A Portable Microscale Cell Culture System with Indirect Temperature Control

A physiologically relevant environment is essential for successful long-term cell culturing in vitro. Precise control of temperature, one of the most crucial environmental parameters in cell cultures, increases the fidelity and repeatability of the experiments. Unfortunately, direct temperature measurement can interfere with the cultures or prevent imaging of the cells. Furthermore, the assessment of dynamic temperature variations in the cell culture area is challenging with the methods traditionally used for measuring temperature in cell culture systems. To overcome these challenges, we integrated a microscale cell culture environment together with live-cell imaging and a precise local temperature control that is based on an indirect measurement. The control method uses a remote temperature measurement and a mathematical model for estimating temperature at the desired area. The system maintained the temperature at 37±0.3 °C for more than 4 days. We also showed that the system precisely controls the culture temperature during temperature transients and compensates for the disturbance when changing the cell cultivation medium, and presented the portability of the heating system. Finally, we demonstrated a successful long-term culturing of human induced stem cell–derived beating cardiomyocytes, and analyzed their beating rates at different temperatures.

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Long-term In Vitro Toxicity Models: Comparisons Between a Flow-cell Bioreactor, a Static-cell Bioreactor and Static Cell Cultures

Alternatives to Laboratory Animals ◽

10.1177/026119290203000505 ◽

2002 ◽

Vol 30 (5) ◽

pp. 515-523 ◽

Cited By ~ 13

Author(s):

Patricia Pazos ◽

Salvador Fortaner ◽

Pilar Prieto

Keyword(s):

Cell Culture ◽

Cell Viability ◽

Flow Cell ◽

Model Systems ◽

In Vitro Toxicity ◽

Protein Determination ◽

Efficient System ◽

Cytotoxicity Studies

In vitro long-term toxicity testing is becoming an important issue in the field of toxicology, and there is a need to develop new model systems that mimic human chronic exposure and its effects. The aim of this work was to test two long-term in vitro toxicity systems which are available, a flow-cell bioreactor (Tecnomouse) and a static cell bioreactor system (CELLine CL 6-well), and to compare them with the use of conventional cell culture flasks. A human cell line, Int 407, was exposed to cadmium chloride (CdCl2; 10–7–10–8M) for 4 weeks. Cell numbers and cell viabilities were determined by the trypan blue (TB) exclusion assay and from exclusion of propidium iodide (PI) as determined by flow cytometry; and cell viability and metabolic activity were determined by the MTT assay. In addition, total protein determination and cadmium uptake measurements were performed. The results obtained with TB and PI exclusion did not show clear differences in cell viability with increasing CdCl2 concentration. However, in the static cell-culture systems, an increase in MTT reduction was found at low concentrations of CdCl2. Expression of heat-shock protein (Hsp27 and Hsp70) increased differently, depending on the CdCl2 concentration applied and the system used. In summary, of the two bioreactors, the CELLine CL 6-well bioreactor was shown to be the more efficient system for performing long-term cytotoxicity studies. It is easy to handle, it permits the assessment of several endpoints, and sufficient replicates can be made available.

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Re: Cryopreservation of Testicular Tissue before Long-Term Testicular Cell Culture Does Not Alter In Vitro Cell Dynamics

The Journal of Urology ◽

10.1016/j.juro.2016.04.027 ◽

2016 ◽

Vol 196 (2) ◽

pp. 521-521

Author(s):

Craig Niederberger

Keyword(s):

Cell Culture ◽

Testicular Tissue ◽

Testicular Cell ◽

Cell Dynamics

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Indirect Temperature Measurement and Control Method for Cell Culture Devices

IEEE Transactions on Automation Science and Engineering ◽

10.1109/tase.2016.2613912 ◽

2018 ◽

Vol 15 (2) ◽

pp. 420-429 ◽

Cited By ~ 2

Author(s):

Antti-Juhana Maki ◽

Tomi Ryynanen ◽

Jarmo Verho ◽

Joose Kreutzer ◽

Jukka Lekkala ◽

...

