Browse
Submit Data
Databases
API
Help

Dataset Information

19 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

3D-printed microplate inserts for long term high-resolution imaging of live brain organoids.

ABSTRACT:

Background

Organoids are a reliable model used in the study of human brain development and under pathological conditions. However, current methods for brain organoid culture generate tissues that range from 0.5 to 2 mm of size, which need to be constantly agitated to allow proper oxygenation. The culture conditions are, therefore, not suitable for whole-brain organoid live imaging, required to study developmental processes and disease progression within physiologically relevant time frames (i.e. days, weeks, months).

Results

Here we designed 3D-printed microplate inserts adaptable to standard 24 multi-well plates, which allow the growth of multiple organoids in pre-defined and fixed XYZ coordinates. This innovation facilitates high-resolution imaging of whole-cerebral organoids, allowing precise assessment of organoid growth and morphology, as well as cell tracking within the organoids, over long periods. We applied this technology to track neocortex development through neuronal progenitors in brain organoids, as well as the movement of patient-derived glioblastoma stem cells within healthy brain organoids.

Conclusions

This new bioengineering platform constitutes a significant advance that permits long term detailed analysis of whole-brain organoids using multimodal inverted fluorescence microscopy.

SUBMITTER: Oksdath Mansilla M

PROVIDER: S-EPMC8015192 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

3D-Printed Bubble-Free Perfusion Cartridge System for Live-Cell Imaging.

Project description:The advent of 3D-printing technologies has had a significant effect on the development of medical and biological devices. Perfusion chambers are widely used for live-cell imaging in cell biology research; however, air-bubble invasion is a pervasive problem in perfusion systems. Although 3D printing allows the rapid fabrication of millifluidic and microfluidic devices with high resolution, little has been reported on 3D-printed fluidic devices with bubble trapping systems. Herein, we present a 3D-printed millifluidic cartridge system with bent and flat tapered flow channels for preventing air-bubble invasion, irrespective of bubble volume and without the need for additional bubble-removing devices. This system realizes bubble-free perfusion with a user-friendly interface and no-time-penalty manufacturing processes. We demonstrated the bubble removal capability of the cartridge by continually introducing air bubbles with different volumes during the calcium imaging of Sf21 cells expressing insect odorant receptors. Calcium imaging was conducted using a low-magnification objective lens to show the versatility of the cartridge for wide-area observation. We verified that the cartridge could be used as a chemical reaction chamber by conducting protein staining experiments. Our cartridge system is advantageous for a wide range of cell-based bioassays and bioanalytical studies, and can be easily integrated into portable biosensors.

| S-EPMC7650622 | biostudies-literature

Dynamic full-field optical coherence tomography: 3D live-imaging of retinal organoids.

Project description:Optical coherence tomography offers astounding opportunities to image the complex structure of living tissue but lacks functional information. We present dynamic full-field optical coherence tomography as a technique to noninvasively image living human induced pluripotent stem cell-derived retinal organoids. Coloured images with an endogenous contrast linked to organelle motility are generated, with submicrometre spatial resolution and millisecond temporal resolution, creating a way to identify specific cell types in living tissue via their function.

| S-EPMC7429964 | biostudies-literature

Adapting the 3D-printed Openflexure microscope enables computational super-resolution imaging.

Project description:We report on a 3D printed microscope, based on a design by the Openflexure project, that uses low cost components to perform fluorescence imaging. The system is sufficiently sensitive and mechanically stable to allow the use of the Super Resolution Radial Fluctuations algorithm to obtain images with resolution better than the diffraction limit. Due to the low-cost components, the entire system can be built for approximately $1200.

| S-EPMC7255852 | biostudies-literature

Low-Cost Microplate Reader with 3D Printed Parts for under 500 USD.

