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List of Publications Ludwig Gutzweiler
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Journal Articles Years: 2022 |
2017 |
2016 |
2011 | show all back to the top of all publications S. Hennig, Z. Shu, L. Gutzweiler, P. Koltay, F. von Stetten, R. Zengerle, S. M. FrühPaper-based open microfluidic platform for protein electrophoresis and immunoprobing 2022 Electrophoresis , volume : 43, issue : 4, pages : 621 - 631 L. Benning, L. Gutzweiler, K. Tröndle, J. Riba, R. Zengerle, P. Koltay, S. Zimmermann, G. B. Stark, G. FinkenzellerAssessment of hydrogels for bioprinting of endothelial cells 2017 J Biomed Mater Res A , pages : 935 - 947» show abstract « hide abstract Abstract In tissue engineering applications, vascularization can be accomplished by co-implantation of
tissue forming cells and endothelial cells (ECs), whereby the latter are able to form functional
blood vessels. The use of three-dimensional (3D) bioprinting technologies has the potential to
improve the classical tissue engineering approach because these will allow the generation of
scaffolds with high spatial control of endothelial cell allocation. This study focuses on a side
by side comparisons of popular commercially available bioprinting hydrogels (matrigel,
fibrin, collagen, gelatin, agarose, Pluronic F-127, alginate and alginate/gelatin) in the context
of their physicochemical parameters, their swelling/degradation characteristics, their
biological effects on vasculogenesis-related EC parameters and their printability. The aim of
this study was to identify the most suitable hydrogel or hydrogel combination for inkjet
printing of ECs to build pre-vascularized tissue constructs. Most tested hydrogels displayed
physicochemical characteristics suitable for inkjet printing. However, Pluronic F-127 and the
alginate/gelatin blend were rapidly degraded when incubated in cell culture medium. Agarose,
Pluronic F-127, alginate and alginate/gelatin hydrogels turned out to be unsuitable for
bioprinting of ECs because of their non-adherent properties and/or their incapability to
support EC proliferation. Gelatin was able to support EC proliferation and viability but was
unable to support endothelial cell sprouting. Our experiments revealed fibrin and collagen to
be most suitable for bioprinting of ECs, because these hydrogels showed acceptable
swelling/degradation characteristics, supported vasculogenesis-related EC parameters and
showed good printability. Moreover, ECs in constructs of preformed spheroids survived the
printing process and formed capillary-like cords. T. Gleichmann, A. Rostas, J. Wörner, E. Schleicher, L. Gutzweiler, B. Hamouda, R. Zengerle, P. Koltay, L. RieggerAtmospheric Photopolymerization of Acrylamide Enabled by Aqueous Glycerol Mixtures: Characterization and Application for Surface-Based Microfluidics 2017 Macromol Mater Eng , page : 1600518» show abstract « hide abstract Abstract Polyacrylamide usually is the material of choice for electrophoretic separation in slab gels, capillaries, and microfluidic devices. So far its polymerization requires anaerobic environments because oxygen impurities inhibit or even terminate the polymerization reaction of acrylamide. Here, it is demonstrated that gel precursor solutions with glycerol contents above 20 vol% enable direct atmospheric photopolymerization of acrylamide with no need for sealing or degassing the solution in advance. The positive effect of glycerol on the polymerization reaction is proven by simulation-validated electron paramagnetic resonance measurements. Nuclear magnetic resonance reveals that glycerol does not interfere with the reaction indicating that the observed enhancement in polymerization is owed to the low oxygen solubility of aqueous glycerol mixtures. Glycerol concentrations of >60 vol% in the gel precursor solution enable complete polymerization of volumes down to 5 nL within less than 5 s. This enables using liquid handling robots to fabricate channel-free open microfluidic structures of solid polyacrylamide hydrogel in a low-cost automated manner in a standard lab