stabilisation
was developed, through the design and
synthesis of combinatorial libraries of
ligands interacting with specific regions on
cutinase surface.
An enzyme technology platform was used
in esters biosyntheses by cutinase in nonconventional media. A novel strategy using a green chemistry approach based on
mini-emulsions has also been developed
for aroma and flavours bioproduction.
Baptista, R.P., Pedersen, S., Cabrita, G.J.,
Otzen, D.E., Cabral, J.M.S., Melo E.P., Biopolymers, 89, 538-547 (2008)
Brissos, V., Eggert, T., Cabral, J.M.S., Jaeger, K.E., Prot. Eng. Design Select., 21, 387393 (2008)
Claudino, M.J.C., Soares, D., Marques,
M.P.C., van Keulen, F., Cabral, J.M.S., Fernandes, P., Bioresource Technol., 99, 23042311 (2008)
Ferreira, I.F., Azevedo, A.M., Rosa, P.A.J.,
Aires-Barros, M.R., J. Chromatogr. A, 1195,
94-100 (2008)
Teles, F.R.R., Fonseca, L.P., Talanta, 77,
606-623 (2008)
Annual Report
2008
7
Nucleic Acid Bioengineering
Objectives
Nucleic Acid Bioengineering is focused on: i)
plasmid vectors and their application in gene
therapy or DNA vaccination; and ii) microchips for DNA detection. The specific objectives are to address the scientific and technological challenges associated with plasmid
biopharmaceuticals by combining biomolecular engineering studies with bioprocess engineering, and to co-develop (with INESC-MN)
thin-film microchip platforms for the manipulation/detection of DNA.
Research Topics
In the case of plasmids, the following research topics are pursued:
1. Structural stability of plasmids - Studies on
the nuclease barriers to gene expression
during plasmid trafficking through the cytosol
of mammalian cells are performed with the
goal of constructing plasmid vectors with an
increased resistance to nucleases and thus
with a higher transfection activity.
2. Manufacturing of plasmid vectors - Processes for the production of plasmids are conceptually designed, developed, optimised and
compared. The impact of specific plasmid
structural elements in the performance of
upstream and downstream processes
(membranes, chromatography, aqueous twophases) and in the quality of the final product
is evaluated. Analytical procedures to monitor
manufacturing and control product quality are
also developed.
3. DNA vaccine prototyping - DNA vaccine
candidates are constructed by cloning para-
sitic antigenic proteins associated with sleeping sickness disease and tested in mice models for their ability to generate cellular and
humoral responses, and to provide immunisation.
In the case of microchips for DNA detection
the following topics are addressed:
1. Immobilisation and handling of DNA - Thin
film technologies, chemical modification, microfluidics and electronic addressing are
used to develop microchips for the molecular
recognition of specific analytes via hybridisation. The core of the chips is a flat surface
with immobilised probe molecules. Other
features include the presence of microelectrodes to generate electric fields that accelerate the kinetics of binding/recognition.
2. Photodetectors - Amorphous silicon
photodetectors are developed for the optoelectronic detection of coloured, chemiluminescent and fluorescent molecules in thin film
chips. The presence of these molecules ultimately reports specific biorecognition events
such as DNA hybridisation.
Main Achievements
The feasibility of exploring DNA-amino
acid (histidine, arginine) interactions in
the context of plasmid DNA purification
by fixed-bed chromatography was demonstrated (in collaboration with UBI).
The recombination of plasmid DNA vectors harbouring direct repeats during replication in E. coli grown under increasing
antibiotic pressure was studied experi-
mentally. A simple non-linear mathematical function was developed to accurately
predict the corresponding recombination
frequencies.
A bead-based hybridisation assay was
developed for detection of traces of E.
coli genomic DNA present in purified
plasmid DNA samples
Miniaturised amorphous silicon thin-film
photodetectors were developed to quantitate the light (colorimetry, fluorescence,
chemiluminescence) generated during
assays for the detection of molecular
recognition events such as DNA hybridisation and antibody-antigen binding (in
collaboration with INESC-MN).
The possibility of using amorphous silicon-based ion-sensitive field-effect transistors (a-Si:H ISFETs) for the label-free
detection of DNA molecules was studied
in detail.
Selected Publications
Gonçalves, D., Prazeres, D.M.F., Chu, V.,
Conde, J.P., Biosensors Bioelectronics, 24,
545-551 (2008)
Martins, S., Prazeres, D.M.F., Monteiro, G.A.,
Anal. Bioanal. Chem., 391, 2179-2187 (2008)
Oliveira, P.H., Lemos, F., Prazeres, D.M.F.,
Monteiro, G.A., Plasmid, 60, 159-165 (2008)
Pimentel, A.C, Prazeres, D.M.F., Chu, V.,
Conde, J.P., J. Applied Physics., 104,
054913 (2008)
Ribeiro, S.C., Oliveira, P.H., Prazeres,
D.M.F., Monteiro, G.A., Mol. Biotechnol., 40,
252-260 (2008)
Annual Report
2008
9
Stem Cell Bioengineering
Objectives
Research Topics
The Stem Cell Bioengineering Laboratory
aims at the development of highly controlled
culture systems (e.g. bioreactors) for the exvivo expansion of stem cells and their controlled differentiation into specific cell types.
As stem cells are rare, their isolation and
expansion/differentiation in vitro significantly
increases the cell population available for
cellular and gene therapy settings, highthroughput drug screening, tissue engineering and stem cell research. Human hematopoietic stem cells (HSC), human mesenchymal stem cells (MSC), human and mouse
embryonic stem cells (ESC), and mouse neural stem cells (NSC) are used as model systems.
1. Expansion of HSC in co-culture with MSC
under serum-free conditions - Current research is focused on understanding the
mechanisms underlying the hematopoietic
supportive capacity of MSC combining proliferative, functional and proteomic analysis.
The elucidation of those mechanisms will
have implications in terms of bioreactor design towards the maximization of human HSC
expansion in vitro.
2. Clinical-scale production of MSC By combining a cross-disciplinary approach
of Stem Cell Bioengineering and Experimental Hematology, culture protocols are optimized for the expansion of human MSC,
while maintaining their multilineage and immunosuppressive capacities, for supplemen-
tation during HSC transplantation. MSC are
isolated from adult bone marrow (BM), adipose tissue (AT) and umbilical cord blood
(UCB).
3. Bioreactor expansion of ESC and NSC The expansion of ESC and ESC-derived
NSC is addressed towards the definition of
highly controlled, efficient, reproducible and
cost-effective bioprocesses to obtain starting
material to generate mature cells (i.e. neurons) for potential use in Regenerative Medicine (e.g. treatment of neurological disorders), as well as for high-throughput drug
screening. In a complementary approach,
high-throughput microarray systems are developed for studying the effect of the microenvironment on self-renewal and neural differentiation of ESC.
4. Gene delivery to stem cells - Aiming at the
maximization of the SC yield to reach meaningful cell numbers for Cell or Gene Therapy
settings or high-throughput screening assays,
efficient plasmid DNA transfection protocols
are developed using non-viral vectors for the
transient over-expression of specific genes
involved in self-renewal or lineage commitment, in collaboration with Nucleic Acid Bioengineering Laboratory.
5. Recombinant protein production for stem
cell research - A platform for recombinant
protein production and purification has been
developed, especially aiming to produce cytokines/growth factors such as Leukemia Inhibitory Factor (LIF) and Bone Morphogenetic
Protein 4 (BMP-4) for stem cell culture.
Main Achievements
● A consortium was established between IBB
-IST and Instituto Português de Oncologia
Francisco Gentil and Centro de Histocompatibilidade do Sul, focusing the isolation and exvivo expansion of MSC under GMP conditions for the treatment of graft versus host
disease, as well as adjuvant in HSC transplantation. The clinical trials already performed are part of the European Blood and
Marrow Transplantation group activities and
represent a pioneer initiative in Portugal,
since ex-vivo expanded allogenic MSC have
been infused into patients for the first time.
● A 3-D high-throughput cell based screening
device was developed for the fast identification of small molecules that can be used to
selectively control mouse ESC fate, in collaboration with Jon Dordick, RPI, USA.
● Two serum-free culture systems were developed for: i) ex-vivo expansion of human
HSC in co-culture with human MSC and ii)
the integrated expansion and neural commitment of mouse ESC.
Selected Publications
Fernandes, T.G., Kwon, S.J., Lee, M.Y.,
Clark, D.S., Cabral, J.M.S., Dordick, J.S.,
Anal. Chem., 80, 6633-6639 (2008)
Diogo, M.M., Henrique D., Cabral, J.M.S.,
Biotechnol. Appl. Biochem., 49, 105-112
(2008)
Frias, A.M., Porada, C.D., Crapnell, K.B.,
Cabral, J.M., Zanjani, E.D., Almeida-Porada,
G., Exp. Hematol., 36, 61-68 (2008)
Annual Report
2008
11
Raquel Aires-Barros and Ana M. Azevedo
The production of monoclonal antibodies (MABs)
For concentrations of NaCl higher than 10%, IgG
has been chosen as a demanding and challenging
partitioned preferentially to the top phase while
example process, since a large number of MABs
the contaminant proteins remained in the bottom
candidates in pre-clinical and clinical trials will
phase. With 15% NaCl, about 90% of the IgG was
reach process development stage in a few years,
recovered in the top PEG rich-phase. A back-
and production capacities will dramatically fall
extraction step was also performed and IgG was
short. The aim of this research is to evaluate the
recovered with a total yield of 76% and a purity of
use of aqueous two-phase systems (ATPS) as a
100%.
generic technology for the selective recovery and
purification of MABs.