Keyword(s):

Cell Culture ◽

Temperature Measurement ◽

Control Method ◽

And Control ◽

Measurement And Control

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A Pilot Study To Establish an In Vitro Model To Study Premature Intestinal Epithelium and Gut Microbiota Interactions

mSphere ◽

10.1128/msphere.00806-21 ◽

2021 ◽

Author(s):

Justin Gibbons ◽

Ji Youn Yoo ◽

Tina Mutka ◽

Maureen Groer ◽

Thao T. B. Ho

Keyword(s):

Cell Culture ◽

In Vitro Model ◽

Bacterial Flora ◽

Mucosal Barrier ◽

Clinical Observations ◽

Culture Models ◽

Vitro Model ◽

Cell Culture Models

The gut bacterial flora influences the development of the immune system and long-term health outcomes in preterm infants. Studies of the mechanistic interactions between the gut bacteria and mucosal barrier are limited to clinical observations, animal models, and in vitro cell culture models for this vulnerable population.

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Long-term in vitro cell culture of sinclair swine melanoma

In Vitro ◽

10.1007/bf02616139 ◽

1979 ◽

Vol 15 (5) ◽

pp. 329-341 ◽

Cited By ~ 5

Author(s):

Shirley H. Kovacs ◽

Paul F. Agris

Keyword(s):

Cell Culture ◽

In Vitro Cell Culture

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Cryopreservation of testicular tissue before long-term testicular cell culture does not alter in vitro cell dynamics

Fertility and Sterility ◽

10.1016/j.fertnstert.2015.07.1134 ◽

2015 ◽

Vol 104 (5) ◽

pp. 1244-1252.e4 ◽

Cited By ~ 26

Author(s):

Yoni Baert ◽

Aude Braye ◽

Robin B. Struijk ◽

Ans M.M. van Pelt ◽

Ellen Goossens

Keyword(s):

Cell Culture ◽

Testicular Tissue ◽

Testicular Cell ◽

Cell Dynamics

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Incubator-independent cell-culture perfusion platform for continuous long-term microelectrode array electrophysiology and time-lapse imaging

Royal Society Open Science ◽

10.1098/rsos.150031 ◽

2015 ◽

Vol 2 (6) ◽

pp. 150031 ◽

Cited By ~ 12

Author(s):

Dirk Saalfrank ◽

Anil Krishna Konduri ◽

Shahrzad Latifi ◽

Rouhollah Habibey ◽

Asiyeh Golabchi ◽

...

Keyword(s):

Cell Culture ◽

Microelectrode Array ◽

Low Cost ◽

Time Lapse ◽

Free Cell ◽

Network Architectures ◽

Hippocampal Networks ◽

Time Lapse Imaging

Most in vitro electrophysiology studies extract information and draw conclusions from representative, temporally limited snapshot experiments. This approach bears the risk of missing decisive moments that may make a difference in our understanding of physiological events. This feasibility study presents a simple benchtop cell-culture perfusion system adapted to commercial microelectrode arrays (MEAs), multichannel electrophysiology equipment and common inverted microscopy stages for simultaneous and uninterrupted extracellular electrophysiology and time-lapse imaging at ambient CO 2 levels. The concept relies on a transparent, replica-casted polydimethylsiloxane perfusion cap, gravity- or syringe-pump-driven perfusion and preconditioning of pH-buffered serum-free cell-culture medium to ambient CO 2 levels at physiological temperatures. The low-cost microfluidic in vitro enabling platform, which allows us to image cultures immediately after cell plating, is easy to reproduce and is adaptable to the geometries of different cell-culture containers. It permits the continuous and simultaneous multimodal long-term acquisition or manipulation of optical and electrophysiological parameter sets, thereby considerably widening the range of experimental possibilities. Two exemplary proof-of-concept long-term MEA studies on hippocampal networks illustrate system performance. Continuous extracellular recordings over a period of up to 70 days revealed details on both sudden and gradual neural activity changes in maturing cell ensembles with large intra-day fluctuations. Correlated time-lapse imaging unveiled rather static macroscopic network architectures with previously unreported local morphological oscillations on the timescale of minutes.

