Compact 3D printed module for fluorescence and label-free imaging using evanescent excitation

Leukocytes have an essential role in patient clinical trajectories and progression. Traditional methods of leukocyte enrichment have many significant limitations for current applications. It is demonstrated a novel 3D printing leukocyte sorting accumulator that combines with centrifugation to ensure label-free initial leukocyte enrichment based on cell density and size. The internal structure of leukocyte sorting accumulator (revealed here in a new design, leukocyte sorting accumulator-3, upgraded from earlier models), optimizes localization of the buffy coat fraction and the length of the period allocated for a second centrifugation step to deliver a higher recovery of buffy coats than earlier models. Established methodological parameters were evaluated for reliability by calculating leukocyte recovery rates and erythrocyte depletion rates by both pushing and pulling methods of cell displacement. Results indicate that leukocyte sorting accumulator-3 achieves a mean leukocytes recovery fraction of 96.2 ± 2.38% by the pushing method of layer displacement. By the pulling method, the leukocyte sorting accumulator-3 yield a mean leukocytes recovery fraction of 94.4 ± 0.8%. New procedures for preliminary enrichment of leukocytes from peripheral blood that avoid cellular damage, as well as avert metabolic and phase cycle intervention, are required as the first step in many modern clinical and basic research assays.

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3D-printed microfluidics integrated with optical nanostructured porous aptasensors for protein detection

Microchimica Acta ◽

10.1007/s00604-021-04725-0 ◽

2021 ◽

Vol 188 (3) ◽

Author(s):

Sofia Arshavsky-Graham ◽

Anton Enders ◽

Shanny Ackerman ◽

Janina Bahnemann ◽

Ester Segal

Keyword(s):

Limit Of Detection ◽

Protein Detection ◽

Target Protein ◽

Superior Performance ◽

Generic Model ◽

Label Free ◽

Uv Curable ◽

Microfluidic Platform ◽

Label Free Detection ◽

3D Printed

AbstractMicrofluidic integration of biosensors enables improved biosensing performance and sophisticated lab-on-a-chip platform design for numerous applications. While soft lithography and polydimethylsiloxane (PDMS)-based microfluidics are still considered the gold standard, 3D-printing has emerged as a promising fabrication alternative for microfluidic systems. Herein, a 3D-printed polyacrylate-based microfluidic platform is integrated for the first time with a label-free porous silicon (PSi)–based optical aptasensor via a facile bonding method. The latter utilizes a UV-curable adhesive as an intermediate layer, while preserving the delicate nanostructure of the porous regions within the microchannels. As a proof-of-concept, a generic model aptasensor for label-free detection of his-tagged proteins is constructed, characterized, and compared to non-microfluidic and PDMS-based microfluidic setups. Detection of the target protein is carried out by real-time monitoring reflectivity changes of the PSi, induced by the target binding to the immobilized aptamers within the porous nanostructure. The microfluidic integrated aptasensor has been successfully used for detection of a model target protein, in the range 0.25 to 18 μM, with a good selectivity and an improved limit of detection, when compared to a non-microfluidic biosensing platform (0.04 μM vs. 2.7 μM, respectively). Furthermore, a superior performance of the 3D-printed microfluidic aptasensor is obtained, compared to a conventional PDMS-based microfluidic platform with similar dimensions. Graphical abstract

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3D-Printed Flow Cells for Aptamer-Based Impedimetric Detection of E. coli Crooks Strain

Sensors ◽

10.3390/s20164421 ◽

2020 ◽

Vol 20 (16) ◽

pp. 4421

Author(s):

Ina G. Siller ◽

John-Alexander Preuss ◽

Katharina Urmann ◽

Michael R. Hoffmann ◽

Thomas Scheper ◽

...

Keyword(s):

High Resolution ◽

3D Printing ◽

Successful Implementation ◽

Label Free ◽

Printing Technology ◽

Flow Cells ◽

E Coli ◽

3D Printing Technology ◽

Sensor Applications ◽

3D Printed

Electrochemical spectroscopy enables rapid, sensitive, and label-free analyte detection without the need of extensive and laborious labeling procedures and sample preparation. In addition, with the emergence of commercially available screen-printed electrodes (SPEs), a valuable, disposable alternative to costly bulk electrodes for electrochemical (bio-)sensor applications was established in recent years. However, applications with bare SPEs are limited and many applications demand additional/supporting structures or flow cells. Here, high-resolution 3D printing technology presents an ideal tool for the rapid and flexible fabrication of tailor-made, experiment-specific systems. In this work, flow cells for SPE-based electrochemical (bio-)sensor applications were designed and 3D printed. The successful implementation was demonstrated in an aptamer-based impedimetric biosensor approach for the detection of Escherichia coli (E. coli) Crooks strain as a proof of concept. Moreover, further developments towards a 3D-printed microfluidic flow cell with an integrated micromixer also illustrate the great potential of high-resolution 3D printing technology to enable homogeneous mixing of reagents or sample solutions in (bio-)sensor applications.