The application of ATPS was expanded to the
initial recovery of human antibodies from both
Non-functionalised systems
Chinese Hamster Ovary (CHO) and hybridoma
A model system containing albumin, myoglobin
cell culture supernatants composed of PEG 6000,
and IgG was used to investigate the feasibility of
phosphate and NaCl (Fig. 2). ATPS was success-
using aqueous two-phase systems (ATPS) of
fully used to partially purify monoclonal antibodies
polyethylene glycol (PEG) and phosphate salts, to
from a hybridoma cell culture supernatant with a
recover human IgG. In order to improve the parti-
total yield of 90% and a purification factor of 4.1.
tion of IgG to the top phase different concentra-
In collaboration with Werner Bäcker from Bayer
tions of NaCl were added to the ATPS (Fig. 1).
Technology Services GmbHa, a counter-current
multi-stage extraction was performed in a mixer-
2.5
2.5
2.0
2.0
1.5
1.5
1.0
1.0
0.5
log KP
0.5
log KP
BEBL
Aqueous Two-Phase Systems for Antibody
Purification
0.0
-0.5
-1.0
0.0
-0.5
-1.0
-1.5
-1.5
-2.0
-2.0
-2.5
-2.5
-3.0
0
5
NaCl %
10
15
Figure 1: Effect of NaCl concentration on the partition
coefficient of IgG (), HSA () and Myo (), with a system composition of 7.04% PEG 6000, 14.37% phosphate
pH 7.0, 0.1% protein.
0
5
10
15
NaCl (%)
Figure 2: Effect of NaCl concentration on the partition
coefficient of IgG () and contaminant proteins (), in a
system composition of 12% PEG 6000, 10% phosphate
pH 6, 40% hybridoma feed stock.
Research Highlights
Figure 3: Multi-stage mixer-settler battery used for aqueous two phase extraction .
settler battery using the optimised ATPS based in
Many of the diseases treated by antibodies re-
PEG/ phosphate/ NaCl (Fig. 3). It was possible to
quire high doses or chronic administration, thus
run successfully this type of system in a multi-
economical process-scale production and purifica-
stage equipment with significant improvements in
tion of these molecules is critical. This work can
the recovery yield, partition coefficient and purity
contribute to important improvements in the down-
of IgG.
stream processing of antibodies, essential in order
to cut down the manufacturing costs, and to use
Functionalised systems
ATPS as a generic and efficient recovery and pu-
To improve the selectivity of aqueous two phase
rification technology on a large scale in the phar-
extraction, the performance of several functional-
maceutical industry.
ised PEG and triethylene glycol (TEG) molecules
1.5
was evaluated for the capture of human antibodies from a CHO cell supernatant. A screening of
1.0
ligands containing charged, hydrophobic and af0.5
free ligands or as phase forming components.
Among the functionalised PEG3350 molecules
(Fig. 4), promising results were obtained with glu-
log KP
finity groups was performed either in the form of
0.0
-0.5
taric acid, amino and benzyl groups, with extraction yields higher than 75% and protein purities
around 90%. Among the free ligands, TEG diglutaric acid (TEG-COOH) displayed the highest affinity towards antibodies and facilitated an extraction yield of 96% and a protein purity of 95%.
-1.0
Control
GA
NH2
Benzyl
Pym
MEP
PEG-Ligand
Figure 4: Partition coefficient of IgG in the presence of
functionalised-PEG in ATPS composed of 8% PEGligand and () 5% dextran or () 8% dextran. GA: glutaric acid; Pym: pyrimidine; MEP: mercaptoethylpyridine.
Annual Report
2008
13
BEBL
Enzymes In BioAnalytical Methods and
Monitoring Tools
Luís P. Fonseca
Rapid development of biotechnology and biotech-
Injection Analysis (FIA) systems. These analytical
nological applications in recent years resulted in
tools allow to improve process optimisation and
an increased need for methods for reliable biopro-
control according to the influence of stringent re-
cess monitoring and control. Various techniques
sponse especially to define the substrate feeding
have been investigated and proposed, but few are
strategy and induction time of biosynthesis in fer-
widely accepted and implemented due to the in-
mentation culture and enzyme yields on down-
trinsic difficulties related to complexity of samples
stream processing, namely of Expanded Bed Ad-
from bioprocesses, asepticity requirements, and
sorption columns (Fig. 1). This led to a significant
the bioprocesses themselves. Enzymes In Bio-
reduction of variability in operation and more pre-
Analytical Methods and Monitoring Tools (EIBAM-
dictable yield and purification degree of enzyme.
MT) project has stemmed on the development of
directly interfaced enzyme based amperometric
The monitoring of nutrients and metabolites in
detection devices for bioprocess monitoring. This
culture medium, (e.g. glucose, galactose, ethanol,
specific project focuses on-line monitoring of ana-
lactate, amino acids), as result of microbial cell
lyte(s), namely carbon source uptake and metabo-
activity, such as S. cerevisiae and E. coli, and
lites formation in medium cultures and enzyme
animal cells, was carried out by multi-enzyme sys-
activity (e.g. cutinase) by assessment with Flow
tems based on immobilised oxidase and horse-
Monitoring tool for EBA
adsorption at pH 4.5
Yeast cells
Sewage
vessel
On/Off Valve
Off
Sample pool
Sewage
Off
EBA column
Diluted sample
Reagent A
Fermentation
broth
Valve
Dilution
Mixing-module
Yeast cells
Clarification
module
FIA system
Sample
injector
Reagent A
Sewage
Pump A
Reaction coil
Reagent B
Pump B
Spectrophotometer
Mixing
chamber
Figure 1: Analytical tool for almost in real time monitoring cutinase activity come out from the
Expanded Bed Adsorption column.
Research Highlights
Figure 2: Mini-analytical columns of CPG
based on immobilized of one oxidase and
Figure 3: Integration of mini-analytical columns into Carrier
of the Flow Injection Analysis (FIA) system
radish peroxidase (HRP) in mini-analytical reac-
It is also expected to design new protein-ionic-
tors (Fig. 2) integrated into a FIA system (Fig. 3).
conducting-based materials with tailor-made prop-
The success of these mini-analytical reactors was
erties and biocompatibility according to the com-
due to their high sensibility and stability, inclu-
position and conditions of preparation. Accuracy
sively in complex culture media. The development
of electrochemical signals, storage and opera-
of an in-situ stabilisation strategy of oxidases
tional stability of planar amperometric biosensors
against hydrogen peroxide, which is instantane-
based on peroxidases, dehydrogenases and
ously and in-situ eliminated, represents a major
some O2 dependent oxidases (e.g. glucose, lac-
breakthrough. This stabilisation strategy is carried
tate, alcohol, among others) and antibodies conju-
out when the oxidation reaction is performed in
gated with peroxidase are being evaluated. These
the presence of peroxidase, 4-aminoantipyrine
new conducting materials, with tailor-made prop-
and phenol-4-sulfonic acid, which are reduced by
erties and efficient direct electron transfer be-
HRP to a quino-imine, a colorimetric compound
tween the biological element and the transducer,
easily monitored in the spectrophotometric detec-
can open a window of new opportunities for appli-
tor of the FIA system. The monitoring of cutinase
cations in chemistry and biology.
biosynthesis is also essential for process control
with great positive impact on process productivity
The Enzymes In Bioanalytical Methods and Moni-
and efficiency, thus reducing production costs
toring Tools (EIBAM-MT) project shows how it is
(Fig. 4).
important the use of enzymes in bioanalytical
methods and opens new perspectives on bioproc-
Recently, this project also focuses on monitoring
essing of enzymes and other biomolecules for
of environmental biological samples and water
quality by determination of total and speciation of
toxic metal elements based on enzymatic digestion of biological samples assisted by probe sonication (EPS), an emerging methodology that minimises interferences on metal determinations in
contrast with traditional analytical methodologies.
industrial
and diagnostic
applications.
Figure
4: Analytical
step-up for
monitoring of nutrients and
metabolites (e.g. glucose, galactose, ethanol, lacate, amino
acids) and cutinase activity in culture medium.
Annual Report
2008
15
Duarte Miguel F. Prazeres and Gabriel A. Monteiro
Gene
therapy
have
h it is possible to reduce RNA without compromis-
emerged in the last two decades as promising
ing the pDNA yield and ii) pDNA is remarkably
alternatives for the treatment and prevention of
stable when stored in cell pellets (>3 weeks at
genetic disorders and acquired diseases. Non-
4ºC, >12 weeks at –20ºC) prior to processing3.
viral vectors, such as naked plasmids, constitute a
Cells are then disrupted by alkaline lysis. If con-
safer gene delivery alternative to viral vectors due
venient, lysates can be stored at –20ºC within the
to their lower toxicity and larger gene capacity.
first 8 weeks without the onset of pDNA degrada-
Plasmid DNA (pDNA) vaccines also offer a credi-
tion. Next, pDNA is concentrated by isopropanol
ble alternative for the prevention and treatment of
precipitation and protein, endotoxin and RNA con-
infectious and acquired diseases. In order for a
tent is reduced by an ammonium sulphate precipi-
DNA vaccine to be successfully developed, sev-
tation step that also acts as a conditioning step for
eral scientific and technological challenges asso-
the subsequent HIC step. HIC is carried out with a
ciated with the design and large scale production
suitable support derivatised with hydrophobic
of pDNA must be addressed. The research pro-
ligands. A typical chromatogram shows a first
gram on “Nucleic Acid Bioengineering” addresses
sharp peak of DNA vaccine followed by a broader
some of these challenges.
peak of weakly retained contaminants: RNA, ge-
and
DNA
vaccination
nomic DNA, proteins (Fig. 1). The process is roDNA vaccine manufacturing
bust, reproducible and amenable to scale-up and
A patented process based on hydrophobic inter-
delivers a product within the standard specifica-
action chromatography (HIC) has been developed
tions and with adequate biological activity (Fig. 2).