Project description:A 96-well microplate reader for absorption spectroscopy was designed, constructed, and tested at a total cost of ca. 500 USD. The reduced cost of the device represents the major technical contribution of this manuscript, as costs were reduced 7 fold from previous reports. The device was able to achieve 3σ limits of detection of ca. 0.01 absorbance units (AU) over a 60 second measurement for the mid-visible wavelength range. Component parts are either commercially available, or 3D printed from plans. Analysis wavelength can be altered throughout the visible region through use of various photographic or theatrical filters. This feature allows the well plate reader to be used for typical laboratory assays such as cell population estimation by optical density (OD) at 600 nm, or enzyme-linked immunosorbent assays (ELISA) at 450 nm. This manuscript reports on the motivation and process of constructing the device, lists required parts, presents data demonstrating device function, and provides the community of scholars with plans to reproduce the work. The device can be reproduced in laboratories lacking sufficient resources to purchase commercially available options and this outcome contributes towards empowerment of individuals and equity of scientific enquiry.

| S-EPMC9103534 | biostudies-literature

Characterization of 3D Printed Stretching Devices for Imaging Force Transmission in Live-Cells.

Project description:IntroductionCell stretch is a method which can rapidly apply mechanical force through cell-matrix and cell-cell adhesions and can be utilized to better understand underlying biophysical questions related to intracellular force transmission and mechanotransduction.Methods3D printable stretching devices suitable for live-cell fluorescent imaging were designed using finite element modeling and validated experimentally. These devices were then used along with FRET based nesprin-2G force sensitive biosensors as well as live cell fluorescent staining to understand how the nucleus responds to externally applied mechanical force in cells with both intact LINC (linker of nucleoskeleton and cytoskeleton) complex and cells with the LINC complex disrupted using expression of dominant negative KASH protein.ResultsThe devices were shown to provide a larger strain ranges (300% uniaxial and 60% biaxial) than currently available commercial or academic designs we are aware of. Under uniaxial deformation, the deformation of the nucleus of NIH 3T3 cells per unit of imposed cell strain was shown to be approximately 50% higher in control cells compared to cells with a disrupted LINC complex. Under biaxial deformation, MDCK II cells showed permanent changes in the nuclear morphology as well as actin organization upon unloading, indicating that failure, plastic deformation, or remodeling of the cytoskeleton is occurring in response to the applied stretch.ConclusionDevelopment and open distribution of low-cost, 3D-printable uniaxial and biaxial cell stretching devices compatible with live-cell fluorescent imaging allows a wider range of researchers to investigate mechanical influences on biological questions with only a minimal investment of resources.

| S-EPMC6816703 | biostudies-literature

Novel 3D-Printed Cell Culture Inserts for Air-Liquid Interface Cell Culture.

Project description:In skin research, widely used in vitro 2D monolayer models do not sufficiently mimic physiological properties. To replace, reduce, and refine animal experimentation in the spirit of '3Rs', new approaches such as 3D skin equivalents (SE) are needed to close the in vitro/in vivo gap. Cell culture inserts to culture SE are commercially available, however, these inserts are expensive and of limited versatility regarding experimental settings. This study aimed to design novel cell culture inserts fabricated on commercially available 3D printers for the generation of full-thickness SE. A computer-aided design model was realized by extrusion-based 3D printing of polylactic acid filaments (PLA). Improvements in the design of the inserts for easier and more efficient handling were confirmed in cell culture experiments. Cytotoxic effects of the final product were excluded by testing the inserts in accordance with ISO-norm procedures. The final versions of the inserts were tested to generate skin-like 3D scaffolds cultured at an air-liquid interface. Stratification of the epidermal component was demonstrated by histological analyses. In conclusion, here we demonstrate a fast and cost-effective method for 3D-printed inserts suitable for the generation of 3D cell cultures. The system can be set-up with common 3D printers and allows high flexibility for generating customer-tailored cell culture plastics.

| S-EPMC9410432 | biostudies-literature

Cell-permeable organic fluorescent probes for live-cell long-term super-resolution imaging reveal lysosome-mitochondrion interactions.