environment. L. Benning, L. Gutzweiler, K. Tröndle, J. Riba, R. Zengerle, P. Koltay, S. Zimmermann, G.B. Stark, G. FinkenzellerCytocompatibility testing of hydrogels toward bioprinting of mesenchymal stem cells 2017 J Biomed Mater Res A , volume : 105, pages : 3231 - 3241» show abstract « hide abstract Abstract Mesenchymal stem cells (MSCs) represent a very attractive cell source for tissue engineering applications aiming at the generation of artificial bone substitutes. The use of three-dimensional bioprinting technologies has the potential to improve the classical tissue engineering approach because bioprinting will allow the generation of hydrogel scaffolds with high spatial control of MSC allocation within the bioprinted construct. In this study, we have performed direct comparisons between commercially available hydrogels in the context of their cytocompatibility toward MSCs and their physicochemical parameters with the aim to identify the most suitable hydrogel for drop-on-demand (DoD) printing of MSCs. In this context, we examined matrigel, fibrin, collagen, gelatin, and gelatin/alginate at various hydrogel concentrations. Matrigel, fibrin, collagen, and gelatin were able to support cell viability, but the latter showed a limited potential to promote MSC proliferation. We concentrated our study on fibrin and collagen hydrogels and investigated the effect of hydroxyapatite (HA) inclusion. The inclusion of HA enhanced proliferation and osteogenic differentiation of MSCs and prevented degradation of fibrin in vitro. According to viscosity and storage moduli measurements, HA-blends displayed physicochemical characteristics suitable for DoD printing. In bioprinting experiments, we confirmed that fibrin and collagen and their respective HA-blends represent excellent hydrogels for DoD-based printing as evidenced by high survival rates of printed MSCs. L. Gutzweiler, S. Kartmann, K. Troendle, L. Benning, G. Finkenzeller, R. Zengerle, P. Koltay, B. Stark, S. ZimmermannLarge scale production and controlled deposition of single HUVEC spheroids for bioprinting applications 2017 Biofabrication , volume : 9 (2), page : 02502» show abstract « hide abstract Abstract We present 1.) a fast and automated method for large scale production of HUVEC spheroids based on the hanging drop method and 2.) a novel method for well-controlled lateral deposition of single spheroids by drop-on-demand printing. Large scale spheroid production is achieved via printing 1536 droplets of HUVEC cell suspension having a volume of 1 µl each within 3 minutes at a pitch of 2.3 mm within an array of 48 x 32 droplets onto a flat substrate. Printing efficiencies between 97.9% and 100% and plating efficiencies between 87.3% and 100% were achieved. Harvested spheroids (consisting of approx. 250 HUVECs each) appear uniform in size and shape. After incubation and harvesting, the spheroids are deposited individually in user-defined patterns onto hydrogels using an automated drop-on-demand dispenser setup. Controlled by an image detection algorithm focusing the dispenser nozzle, droplets containing exactly one spheroid are printed onto a substrate, while all other droplets are discarded. Using this approach an array of 6 x 3 HUVEC spheroids with intermediate distances of 500 µm embedded in fibrin was generated. Successful progress of spheroid sprouting and merging of neighboring sprouts was observed during the first 72 hours of incubation indicating a good viability of the deposited spheroids. L. Gutzweiler, T. Gleichmann, L. Tanguy, P. Koltay, R. Zengerle, L. Riegger, , , Open microfluidic gel electrophoresis: Rapid and low cost separation and analysis of DNA at the nanoliter scale 2017 Electrophoresis , volume : 38, pages : 1764 - 1770» show abstract « hide abstract Abstract Gel electrophoresis is one of the most applied and standardized tools for separation and analysis of
macromolecules and their fragments in academic research and in industry. In this work we present a
novel approach for conducting on-demand electrophoretic separations of DNA molecules in open
microfluidic (OM) systems on planar polymer substrates. The approach combines advantages of slab