and used for the manufacturing of pDNA. E. coli
It has been extensively used in our laboratory to
cells harbouring the pDNA vaccine are cultured in
produce milligram quantities of pDNA vectors for
a bioreactor with a suitable medium. Here we
gene therapy and DNA vaccination, including pro-
found out that: i) by extending cell culture up to 26
totypes for immunization against rabies and sheep
Maedi-Visna virus.
a)
(%)(%)
Abs. 280
280nm
Absnm
NABL
Production and Design of DNA Vaccines
120
Improvement of DNA vaccine stability
DNA
vaccine
Extra- and intra-cellular nuclease degradation of
DNA vaccines after delivery and during trafficking
impurities
60
to the nucleus constitutes a barrier to gene expression and consequently to the elicitation of
immune responses. In vivo clearance of pDNA
0
0
20
40
60
Time
(min)
time
(min)
occurs within a few hours. That barrier may be
circumvented by shielding the DNA vaccines from
the nuclease-rich environments with adjuvants like
Figure 1: Purification of DNA vaccines by hydrophobic interaction chromatography
cationic lipids and other biopolymers, or by using
Research Highlights
Figure 2: CHO cells transfected with GFP expressing plasmid DNA molecules purified by the HIC process.
nuclease inhibitors. Another alternative which is
transcription. The replacement of a few (seven)
explored in our group relies on the construction of
specific nucleotides in the plasmid pMB1 origin of
pDNA variants that are more resistant to nuclease
replication could also increase (up to 2.5-fold) the
action. Although DNA secondary structures play a
nuclease resistance (Fig. 3a), while simultane-
biologically relevant role by facilitating the binding
ously augmenting (1.5 fold) the levels of the ex-
of specific proteins, our studies indicate that
pressed protein in cell culture (Fig. 3b). Moreover,
pDNA vectors can be optimized without loss of
no significant functional loss of the modified origin
functionality, leading to vectors with higher trans-
of replication was detected in E. coli.
fection efficiency. The choice of plasmid vector
sequences is important, not only for mRNA matu-
DNA vaccination and gene therapy have matured
ration/stability, but also for pDNA resistance, and
to the point where a number of products should be
should thus be taken into consideration in the de-
hitting the market in the wake of the first veteri-
sign and evaluation of pDNA vectors. For in-
nary DNA vaccines. Furthermore, new develop-
stances, in vitro and cell culture studies indicate
ments are likely to surface within the coming
that pDNA nuclease resistance can be improved 2
years because of increased investments from
-fold by changing the polyadenylation sequence.
academia and industry in the area. Our work is a
This modification, however, led to a decrease in
contribution to these efforts.
a)
b)
pVAX1GFP
M
0
10 40
pVAX1GFP-O1
0
10 40
pVAX1GFP-O2
0
10 40 (min)
sc
Transfection Efficiency (%)
50
40
30
20
10
0
pVAX1GFP
pVAX1GFP-O1 pVAX1GFP-O2
Figure 3: a) Resistance of plasmid vectors modified in the origin of replication, as measured by incubation with the singlestranded specific S1 nuclease for different times; b) transfection of CHO cells with the modified vectors using Lipofectamine.
Cells were analysed 24 hours post-transfection by flow cytometry. Error bars indicate standard deviation between four replicates.
Annual Report
2008
17
Joaquim MS Cabral, Cláudia Lobato da Silva and Margarida Diogo
Stem Cell Bioengineering Science aims to con-
However, a large number of cells is required,
tribute for a better knowledge of the ex‑vivo ex-
thus strengthening the need to develop large-
pansion of stem cells and their controlled differ-
scale systems using chemically defined media
entiation into specific cell types in bioreactor sys-
for cell expansion and/or controlled differentia-
tems. As stem cells are rare, their isolation and
tion. A stirred culture system was successfully
efficient expansion in vitro significantly increase
used to scale-up mESC expansion in serum-
the cell population available for multiple cell
containing or serum‑free media, using macropor-
therapies. The development of ex-vivo culture
ous microcarriers (Fig. 1).
conditions capable of mimicking stem cell
“niches” in vivo, by facilitating the maintenance
After 8 days, maximal cell densities achieved
and expansion of long-term transplantable stem
were 2.6 and 3.5×106 cells/mL for serum-
cells, as well as their commitment into a specific
containing and serum-free media, respectively,
cell lineage, is a major challenge in stem cell
with fold increases (relatively to day 0) of 50 and
research and its applications. Human hematopoi-
70. Importantly, mESC expanded using serum-
etic stem cells and mesenchymal stem cells, as
free medium retained their pluripotency and the
well as mouse embryonic stem cells (mESC)
ability to commit to the neural lineage (Fig. 2).
have been used as model systems at the Stem
Cell Bioengineering Laboratory.
High-throughput 3-D cell microarray
Although an effective system for the successful
Expansion of ESC
expansion and/or differentiation of stem cells was
In particular, ESC have the ability to differentiate
established, the potential therapeutic cell use is
in vitro into a wide variety of cell types with po-
contingent upon precise control of stem cell fate
tential applications in regenerative medicine.
in culture. We have recently developed in col-
4.0E+06
80
70
3.0E+06
60
Fold Increase
Viable cells/mL
SCBL
Stem Cell Bioengineering Science:
Scaling-up or Scaling-down?
2.0E+06
1.0E+06
50
40
30
20
10
0.0E+00
0
0
1
2
3
4
Time (days)
5
6
7
8
0
1
2
3
4
5
6
7
8
Time (days)
Figure 1: mESC cell expansion on macroporous microcarriers under stirred culture conditions. Growth curve in terms of viable
cell densities per milliliter (A) and cell expansion in terms of fold increase in total cell number (B), respectively, are represented
for serum-containing () and serum-free () media.
Research Highlights
Figure 2: Evaluation of pluripotency and neural commitment potential of mESC cultured on macroporous microcarriers under stirred culture conditions. Pluripotency was evaluated by alkaline phosphatase staining (A). The
percentage of neural progenitors was determined by flow cytometry (B, negative control; C, cells cultured in
neural differentiation medium).
laboration with J. Dordick, Department of Chemi-
their pluripotent and undifferentiated state. In
cal and Biological Engineering at Rensselear
addition, growth rate values obtained for different
Polytechnic Institute, a miniaturized 3-D cell-
culture systems are similar.
culture based chip for high-throughput screening
which consists of mESC encapsulated in 20 nL
Overall, we expect this work will pave an impor-
alginate gels arrayed on a functionalized glass
tant role for the successful control of stem cell
slide (Fig. 3).
fate in vitro, as well as for the design of efficient
culture systems with potential applications in
Our results show that this platform is suitable for
terms of regenerative medicine, as well as for
studying the expansion of mESC, while retaining
drug discovery in the pharmaceutical industry.
Figure 3: 3-D cell culture microarray platform
Annual Report
2008
19
Publications
Articles in International Peer-Reviewed
Journals
Cardoso, M.A.T., Geraldes, V., Cabral, J.M.S., Pa-
Azevedo, A.M., Rosa, P.A.J, Ferreira, I.F., Aires-
lavra, A.M.F., “Characterization of minocycline pow-
Barros, M.R., “Integrated process for the purification
der micronized by a supercritical antisolvent (SAS)
of antibodies combining aqueous two-phase extrac-
process”, J. Supercritical Fluids, 46, 71-76
tion, hydrophobic interaction chromatography and
size-exclusion chromatography”, J. Chromatogr. A,
Cardoso, M.A.T, Cabral, J.M.S., Palavra, A.M.F.,
1213, 154-161
Geraldes, V., “CFD analysis of supercritical antisolvent (SAS) micronization of minocycline hydrochlo-
Baptista, R.P., Pedersen, S., Cabrita, G.J., Otzen,
ride”, J. Supercritical Fluids, 47, 247-258
D.E., Cabral, J.M.S., Melo E.P., “Thermodynamics
and mechanism of cutinase stabilization by treha-
Carvalho, R.H., Lemos, M.A.N.D.A., Lemos, F.,
lose”, Biopolymers, 89, 538-547
Cabral, J.M.S., Ribeiro, F.R., “Electro-oxidation of
phenol on zeolite/graphite composite electrodes -
Brissos, V., Eggert, T., Cabral, J.M.S., Jaeger,
Part 3. Influence of the electrolyte and of nonelectro-
K.E., “Improving activity and stability of cutinase to-
active cations”, Catal. Today, 133, 855-862
wards the anionic detergent AOT by complete saturation mutagenesis”, Prot. Eng. Design Select., 21, 387-
Catarino, I., Minhalma, M., Beal, L.L., Mateus, M.,
393
de Pinho, M.N., "Assessment of saccharide fractionation by ultrafiltration and nanofiltration", J.
