scholarly journals Enhancement of Virus Infection Using Dynamic Cell Culture in a Microchannel

Micromachines ◽  
2018 ◽  
Vol 9 (10) ◽  
pp. 482
Author(s):  
Jeong Kim ◽  
Hye Choi ◽  
Chul Kim ◽  
Hee Jin ◽  
Jae-sung Bae ◽  
...  
Keyword(s):  
Cell Culture ◽  
Virus Infection ◽  
Soft Lithography ◽  
Fluorescent Protein ◽  
Microfluidic Devices ◽  
Petri Dish ◽  
Culture Method ◽  
Dynamic Culture ◽  
Conventional Culture ◽  
Dynamic Cell

With increasing interest in induced pluripotent stem cells (iPSCs) in the field of stem cell research, highly efficient infection of somatic cells with virus factors is gaining importance. This paper presents a method of employing microfluidic devices for dynamic cell culture and virus infection in a microchannel. The closed space in the microchannel provided a better environment for viruses to diffuse and contact cell surfaces to infect cells. The microfluidic devices were fabricated by photolithography and soft lithography. NIH/3T3 fibroblast cells were cultured in the microfluidic device in static and dynamic conditions and compared with the conventional culture method of using Petri dishes. Virus infection was evaluated using an enhanced green fluorescent protein virus as a model. Dynamic culture in the microchannel showed similar growth of cells to that in Petri dish culture, but the virus infection efficiency was four-times higher. The proposed dynamic culture system could be useful in iPSC research by providing efficient virus infection tools.

scholarly journals Microfluidic chambers using fluid walls for cell biology

2018 ◽  
Vol 115 (26) ◽  
pp. E5926-E5933 ◽  
Author(s):  
Cristian Soitu ◽  
Alexander Feuerborn ◽  
Ann Na Tan ◽  
Henry Walker ◽  
Pat A. Walsh ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Biology ◽  
Mammalian Cells ◽  
Soft Lithography ◽  
Petri Dish ◽  
Microtiter Plate ◽  
The Media ◽  
Downstream Analysis ◽  
Microfluidic Chambers ◽  
Made In

Many proofs of concept have demonstrated the potential of microfluidics in cell biology. However, the technology remains inaccessible to many biologists, as it often requires complex manufacturing facilities (such as soft lithography) and uses materials foreign to cell biology (such as polydimethylsiloxane). Here, we present a method for creating microfluidic environments by simply reshaping fluids on a substrate. For applications in cell biology, we use cell media on a virgin Petri dish overlaid with an immiscible fluorocarbon. A hydrophobic/fluorophilic stylus then reshapes the media into any pattern by creating liquid walls of fluorocarbon. Microfluidic arrangements suitable for cell culture are made in minutes using materials familiar to biologists. The versatility of the method is demonstrated by creating analogs of a common platform in cell biology, the microtiter plate. Using this vehicle, we demonstrate many manipulations required for cell culture and downstream analysis, including feeding, replating, cloning, cryopreservation, lysis plus RT-PCR, transfection plus genome editing, and fixation plus immunolabeling (when fluid walls are reconfigured during use). We also show that mammalian cells grow and respond to stimuli normally, and worm eggs develop into adults. This simple approach provides biologists with an entrée into microfluidics.

scholarly journals PDMS-Based Microfluidic Devices for Cell Culture

Inventions ◽  
2018 ◽  
Vol 3 (3) ◽  
pp. 65 ◽  
Author(s):  
Stefania Torino ◽  
Brunella Corrado ◽  
Mario Iodice ◽  
Giuseppe Coppola
Keyword(s):  
Cell Culture ◽  
Cell Cultures ◽  
Soft Lithography ◽  
Microfluidic Devices ◽  
Biological Research ◽  
Culture Chamber ◽  
Culture Methods ◽  
Critical Issues ◽  
Scaling Down ◽  
Cell Culture Methods