gel, capillary- & chip-based methods offering low consumable costs (< 0.1 $) circumventing cost intensive microfluidic chip fabrication, short process times (5 minutes per analysis) and high
sensitivity (4 ng/µl dsDNA) combined with reasonable resolution (17 bases). The open microfluidic
separation system comprises two opposing reservoirs of 2-4 µl in volume, a semi-contact written gel
line acting as separation channel interconnecting the reservoirs and sample injected into the line via
non-contact droplet dispensing and thus enabling the precise control of the injection plug and sample
concentration. Evaporation is prevented by covering aqueous structures with PCR-grade mineral oil
while maintaining surface temperature at 15°C. The liquid gel line exhibits a semi-circular cross
section of adaptable width (~200-600 µm) and height (~30-80 µm) as well as a typical length of 15-55
mm. Layout of such liquid structures is adaptable on-demand not requiring time consuming and
repetitive fabrication steps. The approach was successfully demonstrated by the separation of a
standard label-free DNA ladder (100-1000 bp) at 100 V/cm via in-line staining and laser induced
fluorescent end-point detection using an automated prototype. L. Gutzweiler, F. Stumpf, L. Tanguy, G. Roth, P. Koltay, R. Zengerle, L. RieggerSemi-contact-writing of polymer molds for prototyping PDMS chips with low surface roughness, sharp edges and locally varying channel heights 2016 J Micromech Microeng , volume : 26, issue : 4, pages : 45018 - 45027» show abstract « hide abstract Abstract Microfluidic systems fabricated in polydimethylsiloxane (PDMS) enable a broad variety of applications and are widespread in the field of Lab-on-a-Chip. Here we demonstrate semi-contact-writing, a novel method for fabrication of polymer based molds for casting microfluidic PDMS chips in a highly flexible, time and cost-efficient manner. The method is related to direct-writing of an aqueous polymer solution on a planar glass substrate and substitutes conventional, time- and cost-consuming UV-lithography. This technique facilitates on-demand prototyping in a low-cost manner and is therefore ideally suited for rapid chip layout iterations. No cleanroom facilities and less expertise are required. Fabrication time from scratch to ready-to-use PDMS-chip is less than 5 h. This polymer writing method enables structure widths down to 140 μm and controllable structure heights ranging from 5.5 μm for writing single layers up to 98 μm by stacking. As a unique property, freely selectable height variations across a substrate can be achieved by application of local stacking. Furthermore, the molds exhibit low surface roughness (R a = 24 nm, R RMS = 28 nm) and high fidelity edge sharpness. We validated the method by fabrication of molds to cast PDMS chips for droplet based flow-through PCR with single-cell sensitivity. N Wangler, L Gutzweiler, K Kalkandjiev, C Müller, F Mayenfels, H Reinecke, R Zengerle, N PaustHigh-resolution permanent photoresist laminate TMMF for sealed microfluidic structures in biological applications 2011 J Micromech Microeng , volume : 21, page : 095009» show abstract « hide abstract Abstract We demonstrate the use of photosensitive epoxy laminate TMMF S2045 for the fabrication
and sealing of tapered microfluidic channels. The 45 μm thick resist enables the fabrication of
shallow sealed cavities featuring extreme aspect ratios of less than 1:40 (h = 45 μm, w =
2000 μm). It also provides high resolution and enables minimum feature sizes of 10 μm. For
the fabrication of free-standing structures, an aspect ratio of up to 7:1 was achieved. The
dry-film photoresist can be applied easily by lamination onto structured substrates. The total
thickness variation of the resist across a 100 mm wafer was determined to be less than
±0.6 μm. Process parameters for the fabrication and sealing of various micro-channels are
discussed and optimized in this paper. The main focus was to minimize thermal impact during
lamination, soft-bake, exposure and post–exposure bake, which could lead to lid sagging or
channel clogging due to liquefaction of uncured resist. We tested TMMF according to ISO
10995-5 and found it to be non-cytotoxic, enabling its use for biological applications.