Brissos, V., Melo, E.P., Martinho, J.M.G. , Cabral,
Membr. Sci., 312, 34-40
J.M.S., “Biochemical and structural characterisation
of cutinase mutants in the presence of the anionic
Claudino, M.J.C., Soares, D., Marques, M.P.C., van
surfactant AOT”, Biochem. Biophys. Acta - Prot. Pro-
Keulen,
teom., 1784, 1326-1334
“Immobilization of mycobacterial cells onto silicone -
F.,
Cabral,
J.M.S.,
Fernandes,
P.,
assessing the feasibility of the immobilized biocatalyst
Cardoso, F.A., Germano, J., Ferreira, R., Cardoso,
in the production of androstenedione from sitosterol”,
S., Martins, V.C., Freitas, P.P., Piedade, M.S.,
Bioresource Technol., 99, 2304-2311
Sousa, L., “Detection of 130 nm magnetic particles
by a portable electronic platform using spin valve and
Conde, J.P., Pimentel, A.C., Pereira, A.T., Gouvêa,
magnetic tunnel junction sensors”, J. Appl. Phys.,
A., Prazeres, D.M.F., Chu, V., “Detection of molecu-
103, 07A310
lar tags with an integrated amorphous silicon
photodetector for biological applications,” J. Non-
Cardoso, M.A.T, Monteiro, G.A., Cardoso, J.P.,
Cryst. Solids, 354, 2594-2597
Prazeres, T.J.V., Figueiredo, J.M.F., Martinho,
J.M.G.,
Cabral,
J.M.S.,
Palavra,
A.M.F.,
Costa, L., Brissos, V., Lemos, F., Ribeiro, F.R.,
“Supercritical antisolvent micronization of minocycline
Cabral, J.M.S., “Comparing the effect of immobiliza-
hydrochloride”, J. Supercritical Fluids, 44, 238-244
tion methods on the activity of lipase biocatalysts in
ester hydrolysis”, Bioproc. Biosys. Eng., 31, 323-327
Costa, L., Brissos, V., Lemos, F., Ribeiro, F.R.,
Cabral, J.M.S., “Following multi-component reactions
in liquid medium using spectral band-fitting techniques”, Appl. Spectroscopy, 62, 932-935
Costa, L.F.A., Lemos, F., Ribeiro, F.R, Cabral,
J.M.S., “Zeolite screening for the racemization of 1phenylethanol”, Catal. Today, 133, 625-631
Diogo, M.M., Henrique D., Cabral, J.M.S., “Optimization and integration of expansion and neural commitment of mouse embryonic stem cells”, Biotechnol.
Appl. Biochem., 49, 105-112
Fernandes, T.G., Kwon, S.J., Lee, M.Y., Clark, D.S.,
Cabral, J.M.S., Dordick, J.S., “An on-chip, cellbased microarray immunofluorescence assay for high
-throughput analysis of target proteins”, Anal. Chem.,
Gouvêa, A., Pereira, A.T., Pimentel, A.C., Prazeres,
80, 6633-6639
D.M.F., Chu, V., Conde, J.P., “Colorimetric detection
of molecular recognition reactions with an enzyme
Ferreira, I.F., Azevedo, A.M., Rosa, P.A.J., Aires-
biolabel using a thin-film amorphous silicon photodi-
Barros, M.R., “Purification of human immunoglobulin
ode on a glass substrate”, Sensors Actuators B:
G by thermoseparating aqueous two-phase systems”,
Chemical, 135, 102-107
J. Chromatogr. A, 1195, 94-100
Lienqueo, M.E., Salazar, O., Calado, C.R.C.,
Frias, A.M., Porada, C.D., Crapnell, K.B., Cabral,
Fonseca, L.P., Cabral, J.M.S., “Influence of trypto-
J.M.S., Zanjani, E.D., Almeida-Porada, G., “Gene-
phan tags on the purification of cutinase, secreted by
ration of functional natural killer and dendritic cells in
a recombinant Saccharomyces cerevisiae, using cati-
a human stromal-based serum-free culture system
onic expanded bed adsorption and hydrophobic inter-
designed for cord blood expansion”, Exp. Hematol.,
action chromatography”, Biotechnol. Lett., 30, 1353-
36, 61-68
1358
Gonçalves, D., Prazeres, D.M.F., Chu, V., Conde,
Madeira, C., Loura, L.M.S., Prieto, M., Fedorov, A.,
J.P., “Detection of DNA and proteins using amor-
Aires-Barros, M.R., "Effect of ionic strength and
phous silicon ion-sensitive thin-film field effect transis-
presence of serum on lipoplexes structure monitor-
tors”, Biosens. Bioelectron., 24, 545-551
ized by FRET", BMC Biotechnol., 8: 20
Gonçalves, D., Prazeres, D.M.F., Chu, V., Conde,
Marques, M.P.C., Cabral, J.M.S., Fernandes, P.,
J.P., “Amorphous silicon thin-film transistors gated
“Online oxygen monitoring system - non-conventional
through an electrolyte solution”, IEEE Electron Device
steroid fermentations in microtiter plates”, Bioforum
Lett., 29, 1030-1033
Europe, 9, 42-43
Annual Report
2008
21
Pimentel, A.C, Prazeres, D.M.F., Chu, V., Conde,
J.P., “Fluorescence detection of DNA using an amorphous silicon p-i-n photodiode”, J. Applied Physics.,
104, 054913
Prazeres, D.M.F., “Prediction of diffusion coefficients
of plasmids”, Biotechnol. Bioeng., 99, 1040-1044
Ribeiro, S.C., Oliveira, P.H., Prazeres, D.M.F.,
Monteiro, G.A., “High frequency plasmid recombination mediated by 28 bp direct repeats”, Mol. Biotechnol., 40, 252-260
Sampaio, P.N., Fortes, A.M., Cabral, J.M.S., Pais,
Martins, S., Prazeres, D.M.F., Monteiro, G.A.,
M.S., Fonseca, L.P., “Production and characteriza-
“Chemiluminescent bead-based hybridization assay
tion of recombinant cyprosin B in Saccharomyces
for the detection of genomic DNA from E. coli in puri-
cerevisiae (W303-1A) strain”, J. Biosci. Bioeng., 105,
fied plasmid samples”, Anal. Bioanal. Chem., 391,
305-312
2179-2187
Santos, A.M., Fedorov, A., Martinho, J.M.G., BapOliveira, P.H., Lemos, F., Monteiro, G.A., Prazeres,
tista, R.P., Taipa, M.A., Cabral, J.M.S., “Orientation
D.M.F., “Recombination frequency in plasmid DNA
of cutinase adsorbed onto PMMA nanoparticles
containing direct repeats - predictive correlation with
probed by tryptophan fluorescence”, J. Phys. Chem.
repeat and intervening sequence length”, Plasmid,
B, 112, 3581-3585
60, 159-165
Sousa, F., Prazeres, D.M.F., Queiroz, J.A., “Affinity
chromatography approaches for overcoming the challenges of purifying plasmid DNA”, Trends Biotechnol.,
26, 518-525
Sousa, F., Prazeres, D.M.F., Queiroz, J.A., “Specific
recognition of supercoiled plasmid DNA by arginine
affinity chromatography”, Anal. Biochem., 374, 432434
Taipa, M.A., “Immunoassays: Biological tools for high
throughput screening and characterisation of combinatorial libraries”, CCHTS, 11, 325-335
Teles, F.R.R., Fonseca, L.P., “Trends in DNA bio-
Articles in Conference Proceedings
sensors”, Talanta, 77, 606–623
Barros, D.P.C., Bernardino, S.M.S.A., Fernandes,
P., Cabral, J.M.S., Fonseca, L.P., “Studies of fed-
Teles, F.R.R., Fonseca, L.P., “Applications of poly-
batch operation mode on synthesis of short chain
mers for biomolecule immobilization in electrochemi-
ethyl esters catalyzed by cutinase”, Proceedings of
cal biosensors”, Mat. Sci. Eng. C, 28, 1530–1543
the 10th International Chemical and Biological Engineering Conference - CHEMPOR 2008, E.C. Ferreira
Vale, G., Mota, A., Fonseca, L., Capelo, J.L.,
and M. Mota (eds.), 4-6 September 2008, Braga, Por-
“Ultrasonic assisted enzymatic digestion -USAED- for
tugal, pp 471-476
total elemental determination and elemental speciation: a tutorial”, Talanta, 75, 872-884
Bernardino, S.M.S.A., Gallegos, J.F.M., Maduro,
F., Fernandes, P., Cabral, J.M.S., Fonseca, L.P.,
Vale, G., Pereira, S., Mota, A., Fonseca, L., Capelo,
“Nano and micro-biocatalysts manufacture and their
J.L., “Enzymatic probe sonication as a tool for solid-
impact on the synthesis of β-lactamic antibiotics”,
liquid extraction for total selenium determination by
Proceedings of the 10th International Chemical and
electrothermal-atomic absorption spectrometry”, Ta-
Biological Engineering Conference - CHEMPOR
lanta, 74,198-205
2008, E.C. Ferreira and M. Mota (eds.), 4-6 September 2008, Braga, Portugal, pp 489-494
Vidinha, P., Augusto, V., Nunes, J., Lima, J.C.,
Cabral, J.M.S., Barreiros S., “Probing the microenvi-
Costa, L., Brissos, V., Lemos, F., Ribeiro, F.R.,
ronment of sol-gel entrapped cutinase: The role of
Cabral, J.M.S., “Monitoring multi-component liquid
added zeolite NaY”, J. Biotechnol., 135, 181-189
reaction systems containing highly dispersible heterogeneous catalysts using in situ diode array spectro-
Vidinha, P., Barreiros, S., Cabral, J.M.S., Nunes,
photometry and band-fitting techniques”, Proceedings
T.G., Fidalgo, A., Ilharco, L.M., “Enhanced biocata-
of the 10th International Chemical and Biological Engi-
lytic activity of ORMOSIL-encapsulated cutinase: The
neering Conference - CHEMPOR 2008, E.C. Ferreira
matrix structural perspective”, J. Phys. Chem. C, 112,
and M. Mota (eds.), 4-6 September 2008, Braga, Por-
2008-2015
tugal, pp 213-218
Vidinha, P., Lourenço, N.M.T., Pinheiro, C., Brás,
A.R., Carvalho, T., Silva, T.S., Mukhopadhyay, A.,
Romão, M.J., Parola, J., Dionísio, M., Cabral
J.M.S., Afonso, C.A.M., Barreiros, S., “Ion Jelly: a
tailor-made conducting material for smart electrochemical devices”, Chem. Commun, 44, 5842–5844
Vojinović, V., Cabral, J.M.S., Fonseca, L.P., “Exsitu bioprocess monitoring techniques”, Chemical
Industry & Chemical Engineering Quarterly, 13, 1-15
Annual Report
2008
23
Pereira, A.T., Pimentel, A., Loureiro, J., Freitas,
P.P., Chu, V., Prazeres, D.M.F., Conde, J.P.,
"Colorimetric detection of antigen-antibody recognition in a microfluidic channel with an integrated photodiode", Proceedings of the XXII EUROSENSORS, 710 September, Dresden, Germany, Verein Deutscher
Ingenieure, pp 1212-1215
Patents
Ribeiro,
I.,
Afonso,
C.A.M.,
Cabral,
J.M.S.,
Lourenço, N.M.T., Vidinha, P., Barreiros, S.,
“Synthesis and application of a family of new materials resulting from the chemical cross-linking between
gelatine and organic salts”, European (EP2006321),
United States (US2008/0319164), South Korea (102008-0058173) and Japan (2008-160947) Patent
Applications.
