2018 XV International Scientific Conference on Optoelectronic and Electronic Sensors (COE)

Inkjet 3D printing of labs-on-a-chip with optical

detection
Rafaá Walczak
Faculty of Microsystem Electronics and
Photonics
Wrocáaw University of Science and
Technology
Wrocáaw, Poland
rafal.walczak@pwr.edu.pl

Abstract— Inkjet 3D printing is one of the techniques of
additive manufacturing that is successfully adapted to
requirements of microfluidic structures development. In this
paper result of author’s research works on applicability of inkjet
3D printing is described. Special attention was paid to optical
properties of the printed microstructures. Than examples of
various microfluidic structures are briefly described. Finally a
discussion on limits and potential directions of inkjet 3D printing
development for microfluidics is presented.
Keywords—3D printing, lab-on-a-chip, optical detection
I. INTRODUCTION
Fabrication of microfluidic structures or systems often called
also as labs-on-a-chip includes many microengineering
techniques applied with different materials. Glass
microstructures are fabricated mainly by wet etching [1, 2],
silicon structures by wet or dry etching [3, 4], polymer chips
by milling, moulding or injecting [5, 6], polydimethylsiloxane
(PDMS) by soft lithography [7, 8]. Regardless of applied
material and technology fabrication of the microfluidic
structure usually involves two main steps: (1)
microengineering of the three dimensional microstructure by
one of mentioned above technique (channels, chambers,
mixers etc.) in the substrate(s) followed by (2) encapsulation
of the microstructures by bonding of the substrates. Three
dimensional printing (3Dp) offers new approach to fabrication
of the microfluidic structures where the whole microstructure
is automatically printed from a computer design in a single
process without bonding/assembling of additional
substrates/components.
Increase of interest in applicability of 3D printing as a tool for
fabrication of the microfluidic structure is observed during last
five years [9]. It is main due decrease of the 3D printers price
and increased printing resolution down to dimensions
characteristic microstructures [10]. Among many
commercially available 3D printing techniques only few are
successfully adapted to microfluidic structure fabrication:
sterolitography (STL) [11,12], fused deposition modelling
(FDM) [13, 14] and inkjet 3D printing (i3Dp) [15, 16].
Numerous papers in top-rated journals describe development
of the 3D printed chips for chemistry and bioanalytics [17-
19]. Also interesting reviews are published and summarize
achievements of many groups around the word [20-27]. The
main conclusion from all the papers is that experts identify
four major impacts of 3Dp on R&D works and both
established companies, and new business start-ups involved in
lab-on-a-chip development, fabrication and applications. They
are: economic low volume production, increased geometric
complexity, product personalization, supply chain
compression. The main challenge in successful applicability of
the 3Dp in development of microfluidic structures and
systems is that none of the printing technique is dedicated to
print microfluidic structures. The techniques must be first
adopted to the specific requirements of the microengineering.
These requirements are accurate representation of each
designed microfluidic structures printed with submili- and
microresolutions, roughness of the microstructures (in most
cases microfluidics like smooth surfaces) and possibility of
detection of the optical signals generated inside the printed
structures.
In this paper discussion on the listed above issues is carried
out on the base of short literature review focused on optical
detection in microfluidic structures and author’s works on
applicability of the inkjet 3D printing. Than examples of
microfluidic structures are briefly – all with involved optical
detection. In conclusion some main remarks on i3Dp
properties as well as limits and potential directions of 3Dp
development for microfluidics are discussed.
II. OPTICAL DETECTION IN 3D PRINTED MICROFLUIDICS
Optical detection is one of the most commonly used detection
method applied together with microfluidic structures [28]. It is
also natural tendency to apply optical detection combined with
3D printed microstructures. Three main detection techniques
are considered as techniques that can successfully co-work
with 3D printed chips: optical observation, spectrophotometry
and fluorimetry.
Optical observation of the microfluidic channels seems to be
the simplest method. It can be realized by naked eye, camera
or microscope. The main requirement in relation to the 3D
printed chip is transparency of the printed material, high
enough to identify various colors or changes of colors. This
detection technique is used mainly to verify or visualize proper
operation of the 3D printed microstructure [29, 30], observe
different flow profiles [31], on-chip cell imagining under
phase contras [12] or observation of droplet generation [32].