Swelling of less than 5% for incubation of the dry-film resist in several biologically relevant
solvents, buffers and cleaning solutions was observed. K Kalkandjiev, L Riegger, D Kosse, M Welsche, L Gutzweiler, R Zengerle, P KoltayMicrofluidics in silicon/polymer technology as a cost-efficient alternative to silicon/glas 2011 J Micromech Microeng , volume : 21, page : 025008 (8p» show abstract « hide abstract Abstract We investigate TMMF photopolymer as a cost efficient alternative to glass for the liquid tight sealing of high density silicon microchannels. TMMF enables low temperature sealing and access to structures underneath via lamination and standard UV lithography instead of costly glass machining and anodic bonding. TMMF is highly transparent and has a low autofluorescence for wavelengths larger than 400 nm. As the photopolymer is too thin for implementing bulky World-to-Chip-interfaces, we propose adhesive bonding of COC modules. All materials were tested according ISO 10993-5 and showed no cytotoxic effects on the proliferation of L929 cells. To quantify the cost efficiency of the proposed techniques, we used an established Si/Pyrex nanoliter dispenser as a reference and replaced structured Pyrex wafers by TMMF laminates and COC modules. Thus, consumable costs and time effort were reduced by 90 % respectively 35 % for the sealing and 80 % respectively 75 % for implementing the World-to-Chip interface. Liquid tightness was proved by applying a pressure of 0,2 MPa for 5 h without delamination or cross talk between neighbouring microchannels separated from each other by 100 µm. In contrast to anodic bonding, the proposed techniques are tolerant to surface inhomogenities. They enable manufacturing of silicon/polymer microfluidics at significantly lower costs and without compromising the performance compared to corresponding silicon/glass devices.
Conference papers Years: 2022 |
2020 |
2018 |
2016 |
2014 |
2013 |
2011 |
2010 | show all back to the top of all publications S. Hennig, Z. Shu, L. Gutzweiler, P. Koltay, F. von Stetten, R. Zengerle, S. M. Früh“Paper-based open microfluidics platform for automatic protein analysis 2022 SLAS Europe 2022 Conference and Exhibition, Dublin, Ireland, May 26, 2022 A. Brunauer, B. Breiner, S. Hennig, D. Kainz, R. Verboket, B. Johannsen, D. Baumgartner, K. Mitsakakis, L. Gutzweiler, Z. Shu, P. Koltay, T. Hutzenlaub, N. Paust, R. Zengerle, F. von Stetten, S. M. FrühActuation principles for bioanalytical platforms to combat infectious diseases 2020 Virtual EMBL Conference: Microfluidics: Designing the Next Wave of Biological Inquiry 2020, 13.-15.07.2020 K. Tröndle, S. Kartmann, L. Gutzweiler, R. Zengerle, P. Koltay, S. ZimmermannBioprinting with spheroids: Automated large-scale production and deposition 2018 3D Cell Culture 2018, 5. - 7. Juni 2018, Freiburg T. Gleichmann, B. Hamouda, L. Gutzweiler, M. A. Graewert, R. Zengerle, P. Koltay, L. RieggerAnalytical protein affinity chromatography in nl volumes in on-demand written liquid lines 2016 20th International Conference on Miniaturized Systems for Chemistry and Life Sciences, µTAS 2016, Dublin / Irland, 09. – 13.10.2016 L. Gutzweiler, T. Gleichmann, P. Koltay, R. Zengerle, L. RieggerOpen Microfluidics for Lab-on-a-Chip Applications 2016 2016 International Conference of Microfluidics, Nanofluidics and Lab-on-a--Chip, Dalian/China, June 10-12, 2016 » show abstract « hide abstract Abstract In this work we introduce bioanalytical applications previously realized on lab-on-chip devices that have been successfully transferred to the field of open microfluidics (OM). Open microfluidic systems are microfluidic networks on planar substrates without being enclosed by solid interfaces and thus not having rigid boundaries [1]. Applications in open microfluidic systems can be conducted on-demand with often less amounts of reagents required. Furthermore, time consuming & cost intensive fabrication of microchips can be omitted using planar low-cost polymer substrates. In this work, we present DNA and protein electrophoresis as well as microbatch protein crystallization in an open microfluidic environment. F. Stumpf, L. Gutzweiler, L. Tanguy, P. Koltay, R. Zengerle, L. RieggerA flexible method for rapid-prototyping of PDMS microfluidic chips for droplet based applications using direct-written polymer master structures 2014 Biosensors 2014, Melbourne, Australia, 27.05.2014 – 30.05.2014 J. Riba, L. Gutzweiler, L. Riegger, P. Koltay, R. Zengerle, A. GrossA picoliter dispenser with disposable cartridges for precise and contact-free injection of DNA into open microfluidic structures 2014 2nd MFHS Conference 2014, Freiburg, 08 - 10.Okt. 2014 , pages : 84 - 87» show abstract « hide abstract Abstract We present a new disposable liquid handling