Book chapters
Fernandes,
P.,
Cattorini,
S.,
Carvalho,
F.,
Fernandes, P., Cabral, J.M.S., "Biocatalysis in Bi-
Marques, M.P.C., Bernardino, S., Maduro, F.,
phasic Systems: General", in: Organic synthesis with
Badenes, S., Barros, D., de Carvalho, C.C.C.R.,
enzymes in non-aqueous media, G. Carrea, S. Riva
Fonseca, L.P., Cabral, J.M.S., “A multipurpose hy-
(eds.), Wiley-VCH, Weinheim, pp. 191-209
drogel system for biocatalyst immobilization”, Proceedings of the 10th International Chemical and Bio-
Fernandes, P., Cabral, J.M.S., "Immobilization –
logical Engineering Conference - CHEMPOR 2008,
Microencapsulation", in: Advances in Fermentation
E.C. Ferreira and M. Mota (eds.), 4-6 September
Technology, A. Pandey, C. Larroche, C.R. Soccol, C.-
2008, Braga, Portugal, pp 1935-1940
G. Dussap (eds.), Asiatech Publishers, Inc., New
Delhi, pp. 45-84
Fernandes,
P.,
Cattorini,
S.,
Cabral,
J.M.S.,
“Enzymatic inulin hydrolysis using PVA-based matrices”, Proceedings of the 10th International Chemical
Ph.D. Thesis
and Biological Engineering Conference - CHEMPOR
Dina Isabel Viegas Gonçalves, “Label-free detection
2008, E.C. Ferreira and M. Mota (eds.), 4-6 Septem-
of biomolecules using amorphous silicon ion-sensitive
ber 2008, Braga, Portugal, pp 1873-1878
field-effect transistors”, PhD Thesis, Technical University of Lisbon, IST, Lisbon (advisors: João P.E.R.
Conde and D. Miguel F.T. Prazeres; IST, Lisbon)
Luisella Ruiu, “De novo design, synthesis and
screening of combinatorial libraries of affinity ligands
directed towards the surface of cutinase from Fusarium solani pisi”, PhD Thesis, Technical University of
Lisbon, IST, Lisbon (advisor: M. Ângela C.G. Taipa;
IST, Lisbon)
Fani Pereira Sousa, “Affinity chromatography processes in nucleic acids”, PhD Thesis, Universidade da
Beira Interior, Covilhã (advisors: João A.S.R. Queiroz, UBI, Covilhã and D. Miguel F.T. Prazeres; IST,
Lisbon)
Sofia de Medina Aires Martins, “Development of
quantitative micro-plate-based assays for DNA hybridization”, PhD Thesis, Technical University of Lisbon, IST, Lisbon (advisors: Gabriel A.A. Monteiro and
Luís J.P. Fonseca; IST, Lisbon)
M.Sc. Thesis
Ana Mafalda Nunes Rodrigues, “Application of elec-
Mauro José Castanho Claudino, “Use of miniature
tric field assisted hydridization to peptide nucleic ac-
reactors for the characterization of the side-chain
ids”, MSc Thesis, Technical University of Lisbon, IST,
cleavage of β-sitosterol using immobilized cells”, MSc
Lisbon (advisors: João P.E.R. Conde and D. Miguel
Thesis, Technical University of Lisbon, IST, Lisbon
F.T. Prazeres; IST, Lisbon)
(advisors: Pedro Fernandes, IST and Joaquim M.S.
Cabral; IST, Lisbon)
Joana Brissos Magalhães Lima, “Efeito do grau de
superenrolamento de plasmídeos na sua estabilidade
Milene da Silva Santos, “Cell micropatterning for cell
estrutural e função biológica”, MSc Thesis, Faculdade
chips applications”, MSc Thesis, Technical University
de
of Lisbon, IST, Lisbon (advisors: João P.E.R. Conde
Ciências,
Universidade
de
Lisboa,
Lisbon
(advisors: D. Miguel F.T. Prazeres, IST, Lisbon and
and D. Miguel F.T. Prazeres, IST, Lisbon)
Maria do Céu Correia, FCUL, Lisbon)
Salomé Alexandra de Sá Magalhães, “Construction,
Joana Filipa Sobrinho Boura, “Ex-vivo expansion of
optimization and testing of African Tripanosomiasis
human myoblasts and fibroblasts for potential use in
DNA vaccine prototypes with improved nuclease re-
urinary
Thesis,
sistance”, MSc Thesis, Faculdade de Ciências, Uni-
Faculdade de Ciências, Universidade de Lisboa, Lis-
versidade de Lisboa, Lisbon (advisors: D. Miguel F.
bon (advisors: Cláudia A.M. Lobato da Silva, IST,
T. Prazeres, IST, Lisbon and Maria do Céu Correia,
Lisbon and Maria Gabriela G.M. Rodrigues, FCUL,
FCUL, Lisbon)
incontinence
treatment”,
MSc
Lisbon)
Annual Report
2008
25
Oral Communications
International Conferences
Barros, D.P.C., Bernardino, S.M.S.A., Fernandes,
Afonso, C.A.M., Lourenço, N.M.T., Monteiro, C.A.,
P., Cabral, J.M.S., Fonseca, L.P., “Studies on fed-
“Efficient ionic acylating agents for enzymatic resolu-
batch operation mode on biosynthesis of short chain
th
tion of alcohols in ionic liquids”, 236 National Meet-
ethyl esters catalyzed by cutinase”, 10th International
ing and Exposition of the American Chemical Society,
Chemical and Biological Engineering Conference -
Philadelphia, USA, August
CHEMPOR 2008, Braga, Portugal, September
Azevedo, A.M., Rosa, P.A.J, Ferreira, I.F., de Vries,
Barros, D.P.C., Fonseca, L.P., Cabral, J.M.S.,
J., Korporaal, R., Verhoef, H.J., Visser, T.J., Aires-
Weiss, C., Landfester, K., “Enzymatic synthesis of
Barros, M.R., “Affinity-enhanced partitioning of hu-
fatty acid alkyl esters in miniemulsion”, 7th European
man antibodies in aqueous two-phase systems”, 7th
Symposium on Biochemical Engineering – ESBES 7,
European Symposium on Biochemical Engineering
Faro, Portugal, September
Science – ESBES7, Faro, Portugal, September
Bernardino, S.M.S.A., Fernandes, P., Fonseca,
Azevedo, A.M., Rosa, P.A.J., Ferreira, I.F., Aires-
L.P., “Cephalexin synthesis by Penicillin G Acylase
Barros, M.R., “An alternative process for the purifica-
immobilized in sol-gel”, 7th European Symposium on
tion of therapeutic antibodies comprising aqueous two
Biochemical Engineering – ESBES 7, Faro, Portugal,
th
-phase extraction”, 28 International Symposium on
September
the Separation of Proteins, Peptides and Polynucleotides – ISPPP2008, Baden-Baden, Germany, Sep-
Bernardino, S.M.S.A., Gallegos, J.F.M., Maduro,
tember
F., Fernandes, P., Cabral, J.M.S., Fonseca, L.P.,
“Nano and micro-biocatalysts manufacture and their
impact on the synthesis of β-lactamic antibiotics”, 10th
International Chemical and Biological Engineering
Conference – CHEMPOR 2008, Braga, Portugal,
September
Fernandes, A.M., Diogo M.M., Lobato da Silva, C.,
Henrique, D., Cabral, J.M.S., “Mouse embryonic
stem cell expansion in a microcarrier-based stirred
culture system”, Tissue Engineering and Regenerative Medicine International Society - 2008 Annual
TERMIS, Porto, Portugal, June
Fernandes, A.M., Diogo M.M., Lobato da Silva, C.,
Henrique D., Cabral, J.M.S., “Mouse embryonic
stem cell expansion in a microcarrier-based stirred
culture system”, 10th International Chemical and Biological Engineering Conference – CHEMPOR 2008,
Braga, Portugal, September
Fernandes, T.G., Kwon, S.J., Lee, M., Diogo, M.M.,
Lobato da Silva, C., dos Santos, F., Andrade, P.Z.,
Lobato da Silva, C., Clark, D. S., Cabral, J.M.S.,
Gonçalves, R., Almeida-Porada, G., Cabral, J.M.S.,
Dordick, J.S., " High-throughput 3D cell microarray to
“Maximization of the ex-vivo expansion of human
study stem cell fate", SBE's First International
hematopoietic stem/progenitor cells by direct contact
Conference on Stem Cell Engineering, San Diego,
culture with mesenchymal stem cells", SBE's First
USA, January
International Conference on Stem Cell Engineering,
San Diego, USA, January
Fernandes, T.G., Kwon, S.J., Lee, M.Y., Diogo,
M.M., Lobato da Silva, C., Clark, D.S., Cabral,
Madeira, C., Ferreira, J.A.B., Andrade, S., Cabrita,
J.M.S., Dordick, J.S., "A high-throughput 3D cell
G.J.M., Costa, S.M.B., Melo, E.P., “Fluorescence
th
microarray to study stem cell fate", 7
European
studies of the pair CFP-YFP: FLIM-FRET in vitro
Symposium on Biochemical Engineering – ESBES 7,
studies and expression in neuronal cells to probe the
Faro, Portugal, September
prion protein at the cell surface”, Focus on Microscopy, Osaka-Awaji, Japan, April
Fernandes, T.G., Kwon, S.J., Lee, M.Y., Diogo,
M.M., Lobato da Silva, C., Clark, D.S., Cabral,
Martins, V.C., Cardoso, F.A., Fonseca, L.P.,
J.M.S., Dordick, J.S., "Exploring stem cell fate using
Freitas, P.P., “Femptomolar sensitivity for magneti-
th