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A portable cell culture system with a simple microscope, oxygen sensor and MEA amplifier

10.14293/s2199-1006.1.sor-.pp9qfb1.v1 ◽

2019 ◽

Author(s):

Dhanesh Kattipparambil Rajan

Keyword(s):

Cell Culture ◽

Chemical Sensing ◽

Cell Imaging ◽

Modular System ◽

Disease Models ◽

Environment Control ◽

In Situ Imaging

Cell culture in-vitro is a well-known method to develop cell and disease models for studying physiologically relevant mechanisms and responses for various applications in life sciences. Conventional methods for instance, using static culture flasks or well plates, have limitations, as these cannot provide accurate tractable models for advanced studies. However, microscale systems can overcome this since they mimic the cells' natural microenvironment adequately. We have developed a portable live-cell imaging system with an invert-upright-convertible architecture and a mini-bioreactor for long-term simultaneous cell imaging and analysis, chemical sensing and electrophysiological recording. Our system integrates biocompatible cell-friendly materials with modular measurement schemes and precise environment control and can be automated. High quality time-lapse cell imaging is hugely useful in cell/disease models. However, integration of advanced in-vitro systems into benchtop microscopes for in-situ imaging is tricky and challenging. This is especially true with device based biological systems, such as lab/organ/body-on-chips, or mini-bioreactors/microfluidic systems. They face issues ranging from optical and physical geometry incompatibilities to difficulties in connectivity of flow and perfusion systems. However, the novel modular system we have developed either as an inverted or as an upright system can easily accommodate virtually any in-vitro devices. Furthermore, it can accept additional sensor or measurement devices quite freely. Cell characterization, differentiation, chemical sensing, drug screening, microelectrode-array-electrophysiological recordings, and cell stimulation can be carried out with simultaneous in-situ imaging and analysis. Moreover, our system can be configured to capture images from regions that are otherwise inaccessible by conventional microscopes, for example, cells cultured on physical or biochemical sensor systems. We demonstrate the system for video-based beating analysis of cardiomyocytes, cell orientation analysis on nanocellulose, and simultaneous long-term in-situ microscopy with pO2 and temperature sensing. The compact microscope as such is comparable to standard phase-contrast-microscopes without any detectable aberrations and is useful practically for any in-situ microscopy demands.

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Increased transcript expression levels of DNA methyltransferases type 1 and 3A during cardiac muscle long-term cell culture

Medical Journal of Cell Biology ◽

10.2478/acb-2021-0005 ◽

2021 ◽

Vol 9 (1) ◽

pp. 27-32

Author(s):

Mariusz J. Nawrocki ◽

Rut Bryl ◽

Sandra Kałużna ◽

Katarzyna Stefańska ◽

Bogumiła Stelmach ◽

...

Keyword(s):

Heart Failure ◽

Dna Methylation ◽

Cell Culture ◽

Cardiac Muscle ◽

Myocardial Cell ◽

Dna Methyltransferases ◽

Transcript Expression ◽

Expression Levels

Abstract Heart failure (HF) is one of the main causes of death worldwide. Recent studies reported altered DNA methylation in failing human hearts. This may suggest a role of DNA methylation, most frequently implicated in epigenetic control, in the development of heart failure. Here, employing RT-qPCR, we characterized transcript levels for main DNA methyltransferases (DNMTs), DNMT1, DNMT3A, and DNMT3B, mediate DNA methylation, and they have different functions that complement each other during methylation. All analyzes were performed at different stages of porcine myocardial cell primary culture. In the present study we demonstrated increasing transcript expression levels for all analyzed genes during in vitro cultivation. The changes for DNMT1 and DNMT3A seem to be particularly important, where statistically significant changes were observed. Running title: DNMTs role in cardiac muscle cell culture

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