Microfluidic technology has affirmed itself as a powerful tool in medical and biological research by offering the possibility of managing biological samples in tiny channels and chambers. Among the different applications, the use of microfluidics for cell cultures has attracted much interest from scientists worldwide. Traditional cell culture methods need high quantities of samples and reagents that are strongly reduced in miniaturized systems. In addition, the microenvironment is better controlled by scaling down. In this paper, we provide an overview of the aspects related to the design of a novel microfluidic culture chamber, the fabrication approach based on polydimethylsiloxane (PDMS) soft-lithography, and the most critical issues in shrinking the size of the system.

scholarly journals Electrospun hydrogels for dynamic culture systems: advantages, progress, and opportunities

2021 ◽  
Author(s):  
M. Gregory Grewal ◽  
Christopher B. Highley
Keyword(s):  
Cell Culture ◽  
Culture Systems ◽  
Dynamic Culture ◽  
Cell Culture Systems ◽  
Dynamic Cell ◽  
And Control ◽  
Soft Biomaterials

Hydrogel nanofibers build on established soft biomaterials to enable design and control of unique, dynamic cell culture systems.

scholarly journals A rapid-prototyped moulding system towards optimised scalable fabrication of elastomeric microfluidic devices

2021 ◽  
Author(s):  
Christine Poon ◽  
Albert Fahrenbach
Keyword(s):  
3D Printing ◽  
Microfluidic Device ◽  
Lean Manufacturing ◽  
Soft Lithography ◽  
Low Cost ◽  
Microfluidic Devices ◽  
Petri Dish ◽  
Device Fabrication ◽  
Alternative Techniques ◽  
Optimisation Techniques

3D printing and makerspace technologies are increasingly explored as alternative techniques to soft lithography for making microfluidic devices, and for their potential to segue towards scalable commercial fabrication. Here we considered the optimal application of current benchtop 3D printing for microfluidic device fabrication through the lens of lean manufacturing and present a straightforward but robust rapid prototyped moulding system that enables easy estimation of more precise quantities of polydimethylsiloxane (PDMS) required per device to reduce waste and importantly, making devices with better defined depths and volumes for (i) modelling gas exchange and (ii) fabrication consistency as required for quality-controlled production. We demonstrate that this low-cost moulding step can enable a 40 – 300% reduction in the amount of PDMS required for making individual devices compared to the established method of curing approximately 30 grams of PDMS prepolymer overlaid on a 4” silicon wafer master in a standard plastic petri dish. Other process optimisation techniques were also investigated and are recommended as readily implementable changes to current laboratory and foundry-level microfluidic device fabrication protocols for making devices either out of PDMS or other elastomers. Simple calculators are provided as a step towards more streamlined, software controlled and automated design-to-fabrication workflows for both custom and scalable lean manufacturing of microfluidic devices.

scholarly journals A rapid-prototyped moulding system towards optimised scalable fabrication of elastomeric microfluidic devices

2021 ◽  
Author(s):  
Christine Poon ◽  
Albert Fahrenbach
Keyword(s):  
3D Printing ◽  
Microfluidic Device ◽  
Lean Manufacturing ◽  
Soft Lithography ◽  
Low Cost ◽  
Microfluidic Devices ◽  
Petri Dish ◽  
Device Fabrication ◽  
Alternative Techniques ◽  
Optimisation Techniques

3D printing and makerspace technologies are increasingly explored as alternative techniques to soft lithography for making microfluidic devices, and for their potential to segue towards scalable commercial fabrication. Here we considered the optimal application of current benchtop 3D printing for microfluidic device fabrication through the lens of lean manufacturing and present a straightforward but robust rapid prototyped moulding system that enables easy estimation of more precise quantities of polydimethylsiloxane (PDMS) required per device to reduce waste and importantly, making devices with better defined depths and volumes for (i) modelling gas exchange and (ii) fabrication consistency as required for quality-controlled production. We demonstrate that this low-cost moulding step can enable a 40 – 300% reduction in the amount of PDMS required for making individual devices compared to the established method of curing approximately 30 grams of PDMS prepolymer overlaid on a 4” silicon wafer master in a standard plastic petri dish. Other process optimisation techniques were also investigated and are recommended as readily implementable changes to current laboratory and foundry-level microfluidic device fabrication protocols for making devices either out of PDMS or other elastomers. Simple calculators are provided as a step towards more streamlined, software controlled and automated design-to-fabrication workflows for both custom and scalable lean manufacturing of microfluidic devices.