three-dimensional cellular microarrays", 7 European
cally assisted DNA hybridisation”, 1st International
Symposium on Biochemical Engineering Science –
Conference from Nanoparticles and Nanomaterials to
ESBES 7, Malcolm Lilly Award Lecture, Faro,
Nanodevices
Portugal, September
Halkidiki, Greece, June
Fernandes, P., “Whole cell biocatalysis”, III Congreso
Martins, V.C., Cardoso, F.A., Loureiro, J., Ger-
Interuniversitario de Biotecnología, Léon, Spain, July
mano, J., Cardoso, S., Ferreira, R., Fonseca, L.P.,
and
Nanosystems
-
IC4N
2008,
Sousa, L., Piedade, M.S., Freitas, P.P., “Integrated
Fonseca, L.P., Fernandes, P., Lourenço, N.,
spintronic platforms for biomolecular detection, sepa-
Cordas, C., Bernadino, S., Barros, D., Marques,
ration and counting”, Joint European Magnetic Sym-
M., “Recent trends in enzyme and cell immobilization
posia - JEMS08, Dublin, Ireland, September
by entrapment and encapsulation”, XVI International
Conference on Bioencapsulation, Dublin, Irland,
September
Gomes, A.G., Azevedo, A.M., Aires-Barros, M.R.,
Prazeres, D.M.F., “Plasmid DNA purification by
phenyl boronate affinity chromatography”, 7th European Symposium on Biochemical Engineering Science - ESBES7, Faro, Portugal, September
Annual Report
2008
27
Monteiro, G.A., “Bringing DNA vaccines closer to the
Rosa, P.A.J, Azevedo, A.M., Ferreira, I.F., Aires-
bedside”, Vaccines Europe, Brussels, Belgium, De-
Barros, M.R., “Purification of human antibodies using
cember
affinity aqueous-two phase systems”, 10th International Chemical and Biological Engineering Confer-
Prazeres, D.M.F., Pereira, A.T., Pimentel, A.C.,
ence - CHEMPOR 2008, Braga, Portugal, September
Gouvêa, A., Chu, V., Conde, J.P., “Integrated optoelectronic detection of molecular recognition reacth
Rosa, P.A.J., Azevedo, A.M., Ferreira, I.F., Som-
tions”, 7 European Symposium on Biochemical Engi-
merfeld, S., Aires-Barros, M.R., Bäcker, W., “Multi-
neering Science - ESBES7, Faro, Portugal, Septem-
stage aqueous-two phase extraction of human anti-
ber
bodies”, Jahrestreffen des ProcessNet – Fachausschusses Extraktion und des Arbeitskreises Phytoex-
Rosa, P.A.J., Azevedo, A.M., de Vries, J., Korpo-
trakte, Clausthal-Zellerfeld, Germany, April
raal, R., Verhoef, H.J., Visser, T.J., Aires-Barros,
M.R., “Optimisation of affinity-enhanced purification of
Samatou, J. A., Wentink, E.A., Hoffmann, A., Rosa,
th
antibodies using aqueous two-phase extraction”, 12
P.A.J., Azevedo, A.M., Aires-Barros, M.R., Bäcker,
International Symposium on Preparative and Indus-
W., Górak, A., “Modellierung und simulation der
trial Chromatography and Allied Techniques - SPICA
mehrstufigen
2008, Zurich, Switzerland, September - October
antikörpern
extraktion
in
Jahrestreffen
wässrigen
der
und
von
monoklonalen
zweiphasensystemen”,
Fachgemeinschaft
Rosa, P.A.J, Azevedo, A.M., Ferreira, I.F., Som-
Apparate-
Anlagentechnik,
merfeld, S., Aires-Barros, M.R., Bäcker, W., “Multi-
Germany, November
Bad
Prozess-,
Honnef,
stage-enhanced recovery of human antibodies by
aqueous two-phase extraction”, 7th European Sympo-
Sousa,
sium on Biochemical Engineering Science - ESBES
“Temperature-induced conformational changes of
7, Faro, Portugal, September
pDNA and their influence on histidine-agarose reten-
F.,
Prazeres,
D.M.F.,
Queiroz,
J.A.,
tion: A circular dichroism study”, 7th European Symposium on Biochemical Engineering Science - ESBES7,
Faro, Portugal, September
Vidinha, P., Lourenço, N.M.T., Carvalho, T., Brás,
A.R., Silva, T.S., Mukhopadhyay, A., Cordas, C.M.,
Romão, M.J., Dionisio, M., Cabral, J.M.S., Fonseca, L.P., Afonso, C.A.M., Barreiros, S., "Ion jelly®:
A tailor-made conducting material for smart electrochemical devices", 7th International Symposium on
Polyelectrolytes - Polyelectrolytes 2008, Coimbra,
Portugal, June
National Conferences
Andrade,
P.Z., Temtem, M., dos Santos, F.,
Lobato da Silva, C., Aguiar-Ricardo, A., Cabral,
J.M.S., “"Green" chitosan membranes for the ex-vivo
expansion of human bone marrow mesenchymal
stem cells”, 3rd Annual International Meeting of the
Portuguese Society for Stem Cells and Cellular
Therapies, Faro, Portugal, April
Diogo, M.M., Fernandes, A.M., Fernandes, T.G.,
Lobato da Silva, C., Henrique, D., Dordick, J.S.,
Cabral,
J.M.S.,
"Large-scale
and
nano-scale
approaches for the expansion of mouse embryonic
stem (mES) cells", 3rd Annual International Meeting
of the Portuguese Society for Stem Cells and Cellular Therapies, Faro, Portugal, April
dos Santos, F., Lobato da Silva, C., Andrade,
P.Z., Miranda, N., Teixeira, G., Guimarães, A., Fer-
Fernandes, P., “Biocatálise industrial”, Workshop IV
reira, I., Rodriguez, E., Oiveira, J., Trindade, H.,
Dia de Biologia Marinha e Biotecnologia, Peniche,
Abecassis, M., Cabral, J.M.S., “Treatment of steroid
Portugal, May
and extracorporeal resistant acute graft-versus-host
disease with donor mesenchymal stem cells”, 3rd
Fernandes, P., Marques, M.P.C., Cattorini, S., Car-
Annual International Meeting of the Portuguese Soci-
valho, F., Cabral, J.M.S., “A dual purpose immobili-
ety for Stem Cells and Cellular Therapies, Faro, Por-
zed biocatalyst for inulin and sucrose hydrolysis”,
tugal, April
Carbohydrates as Organic Raw Materials V - CORM
V, Lisbon, Portugal, February
Eibes, G., Fernandes, A.M., Diogo, M.M., Lobato
da Silva, C., Cabral, J.M.S., “Modeling of mouse
Lourenço,
embryonic stem cell expansion in a stirred culture
Carvalho, T., Silva, T.S., Mukhopadhyay, A.,
rd
N.M.T.,
Vidinha,
P.,
Brás,
A.R.,
system”, 3 Annual International Meeting of the Por-
Cordas, C., Dionisio, M., Romão, M.J., Cabral,
tuguese Society for Stem Cells and Cellular Thera-
J.M.S., Fonseca, L.P., Afonso, C.A.M., Barreiros,
pies, Faro, Portugal, April
S.,
"Ion
Jelly®-
A
tailor-made
Material
for
st
Electrochemical Applications", 1 Portuguese Young
Estrela, N.L., Chen, L.Y., Gunna, S.M.C., Cabrita,
Chemists Meeting, Lisbon, Portugal, October
G.J.M., Otzen, D.E., Melo, E.P., “Folding and amyloidosis of proteins: The prevention of amyloidosis by
Marques, M.P.C., Caramujo, M.J., de Carvalho,
osmolytes”, XVIth National Congress of Biochemis-
C.C.C.R., "Bioremediação de amostras da Base
try, São Miguel, Portugal, October
Naval de Lisboa", Jornadas do Mar 2008, Escola
Naval, Lisbon, Portugal, November
Annual Report
2008
29
Poster Presentations
International Conferences
Bernardino, S.M.S.A., Fernandes, P., Fonseca,
Azevedo, A.M., Rosa, P.A.J., Ferreira, I.F., Aires-
L.P., “Preparation of micro-porous silica xerogel with
Barros, M.R., “Aqueous two-phase extraction of hu-
magnetic properties and its application to penicillin G
man antibodies”, Bioprocess Technology Europe:
acylase immobilization”, XVI International Conference
Development and Production of Antibodies, Vaccines
on Bioencapsulation, Dublin, Ireland, September
and Gene Vectors, Amsterdam, The Netherlands,
June-July
Botelho-Cunha, V., Mateus, M., Petrus, J.C.C., de
Pinho, M.N., “Membrane fractionation of galacto-
Badenes, S.M., Lemos, F., Cabral, J.M.S., “Cata-
oligosaccharides produced enzymatically in lactose
lysed transesterification of triolein using microencap-
solutions”, Engineering with Membranes - Membrane
sulated cutinase in AOT-reversed micelles for bio-
Processes: Development, Monitoring and Modelling;
th
diesel production”, 4 International Congress on Bio-
From the nano to the macroscale - EWM2008, Vale
catalysis - BIOCAT 2008, Hamburg, Germany, Au-
de Lobo, Portugal, May
gust-September
Cabeça, R., Prazeres, D.M.F., Chu, V., Conde, J.P.,
Badenes, S.M., Lemos, F., Cabral, J.M.S., “Opti-
“Electrical and Chemical Control of Surfaces for DNA
mization of biodiesel production by triolein transesteri-
Immobilization and Hybridization”, 2008 Materials
fication using microencapsulated cutinase in AOT-