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 2018 XV International Scientific Conference on Optoelectronic and Electronic Sensors (COE)
Spectrophotometry requires source of light in wide range of
wavelengths combined with measurement cell (cuvette) and
spectrophotometer. Two approaches of spectrophotometry can
be considered with microfluidic structures. In the firs approach
3D printed measurement cell is a part of optical path without
integrated optical components. The light passes through the
measurement cell walls. Thus high transparency of the walls
are strongly recommended (i.e. 400 μm thick walls and 500
μm long optical path in microcuvette [33]) or post-printing
assembling of optically transparent window is necessary (i.e.
polystyrene window glued to 3D printed low-volume cuvette
with 10 mm long optical path [34]). In the second approach
some optical components (i.e. optical fibres) are integrated
with the microfluidic circuit. Walczak et al presented
successful beverages identification in microfluidic flow-
through cell with integrated plastic optical fibers [35].
Fluorimetry requires two main components with spectral
characteristics tailored each to other. Fluorescence inducing
light source which emission spectral characteristics is covering
absorbance spectra of the fluorochrome and photodetector
with optical filters with spectral characteristic matched to
emission spectra of the fluorochrome. Many aspects of the
fluorescence detection in various microfluidic chips were in
details discussed by Walczak et al [36 - 38]. In case of 3D
printed chips important issues are transparency of the chip and
autofluorescence of the chip body material. Both of them are
in details discussed in the next chapter. The 3D printed chips
are used to fluorescence cell imagining [12] or recently also to
separate DNA ladder with on-chip fluorescence detection [39].
One of the key elements in successful application of 3D
printing in microfluidics is knowledge on optical properties of
the material used as building material of 3D printed
microstructures. Other important issues like geometrical or
mechanical play also important role and are briefly discussed
in the next chapter.
Inkjet 3D printing utilize drop-by-drop building of a single
layer (slice) of the printed element. Each droplet layer is
flattened by heated roller and cured by UV light. The
deposited layers form three-dimensional structures. A
comparison between printouts obtained from four printers was
conducted which considered the dimensional fidelity, shape
conformity and surface quality of the printed test structures
[35]. Based on the results of the preliminary works an inkjet
3D printer ProJet3510SD (3D Systems Inc., USA) was
selected as an optimal solution compromising resolution,
accuracy, fidelity and surface roughness important from the
point of view of microfluidic structures fabrication [35].
Applied in the experiments building material (VisiJet M3
Crystal) is described by the manufacturer as transparent
material. On the other hand it is well known that optical
detection is commonly used method of detection in lab-on-a-
chip technique with special attention paid to fluorescence
detection. Successful fluorescence detection require high
transparency of the material (introduction and collection of the
fluorescence light through microfluidic structure walls) and
low or negligible autofluorescence of the material due to
possible falsification of the optical signals readout [37]. Both
parameters – transparency and autofluorescence – were
studied for Visijet M3 Crystal material [42]. It was found that
the printed microstructures are semi-transparent. The highest
transmittance for visible light region (61% for λ=650 nm) was
for 64 μm thick layer printed with layer orientation parallel to
light propagation direction (Fig. 1). The transmittance
decreased almost linearly down to 20-30% for 320 μm thick
layers. Thus optical detection is possible but thickness of the
printed walls should be as small as possible.
Although theoretical minimal thickness of a single printed
layer is 32 μm the minimal properly (without defects and
perforation) printed layer was two times thicker. Thickness of
the layer was measured by the precise thickness meter after
printout. Obtained values of the transmittance are significantly
lower than for glass applied in microfluidics (i. e. borofloat
glass visible light transmittance of 0.2 mm thick layer is above
90% according to manufacturers’ data). It is also difficult to
compare obtained values to other groups’ results because
transmittance values are reported very rarely. Tothill et al
measured attenuation level of polyactic acid layer (PLA)
printed by fused deposition modelling technique (FDM) [33].
They reported attenuation to be about 20% for 0.25 mm thick
layer and 633 nm wavelength, and varying slightly on printing
speeds. It corresponds to 80% transmittance. It is significantly
higher transmittance than for the inkjet printed layer (~27%
for 0.25 mm thick layer) but printing techniques (FDM vs
i3Dp) and printed materials (PLA vs UV curable resin) are
different.
The autofluorescence of the polymer was investigated for three
most commonly used fluorescence inducing wavelengths
(light emitting diode λ=488 nm and lasers λ=532 nm, and
λ=635 nm, optical power 5-25 mW). The autofluorescence
was not observed. Thus thin layers of the inkjet printed
structures may be used as parts of the optical detection
systems in microfluidic structures were optical detection is
involved. Zhu et al concluded theirs works that fluorescence
microscopy revealed significant autofluorescence and sub-
standard optical transparency of the majority of 3D printed
polymers [43]. Resin used in sterolitography (VisiJet SL Clear
by Stratasys, USA) was found as the only suitable polymer for
fabricating imagining grade structures comparable to optical
characteristics of PDMS (material commonly used in
microfluidics). But they did not gave any numerical values of
the autofluorescence or transmittance. Thus the following
question arise: what is minimal level of transmittance or
maximal tolerable level of autofluorescence for 3D printed
structures combined with optical detection? From practical
point of view it can be concluded that as long as optical
signals are properly identified and detected for standard
measurement conditions (instrumental and biochemical) the
optical properties of the chip (and material) are acceptable.
Optimization of the chip construction (i.e. geometry, printing
conditions) is necessary when the signals are close to limits of
detection caused by the chip properties reasons.
Main conclusion of the carried out works are that i3Dp has
resolution and accuracy that enables fabrication of
microfluidic structures but with minimal diameter of around
 2018 XV International Scientific Conference on Optoelectronic and Electronic Sensors (COE)