system addressing the lower picoliter volume
range and allowing for precise and contact-free
injection of biopolymer samples into open
microfluidic structures. Droplet placement
precision of single stranded DNA (ssDNA)
solutions up to 50 μM and a DNA reference
ladder is investigated by high resolution optical
monitoring. We present three different sample
injection methods that account for evaporation
protection of picoliter samples. Finally, we show
the applicability of the dispenser by a successful
electrophoretic separation via injecting ~70 pL of
a DNA solution into a 200-300 μm wide gel line. L. Gutzweiler, T. Gleichmann, P. Koltay, R. Zengerle, L. RieggerSemi-contact writing technology & applications 2014 2nd MFHS Conference 2014, Freiburg, 08 - 10.Okt. 2014 , pages : 96 - 99» show abstract « hide abstract Abstract We present an adaption of a direct ink writing
approach to facilitate prevalent standard
applications like fabricating masters for PDMS
casting, conducting path structuring and
generation of open microfluidic structures
towards gel electrophoresis. In all presented
applications, flexibility is increased and standard
structuring processes can be substituted offering
the ability for low-cost fabrication with less
required expertise. In contrast to existing
approaches low to medium viscous liquids are
applied due to passive capillary forces. T. Gleichmann, L. Gutzweiler, R. Zengerle, P. Koltay, L. RieggerSmart open microfluidics: an automated platform for the dynamic generation of fluidic structures down to the sub nl-range 2014 2nd MFHS Conference 2014, Freiburg, 08 - 10.Okt. 2014 , pages : 161 - 164» show abstract « hide abstract Abstract We developed an open microfluidic (OM) [1]
platform for the computer-assisted generation of
fluidic structures virtually on demand. On using a
set of piezo-driven dispenser modules, namely
PipeJetTM [2] and Nano-Jet [3] (BioFluidix
GmbH, Freiburg, Germany), and a peristaltic
pump as well the system enables handling of
liquids from the mL- down to the pL-range covering
up to nine orders of magnitude in volume.
The integration of multiple structuring methods
like semi-contact writing (SCW) or non-contact
dispensing allows for processing droplet arrays or
continuous geometries of desired dimension,
using low to medium viscous fluids and a number
of different substrate materials. L. Gutzweiler, F. Stumpf, L. Riegger, P. Koltay, R. Zengerle, L. TanguyA flexible method for rapid-prototyping of PDMS microfluidic chips using direct-written polymer-master-structures 2013 MicroTAS 2013, Freiburg, 27.10.- 31.10.2013 , pages : 1409 - 1411 L. Gutzweiler, L. Riegger, P. Koltay, R. Zengerle, L. TanguyDNA Gelelektrophorese in offenen mikrofluidischen Systemen 2013 Microsystemtechnik (MST) Kongress 2013, Aachen, 14. - 16.10.2013 , pages : 380 - 383» show abstract « hide abstract Abstract In diesem Beitrag wird die elektrophoretische Trennung von DNA in digital gedruckten offenen mikrofluidischen Linien präsentiert. Im Gegensatz zur gängigen, Chip-basierten Kapillarelektrophorese in mikrostrukturierten Glas-Chips, kommen bei diesem Ansatz mit Platin-Elektroden versehene, planare Polyimid-Substrate zum Einsatz. Eine mittels Teilkontaktverfahren generierte 200 μm breite Gellinie verbindet die Elektroden und fungiert als Trennkanal, in wel-chen kontaktfrei 500 pL der zu untersuchenden Probe dosiert werden. Um Verdunstungseffekte zu unterbinden wird das Gel bis zum Taupunkt gekühlt und mit Mineralöl überschichtet. Durch das Anlegen eines elektrischen Feldes an den Elektroden für 80 s konnte die Trennung der DNA-Fragmente (56 bp-Cy5 und 112 bp-Cy5, 10 μM) erfolgreich gezeigt werden. L. Tanguy, L. Gutzweiler, P. Koltay, R. Zengerle, L. RieggerOn-demand electrophoretic separation of DNA in written gel lines on planar substrates 2013 Transducers 2013, Barcelona, Spain, 16. – 18.06.2013 , pages : 1223 - 1226» show abstract « hide abstract Abstract We report a new approach to perform on-demand
electrophoretic separation of DNA. In contrast to standard
chip-based capillary electrophoresis in micromachined
glass chips, we apply a planar polyimide substrate, write
200 μm wide gel lines bridging two Pt-electrodes and
inject 500 pl sample volumes in non-contact manner. The
gel is covered with mineral oil to inhibit evaporation.