Research Society Spring Meeting, São Francisco,
th
reversed micelles”, 7 European Symposium on Bio-
USA, March
chemical Engineering Science – ESBES 7, Faro, Portugal, September
Carapuça, E., Azzoni, A., Prazeres, D.M.F., Monteiro, G.A., Mergulhão, F.J.M., “In vivo plasmid sta-
Barros, D.P.C., Bernardino, S.M.S.A., Fernandes,
bility: production host versus target cell. Implications
P., Fonseca, L.P., Cabral, J.M.S., “Comparison of
on DNA vaccine development and protein produc-
cutinase bioencapsulation in sol-gel and PVA versus
tion”, 5th Recombinant Protein Production Meeting: An
lyophilized form on biosynthesis of ethyl caproate in
integrate view of host physiology, Sardinia, Italy, Sep-
organic solvent”, XVI International Conference on
tember
Bioencapsulation, Dublin, Ireland, September
Carvalho, J.A., Monteiro, G.A., Atouguia, J., PraBarros, D.P.C., Fonseca, L.P., Cabral, J.M.S.,
zeres, D.M.F., Rodgers, J., “Developing a vaccine
Weiss, C., Landfester, K., “Mini-emulsion and or-
for African trypanosomiasis: only wishful thinking or a
ganic solvent media in biosynthesis of flavours esters
definite possibility?”, Infectious diseases of the nerv-
th
- studies on stepwise addition of substrates”, 4 Inter-
ous system: pathogenesis and worldwide impact,
national Congress on Biocatalysis – BIOCAT 2008,
Paris, France, September
Hamburg, Germany, August-September
Carvalho, J.A., Rodgers, J., Atouguia, J., Prazeres, D.M.F., Monteiro, G.A, “Screening of DNA
vaccines prototypes encoding antigen targeting sequences against African Trypanosomiasis”, Vaccine
Technology, Albufeira, Portugal, June
Cordas, C.M., Lourenço, N.M.T., Vidinha, P.,
Afonso, C.A.M., Barreiros, S., Cabral, J.M.S.,
Fonseca, L.P., “Immobilization of proteins in newionic-conducting-based materials - Ion Jelly®”, XVI
International Conference on Bioencapsulation, Dublin, Ireland, September
Fernandes, P., Marques, M.P.C., Carvalho, F.,
Cabral, J.M.S., “A simple method for biocatalyst im-
Cordas, C.M., Lourenço, N.M.T., Vidinha, P.,
mobilization using PVA based hydrogel particles”, 7th
Afonso, C.A.M., Barreiros, S., Fonseca, L.P.,
European Symposium on Biochemical Engineering
Cabral, J.M.S., "Electrochemical detection of immobi-
Science - ESBES 7, Faro, Portugal, September
th
lized proteins in ion jelly films", 7 International Symposium on Polyelectrolytes - Polyelectrolytes 2008,
Fernandes,
Coimbra, Portugal, June
“Enzymatic
P.,
Cattorini,
inulin
S.,
hydrolysis
Cabral,
using
J.M.S.,
PVA-based
th
matrices”, 10 International Chemical and Biological
de Carvalho, C.C.C.R., Marques, M.P.C., "Bioremeth
diation of samples from a naval base", 7 European
Engineering Conference - CHEMPOR 2008, Braga,
Portugal, September
Symposium on Biochemical Engineering Science ESBES 7, Faro, Portugal, September
Fernandes, A.M., Diogo, M.M., Lobato da Silva, C.,
Henrique D., Cabral, J.M.S., “Mouse embryonic
dos Santos, F., Lobato da Silva, C., Andrade, P.Z.,
stem cell expansion in a microcarrier-based stirred
Abecasis, M., Cabral, J.M.S, “Effect of hypoxia on
culture system”, SBE's First International Conference
human mesenchymal stem cell (MSC) expansion and
on Stem Cell Engineering, San Diego, USA, January
th
metabolism”, 7 European Symposium on Biochemical Engineering Science – ESBES 7, Faro, Portugal,
Fernandes, T.G., Kwon, S.J., Lee, M.Y., Clark,
September
D.S., Cabral, J.M.S., Dordick, J.S., “An on-chip, cellbased microarray immunofluorescence assay for high
dos Santos, F., Lobato da Silva, C., Andrade, P.Z.,
-throughput analysis of target proteins” 7th European
Abecasis, M., Cabral, J.M.S, “Effect of hypoxia on
Symposium on Biochemical Engineering Science –
human mesenchymal stem cell (MSC) expansion and
ESBES 7, Faro, Portugal, September
metabolism”, 36
th
International Annual Scientific
Meeting, ISEH Society for Hematology and Stem
Fernandes, T.G., Kwon, S.J., Lee, M.Y., Diogo,
Cells, Boston, USA, July
M.M., Lobato da Silva, C., Clark, D.S., Cabral,
J.M.S., Dordick, J.S., "Expansion and neural com-
Fernandes,
P.,
Cattorini,
S.,
Carvalho,
F.,
mitment of mouse embryonic stem cells on a microar-
Marques, M.P.C., Bernardino, S., Maduro, F.,
ray platform", SBE's First International Conference on
Badenes, S., Barros, D., Carvalho, C.C.C.R.,
Stem Cell Engineering, San Diego, USA, January
Fonseca, L.P., Cabral, J.M.S., “A multipurpose
hydrogel system for biocatalyst immobilization”, 10th
Fernandes, T.G., Kwon, S.J., Lee, M.Y., Diogo,
International Chemical and Biological Engineering
M.M., Lobato da Silva, C., Clark, D.S., Cabral,
Conference - CHEMPOR 2008, Braga, Portugal, Sep-
J.M.S., Dordick, J.S., "A high-throughput 3D cell
tember
microarray to study stem cell fate", 6th ISSCR Annual
Meeting, Philadelphia, USA, June
Annual Report
2008
31
Ferreira, I.F., Azevedo, A.M., Rosa, P.A.J, Aires-
Freitas, S.S., Wu, M., Monteiro, G.A., Prazeres,
Barros, M.R., “Temperature induced affinity aqueous
D.M.F., Santos, J.A.L., “Intermediate Recovery of
two-phase extraction for the purification of human
Plasmid DNA Using Tangential Flow Filtration”, Engi-
th
immunoglobulin G”, 7 European Symposium on Bio-
neering with Membranes 2008, Vale do Lobo, Portu-
chemical Engineering Science – ESBES 7, Faro, Por-
gal, May
tugal, September
Gomes, A.G., Azevedo, A.M., Aires-Barros, M.R.,
Ferreira, I.F., Azevedo, A.M., Rosa, P.A.J, Aires-
Prazeres, D.M.F., “Plasmid DNA purification by
Barros, M.R., “Optimisation of temperature induced
phenyl boronate affinity chromatography”, 28th Inter-
aqueous-two phase extraction of human antibodies”,
national Symposium on the Separation of Proteins,
th
28
International Symposium on the Separation of
Proteins, Peptides and Polynucleotides – ISPPP
Peptides and Polynucleotides – ISPPP2008, BadenBaden, Germany, September
2008, Baden-Baden, Germany, September
Lobato da Silva, C., dos Santos, F., Andrade, P.Z.,
Ferreira, I.F., Rosa, P.A.J, Azevedo, A.M., Aires-
Miranda,
Barros, M.R., “Optimisation of temperature induced
Rodriguez, E., Oliveira, J., Trindade, H., Cabral,
aqueous-two phase extraction of human antibodies”,
J.M.S, Abecasis, M., “Treatment of steroid and
th
12
International Symposium on Preparative and
N.,
extracorporeal
Teixeira,
resistant
G.,
Guimarães,
acute
I.,
graft-versus-host
Industrial Chromatography and Allied Techniques -
disease with donor mesenchymal stem cells”, 36th
SPICA
International Annual Scientific Meeting, ISEH Society
2008,
Zurich,
Switzerland,
September-
October
for Hematology and Stem Cells, Boston, USA, July
Fonseca, L.P., Barros, D.P.C., “Biosynthesis of ethyl
Lobato da Silva, C., dos Santos, F., Andrade, P.Z.,
caproate and other short alkyl esters catalyzed by
Miranda, N., Teixeira, G., Guimarães, I., Rodri-
cutinase”, COST 865 - Spring 2008 Workshop, Bioen-
guez, E., Oliveira, J., Trindade, H., Cabral, J.M.S,
capsulation Sciences to Applications, Ljubljana, Slo-
Abecasis,
venia, April
extracorporeal
M.,
“Treatment
resistant
acute
of
steroid
and
graft-versus-host
disease with donor mesenchymal stem cells”, 7th
Freitas, S.S., Monteiro, G.A., Prazeres, D.M.F.,
European Symposium on Biochemical Engineering
Santos, J.A.L., “Recovery of Plasmid DNA from Pre-
Science – ESBES 7, Faro, Portugal, September
treated Lysates Using Tangential Flow Filtration”, 7
th
European Symposium on Biochemical Engineering
Lourenço, N.M.T., Vidinha, P., Cordas, C.M.,
Science – ESBES 7, Faro, Portugal, September
Afonso, C.A.M., Barreiros, S., Cabral, J.M.S.,
Fonseca, L.P., “Ion Jelly® - A suitable material for
Freitas, S.S., Monteiro, G.A., Prazeres, D.M.F.,
biosensing detection", 7th International Symposium on
Santos, J.A.L., “Purification of plasmid DNA vectors
Polyelectrolytes – Polyelectrolytes 2008, Coimbra,
by hydrophobic interaction chromatography using