200 μm and length limited by efficiency of the support necessary. It is impossible by the use of traditional
material removing [44, 45]. Optical properties of the building microfluidic chips that have closed architecture.
material are acceptable – thin walls can be used to transmit a)
visible light and autofluorescence of the polymer was not
observed. 3D printed labs-on-a-chip with optical detection
One of the most spectacular application of i3Dp is
development of disposable chips for on-chip gel
electrophoresis [39]. The lab-on-a-chip is printed in a single
process and cleaned according to the optimal procedure. The
chip contains dosing and separation microchannels with 500
μm diameter and 10/23 mm length respectively, four
microreservoirs for buffer and sample introduction (10 μL
volume each) and fluorescence detection area with thinned
walls at the end of the separation microchannel (Fig. 1a).
Successful separation of DNA 50-800 bp ladder in POP-4 gel
(A&A Biotechnology, Poland) has been achieved in less than
10 minutes. Theoretical number of separation plates achieved
for the developed chip was at level of 10 000 (Fig. 1b). It is b)
significantly lower than the best result reported by Walczak et
al in all glass chip (up to 600 000 of plates for 30 μm deep,
500 μm width and 25 mm long separation channel) but printed
microchannels have one order of magnitude larger channel in
diameter [38].

a)

Fig. 2. Modular chips for gel electrophoresis with optical (fluorescence)

b)
Fig. 1. Inkjet 3D printed chip for gel electrophoresis with fluorescence
detection: a) view of the chip during main separation stages: sample injection
into separation channel, sample flow inside separation channel and sample at
the detection area, b) electropherogram of 50-800 bp DNA ladder separation
Further improvement of the separation efficiency can be
obtained by decrease of the separation channel diameter or
increase of the channel length. It cannot be easily achieved
due to limited efficiency of the support material removing
from narrow and long microchannels. This problem can be
solved by prosed recently modular configuration of the lab-on-
a-chip for gel electrophoresis (Fig. 2) [46]. In this concept the
chip is configured with functional modules: T-type cross-
section, separation microchannel(s), optical detection module
(also with standard optical fibers connections), outlet module
etc. (Fig. 2a). In the first experiments proper separation of the
DNA ladder was successfully achieved (Fig. 2b). What more,
proposed modular configuration enables also transfer of the
sample or separated fraction between various configurations if
detection modules: a) types of modules and view of assembled chip,
b) example of 20-200 bp DNA ladder separation
III. CONCLUSIONS AND DISCUSSION
Inkjet 3D printing was originally developed for rapid and
precise fabrication of some mechanical elements but it was
also successfully adopted to the needs of microengineering.
Due to specific requirements of microfluidics fabrication
development of the microstructures require knowledge on
main properties of the i3Dp. Comprehensive research works
of the team headed by author resulted in determination of main
morphological, optical and mechanical features of the i3DP
technique. Collected knowledge and skills were next used to
develop some microfluidic structures and more complicated
systems. In conclusions of these works, i3Dp is powerful tool
that offers great flexibility in designing and fabrication of truly
3D structures with dimension at submillimetre to tenths of
micrometers range. Completely new approach is used in
comparison to traditional techniques – the microstructure is
printed directly from computer design in single process as
fully functional element without additional assembling and in
time counted in hours. The disadvantages of i3Dp are
influence of printing resolution and orientation on morphology
and optical/mechanical properties of the printouts and limited
diameter/length of the microchannel caused by efficiency of
support material remove. Aside from fabrication of the
structures directly from computer design that is without doubts
added value of 3D printing, the next breakthrough step in i3Dp
proliferation (other 3D printing techniques too) can be
 2018 XV International Scientific Conference on Optoelectronic and Electronic Sensors (COE)
printing at the same time with materials with different
properties and functions (multimaterial printing). Although
multicolor printing is already available inkjet 3D printing of
materials with different properties (for example conductive,
dielectric or optically passive/active) is still a major challenge.
ACKNOWLEDGMENT
The works are financed by Polish National Science Centre
(NCN) under frame of the project no 2013/10/E/ST8/0342.
Special thanks to Mr. Krzysztof Adamski for fruitful co-
operation in development of presented here printed structures.
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