Project description:Characterizing the long-term nanometer-scale interactions between lysosomes and mitochondria in live cells is essential for understanding their functions but remains challenging due to limitations of the existing fluorescent probes. Here, we develop cell-permeable organic fluorescent probes for lysosomes with excellent specificity and high photostability. We also use an existing Atto 647N dye with high brightness and excellent photostability to achieve specific labeling of mitochondria in live cells. Using these probes, we obtain dual-color structured illumination microscopy (SIM) images of dynamic physical lysosome-mitochondrion interactions in live cells at an ~90-nm resolution over a long time course of ~13?min. We successfully record the consecutive dynamic processes of lysosomal fusion and fission, as well as four types of physical lysosome-mitochondrion interactions by super-resolution imaging. Our probes provide an avenue for understanding the functions and the dynamic interplay of lysosomes and mitochondria in live cells.

| S-EPMC5670236 | biostudies-other

Mirror-enhanced scanning light-field microscopy for long-term high-speed 3D imaging with isotropic resolution.

Project description:Various biological behaviors can only be observed in 3D at high speed over the long term with low phototoxicity. Light-field microscopy (LFM) provides an elegant compact solution to record 3D information in a tomographic manner simultaneously, which can facilitate high photon efficiency. However, LFM still suffers from the missing-cone problem, leading to degraded axial resolution and ringing effects after deconvolution. Here, we propose a mirror-enhanced scanning LFM (MiSLFM) to achieve long-term high-speed 3D imaging at super-resolved axial resolution with a single objective, by fully exploiting the extended depth of field of LFM with a tilted mirror placed below samples. To establish the unique capabilities of MiSLFM, we performed extensive experiments, we observed various organelle interactions and intercellular interactions in different types of photosensitive cells under extremely low light conditions. Moreover, we demonstrated that superior axial resolution facilitates more robust blood cell tracking in zebrafish larvae at high speed.

| S-EPMC8568963 | biostudies-literature

Long-Term Maintenance and Meiotic Entry of Early Germ Cells in Murine Testicular Organoids Functionalized by 3D Printed Scaffolds and Air-Medium Interface Cultivation.

Project description:Short-term germ cell survival and central tissue degeneration limit organoid cultures. Here, testicular organoids (TOs) were generated from two different mouse strains in 3D printed one-layer scaffolds (1LS) at the air-medium interface displaying tubule-like structures and Leydig cell functionality supporting long-term survival and differentiation of germ cells to the meiotic phase. Chimeric TOs, consisting of a mixture of primary testicular cells and EGFP+ germline stem (GS) cells, were cultured in two-layer scaffolds (2LSs) for better entrapment. They showed an improved spheroidal morphology consisting of one intact tubule-like structure and surrounding interstitium, representing the functional unit of a testis. However, GS cells did not survive long-term culture. Consequently, further optimization of the culture medium is required to enhance the maintenance and differentiation of germ cells. The opportunities TOs offer to manipulate somatic and germ cells are essential for the study of male infertility and the search for potential therapies.

| S-EPMC8739976 | biostudies-literature

3D printed cellular solid outperforms traditional stochastic foam in long-term mechanical response.

Project description:3D printing of polymeric foams by direct-ink-write is a recent technological breakthrough that enables the creation of versatile compressible solids with programmable microstructure, customizable shapes, and tunable mechanical response including negative elastic modulus. However, in many applications the success of these 3D printed materials as a viable replacement for traditional stochastic foams critically depends on their mechanical performance and micro-architectural stability while deployed under long-term mechanical strain. To predict the long-term performance of the two types of foams we employed multi-year-long accelerated aging studies under compressive strain followed by a time-temperature-superposition analysis using a minimum-arc-length-based algorithm. The resulting master curves predict superior long-term performance of the 3D printed foam in terms of two different metrics, i.e., compression set and load retention. To gain deeper understanding, we imaged the microstructure of both foams using X-ray computed tomography, and performed finite-element analysis of the mechanical response within these microstructures. This indicates a wider stress variation in the stochastic foam with points of more extreme local stress as compared to the 3D printed material, which might explain the latter's improved long-term stability and mechanical performance.

| S-EPMC4846814 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data