Portugal, June
th
sodium citrate in the mobile phase”, 28 International
Symposium on the Separation of Proteins, Peptides
Marques, M.P.C., Cabral, J.M.S., Fernandes, P., “A
and Polynucleotides – ISPPP 2008, Baden-Baden,
novel online-monitoring system for oxygen in non-
Germany, September
conventional fermentations”, 7th European Symposium on Biochemical Engineering Science – ESBES
7, Faro, Portugal, September
Marques, M.P.C., Carvalho, F., Claudino, M.J.C.,
Prazeres, D.M.F., Azzoni, A.R, Freitas, S.S., San-
Fernandes, P., Cabral, J.M.S., “Multi-step biotrans-
tos, J.A.L., Monteiro, G.A., “On the design and pro-
th
formations throughout scales”, 7 European Sympo-
duction of more stable and efficient plasmid DNA
sium on Biochemical Engineering Science - ESBES
vectors”, Vaccine Technology II, Albufeira, Portugal,
7, Faro, Portugal, September
June
Martins,
V.C.,
Cardoso,
F.A.,
Cardoso,
S.,
Prazeres, D.M.F., “Prediction of Diffusion Coefficients
Fonseca, L.P., Freitas, P.P., “Biological detection
of Plasmids”, 28th International Symposium on the
limit of a SV-based biochip for pathogenic analysis”,
Separation of Proteins, Peptides and Polynucleotides
Nanoiberian Conference – Nanospain 2008, Braga,
– ISPPP 2008, Baden-Baden, Germany, September
Portugal, April
Prazeres, D.M.F., Santos, J.A.L., Freitas, S.S., “A
Monteiro, C.M., Lourenço, N.M.T, Afonso, C.A.M.,
process for the manufacturing of plasmid biopharma-
“Enzymatic resolution and separation of sec-alcohols
ceuticals: alkaline lysis, microfiltration, salt precipita-
th
based on sustainable acylating agents”, 10 Interna-
tion and hydrophobic interaction chromatography”,
tional Chemical and Biological Engineering Confer-
Bioprocess Technology Europe: Development and
ence, CHEMPOR 2008, Braga, Portugal, September
Production of Antibodies, Vaccines and Gene Vectors, Amsterdam, The Netherlands, June-July
Monteiro, G.A., Carvalho, J., Rodgers, J., Prazeres, D.M.F., “Screening of DNA Vaccine Prototypes
Raiado-Pereira,
L.,
Santos,
J.A.L.,
Prazeres,
Encoding Antigen Targeting Sequences Against
D.M.F., Mateus, M., “Prospective hydrophobic inter-
Sleeping Sickness”, Vaccine Technology II, Albufeira,
action and pseudo-affinity membranes for chroma-
Portugal, June
tographic purification of plasmid DNA”, 7th European
Symposium on Biochemical Engineering Science –
Nascimento, K.S., Azevedo, A.M., Cavada, B.S.,
ESBES 7, Faro, Portugal, September
Aires-Barros, M.R., “Partitioning of Canavalia brasiliensis lectin in polyethylene glycol – sodium citrate
th
Silva, C.S.O., Lansalot, M., Afonso, C.A.M., Mar-
aqueous two-phase systems”, 7 European Sympo-
tinho, J.M.G., Taipa, M.A., “Biomimetic affinity poly-
sium on Biochemical Engineering Science – ESBES
mer-particles for antibody recognition”, 7th European
7, Faro, Portugal, September
Symposium on Biochemical Engineering Science –
ESBES 7, Faro, Portugal, September
Oliveira, P.H., Prazeres, D.M.F., Monteiro, G.A.,
“Plasmid DNA instability mediated by direct repeats
and type-2 insertion sequences”, EMBO conference
National Conferences
Recombination Mechanisms, Il Ciocco, Italy, May
Marques, M.P.C., Carvalho, F., Monteiro, G.A.,
Cabral, J.M.S., Fernandes, P., “Maintenance of
Pereira, A.T., Pimentel, A., Loureiro, J., Freitas,
catalytic activity in complex steroid bioconversion”,
P.P., Chu, V., Prazeres, D.M.F., Conde, J.P.,
XVI National Congress of Biochemistry, Ponta
“Colorimetric detection of antigen-antibody recogni-
Delgada, S. Miguel, Portugal, October
tion in a microfluidic channel with an integrated photodiode”, Eurosensors XXII, Dresden, Germany, September
Annual Report
2008
33
Prizes
Malcolm Lilly Award
UTL/Santander Totta Scientific Award
Tiago G. Fernandes was awarded with the 2008 Malth
Professor Joaquim M. Sampaio Cabral won the sci-
colm Lilly award on the occasion of the 7 European
entific award from the Technical University of Lisbon
Symposium on Biochemical Engineering Science
(UTL) in Biological Engineering, Biochemistry and
(ESBES-7, September 7-10, 2008), in Faro, Portugal,
Biotecnology area. This award is the recognition of
for the contribution “Exploring stem cell fate using
the scientific work and career and is sponsored by
three-dimensional cellular microarrays”, supervised
Santander Totta bank. The award ceremony took
by J.M.S. Cabral (IBB-IST) and
place on the 4th December 2008 at IST.
J.S. Dordick
(Renssealaer Polytechnic Institute, Troy, NY, USA).
FLAD Educational Innovation Awards
Best Oral Presentation
Cláudia Lobato da Silva (Professor at IST) was distin-
Martins, V.C., Cardoso, F.A., Loureiro, J., Germano,
guished with the First Annual FLAD Educational Inno-
J., Cardoso, S., Ferreira, R., Fonseca, L.P., Sousa,
vation Award. This award from the Luso-American
L., Piedade, M.S., Freitas, P.P., “Integrated spintronic
Foundation based in Lisbon recognizes the extraordi-
platforms for biomolecular detection, separation and
nary efforts of Portuguese faculty members in the MIT
counting”, Joint European Magnetic Symposia -
-Portugal Program to create educational programs at
JEMS08, Dublin, Ireland.
the highest standards of excellence.
Best Poster Awards
Idea to Product Competition
Ferreira, I.F., Azevedo, A.M., Rosa, P.A.J, Aires-
Nuno Lourenço won the 2nd Place on 6th Annual Idea
Barros, M.R., “Optimisation of temperature induced
to Product Competition -Cockrell School of Engineer-
aqueous-two phase extraction of human antibodies”,
ing Int'l Challenge with the project “Ion Jelly®- Thin-
th
28
International Symposium on the Separation of
Film Batteries”, held at University of Texas, Austin.
Proteins, Peptides and Polynucleotides - ISPPP2008,
Baden-Baden, Germany.
BES Innovation Award
Ferreira, I.F., Rosa, P.A.J., Azevedo, A.M., Aires-
Nuno Lourenço was distinguished with the BES Inno-
Barros, M.R., “Optimisation of temperature induced
vation award, area of Energy, with the Project “Ion
aqueous-two phase extraction of human antibodies”,
Jelly®- Development of Thin-Film Flexible Batteries”.
th
12
International Symposium on Preparative and
Industrial Chromatography and Allied Techniques SPICA 2008, Zurich, Switzerland.
Young researcher Award
M. Margarida Diogo and Cláudia A. M. Lobato da
Raiado-Pereira, L., Santos, J.A.L., Prazeres, D.M.F.,
Silva received Honourable Mentions in “Prémio Jov-
Mateus, M., “Prospective hydrophobic interaction and
ens Investigadores UTL/Deloitte” in the scientific area
pseudo-affinity membranes for chromatographic puri-
of Biological Engineering, Biochemistry and Biotech-
th
fication of plasmid DNA”, 7 European Symposium on
Biochemical Engineering Science - ESBES 7, Faro,
Portugal,
nology.
Staff
Faculty
Catarina Madeira
Kelany Nascimento
Joaquim M. S. Cabral
Sofia Ribeiro
Pedro Oliveira
M. Raquel Aires-Barros
Carlos Rodrigues
Luís P. Fonseca
PhD Students
Paula Rosa
D. Miguel F. Prazeres
Pedro Andrade
Francisco Santos
Marília Mateus
Susana Bernardino
Cláudia Silva
Gabriel A. Monteiro
Sara Badenes
Ana Isabel Silva
José L. A. Santos
Dragana Barros
Isabel Sousa
Cláudia Lobato da Silva
Luís Borlido
M. Ângela Taipa
Joana Carvalho
Master Students
Nídia Estrela
Joana Boura
Research Scientists
Ana Fernandes
Joana Lima
Ana M. Azevedo
Tiago Fernandes
Salomé Magalhães
M. Margarida Diogo
I. Filipa Ferreira
Pedro Fernandes
José Forte
Research Assistants
Gabriela Gomes
Luís Raiado-Pereira
Post-doctoral Fellows
João Guerreiro
Evandro Tavares
Ricardo Baptista
David Malta
Cristina Cordas
Marco Marques
Technicians
Gemma Eibes
Sofia Martins
Ricardo Pereira
Nuno Lourenço
Verónica Martins
Annual Report
2008
35
BERG
BioEngineering Research Group
Institute for Biotechnology and Bioengineering
Instituto Superior Técnico
Av. Rovisco Pais
1049-001 Lisbon
Portugal
www.ibb.pt/berg