Subsequently, an electrical field is applied for 80 s and the
separation of the DNA molecules (56 bp-Cy5 and
112 bp-Cy5, 10 μM) is successfully demonstrated. Kiril Kalkandjiev, Ludwig Gutzweiler, Roland Zengerle, Peter KoltayKostengünstige Fertigung großflächiger Druckköpfe durch Lamination von Trockenlacken auf vorstrukturierte Polymersubstrate 2011 Mikrosystemtechnik-Kongress, Darmstadt, Deutschland, October 10-12 , pages : 883 - 886» show abstract « hide abstract Abstract In diesem Beitrag wird ein mikrofluidischer Kunststoff-Druckkopf als Alternative zu seinem etablierten Silizium/Glas-Äquivalent beschrieben. Der Druckkopf wurde durch die Kombination spanabhebender und lithographischer Verfahren hergestellt, welche für großflächige Bauteile mit mikrostrukturierten Durchgangsöffnungen besonders geeignet sind. Er beinhaltet 24 Düsen mit einem Durchmesser von 50 μm, die über mikrofluidische Kanäle mit jeweils einem Reservoir verbunden sind. Alle Mikrostrukturen wurden im Trockenlack TMMF strukturiert, das Druckkopfinterface besteht aus PMMA. Die verhältnismäßig große Grundfläche von 36 x 18 mm wird durch die standardisierte Anordnung der Reser-voire im Rastermaß der 384er Mikrotiterplatte und die notwendige Interaktion mit einem externen Piezoaktor vorgege-ben. Grundlegende Anforderungen wie die leckagefreie Verbindung zwischen den Reservoiren und den entsprechenden Düsen, die Realisierung eines gedeckelten mikrofluidischen Kanalsystems sowie die Durchgängigkeit und Homogenität der Düsen wurden reproduzierbar umgesetzt. Durch entsprechende Nachbehandlung wurden die mikrofluidischen Strukturen mit entsprechenden Oberflächenfunktionalitäten versehen, die eine kapillare Befüllung der Düsen und den parallelen Ausstoß von Einzeltropfen nach dem etablierten TopSpot® Verfahren ermöglichen. Kiril Kalkandjiev, Ludwig Gutzweiler, Mathias Welsche, Roland Zengerle, Peter KoltayA novel approach for the fabrication of all-polymer microfluidic devices 2010 roc. of IEEE-MEMS, Hong-Kong, China, January 24-28 , pages : 1079 - 1082» show abstract « hide abstract Abstract This paper describes a process sequence for the fabrication of all-polymer microfluidic chips based on the multilayer lamination of TMMF dry resist (TOK, Japan) on a pre-patterned PMMA substrate. The sequence provides a simple way to meet major microfluidic requirements like the fabrication of embedded microchannels, nozzles and intercon-necting vias as well as their accurate integration into a chip-interface without additional materials and bonding procedures. We demonstrate the appli-cability of the sequence by manufacturing and test-ing a 24-channel TopSpot printhead. Additionally, non-cytotoxicity of TMMF was confirmed in cell culture experiments and different methods for sur¬face modification were investigated. Credits: SILK Icons by http://www.famfamfam.com/lab/icons/silk/