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Photoacoustic imaging of 3D-printed vascular networks

Biofabrication ◽

10.1088/1758-5090/ac49d5 ◽

2022 ◽

Author(s):

Chenshuo Ma ◽

Wanlu Li ◽

Daiwei Li ◽

Maomao Chen ◽

Mian Wang ◽

...

Keyword(s):

3D Printing ◽

Photoacoustic Imaging ◽

Optical Resolution ◽

Blood Oxygenation ◽

Circulation System ◽

Photoacoustic Microscopy ◽

Label Free ◽

Digital Light Processing ◽

3D Printed

Abstract Thrombosis in the circulation system can lead to major myocardial infarction and cardiovascular deaths. Understanding thrombosis formation is necessary for developing safe and effective treatments. In this work, using digital light processing (DLP)-based 3D printing, we fabricated sophisticated in vitro models of blood vessels with internal microchannels that can be used for thrombosis studies. In this regard, photoacoustic microscopy (PAM) offers a unique advantage for label-free visualization of the 3D-printed vessel models, with large penetration depth and functional sensitivity. We compared the imaging performances of two PAM implementations: optical-resolution PAM and acoustic-resolution PAM, and investigated 3D printed- vessel structures with different patterns of microchannels. Our results show that PAM can provide clear microchannel structures at depths up to 3.6 mm. We further quantified the blood oxygenation in the 3D-printed vascular models, showing that thrombi had lower oxygenation than the normal blood. We expect that PAM can find broad applications in 3D printing and bioprinting for in vitro studies of various vascular and other diseases.

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Label-free counting of Escherichia coli cells in nanoliter droplets using 3D printed microfluidic devices with integrated contactless conductivity detection

Analytica Chimica Acta ◽

10.1016/j.aca.2019.04.045 ◽

2019 ◽

Vol 1071 ◽

pp. 36-43 ◽

Cited By ~ 14

Author(s):

Lucas C. Duarte ◽

Federico Figueredo ◽

Luiz E.B. Ribeiro ◽

Eduardo Cortón ◽

Wendell K.T. Coltro

Keyword(s):

Escherichia Coli ◽

Microfluidic Devices ◽

Label Free ◽

Conductivity Detection ◽

Contactless Conductivity Detection ◽

Escherichia Coli Cells ◽

3D Printed

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Label-free, mass-sensitive single-molecule imaging using interferometric scattering microscopy

Essays in Biochemistry ◽

10.1042/ebc20200023 ◽

2020 ◽

Author(s):

Nikolas Hundt

Keyword(s):

Single Molecule ◽

Cellular Systems ◽

Single Molecule Imaging ◽

Label Free ◽

Measurement Sensitivity ◽

Mass Distributions ◽

Quantitative Measurements ◽

Contrast Mechanism ◽

Cellular Components ◽

Scattering Signal

Abstract Single-molecule imaging has mostly been restricted to the use of fluorescence labelling as a contrast mechanism due to its superior ability to visualise molecules of interest on top of an overwhelming background of other molecules. Recently, interferometric scattering (iSCAT) microscopy has demonstrated the detection and imaging of single biomolecules based on light scattering without the need for fluorescent labels. Significant improvements in measurement sensitivity combined with a dependence of scattering signal on object size have led to the development of mass photometry, a technique that measures the mass of individual molecules and thereby determines mass distributions of biomolecule samples in solution. The experimental simplicity of mass photometry makes it a powerful tool to analyse biomolecular equilibria quantitatively with low sample consumption within minutes. When used for label-free imaging of reconstituted or cellular systems, the strict size-dependence of the iSCAT signal enables quantitative measurements of processes at size scales reaching from single-molecule observations during complex assembly up to mesoscopic dynamics of cellular components and extracellular protrusions. In this review, I would like to introduce the principles of this emerging imaging technology and discuss examples that show how mass-sensitive iSCAT can be used as a strong complement to other routine techniques in biochemistry.

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