Automated cell counting of astrocytes on patterned substrates containing aliphatic and charged properties

1995 ◽  
Vol 53 ◽  
pp. 974-975
Author(s):  
W. Shain ◽  
H. Ancin ◽  
H.C. Craighead ◽  
M. Isaacson ◽  
L. Kam ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Attachment ◽  
Culture Method ◽  
Cell Counting ◽  
Data Sets ◽  
Nuclear Staining ◽  
Double Positive ◽  
A Cell ◽  
Wafer Test ◽  
Cell Densities

Neural protheses have potential to restore nervous system functions lost by trauma or disease. Nanofabrication extends this approach to implants for stimulating and recording from single or small groups of neurons in the spinal cord and brain; however, tissue compatibility is a major limitation to their practical application. We are using a cell culture method for quantitatively measuring cell attachment to surfaces designed for nanofabricated neural prostheses.Silicon wafer test surfaces composed of 50-μm bars separated by aliphatic regions were fabricated using methods similar to a procedure described by Kleinfeld et al. Test surfaces contained either a single or double positive charge/residue. Cyanine dyes (diIC18(3)) stained the background and cell membranes (Fig 1); however, identification of individual cells at higher densities was difficult (Fig 2). Nuclear staining with acriflavine allowed discrimination of individual cells and permitted automated counting of nuclei using 3-D data sets from the confocal microscope (Fig 3). For cell attachment assays, LRM5 5 astroglial cells and astrocytes in primary cell culture were plated at increasing cell densities on test substrates, incubated for 24 hr, fixed, stained, mounted on coverslips, and imaged with a 10x objective.

Comparison of Cell Culture and a Poliovirus Gene Probe Assay for the Detection of Enteroviruses in Environmental Water Samples

1993 ◽  
Vol 27 (3-4) ◽  
pp. 311-314 ◽  
Author(s):  
Aaron B. Margolin ◽  
Charles P. Gerba ◽  
Kenneth J. Richardson ◽  
Jaime E. Naranjo
Keyword(s):  
Cell Culture ◽  
Activated Sludge ◽  
Water Samples ◽  
Cdna Probe ◽  
Culture Method ◽  
Tidal River ◽  
Nucleic Acid Hybridization ◽  
Treated Wastewater ◽  
Gene Probe ◽  
Naturally Occurring

Nucleic acid hybridization provides a rapid non-cell culture method for the detection of enteric viruses in water. The purpose of this work was to compare the detection of naturally occurring enteroviruses by cell culture with their detection by a poliovirus gene probe in various types of water samples. Samples of activated sludge effluent, tertiary treated wastewater (activated sludge, filtration and passage through reverse osmosis), ground water, surface water and tidal river water were processed through 1 MDS Virozorb filters to concentrate any naturally occurring virus. Viruses were eluted from the filters with pH 9.5 beef extract and reduced in volume by flocculation to 20-30 ml. These concentrates were then assayed in the BGM cell line by the cytopathogenic effects (CPE) method and by a poliovirus cDNA probe (base pairs 115-7440) labeled with 32P. A total of 233 samples were assayed in this manner. In slightly more than 93% of the samples gene probe and cell culture yielded the same results. Of these samples 36 were positive by gene probe and 28 by cell culture assay. Positive samples for gene probe were confirmed by treatment with NaOH or RNAse and then reprobed. Samples demonstrating CPE upon primary passage were confirmed positive by subsequent passage of cell lysate on a new monolayer of BGM cells. Ten samples were positive by gene probe and negative by cell culture, and 4 samples were negative by gene probe and positive by cell culture.

scholarly journals Effect of cat litters on feline coronavirus infection of cell culture and cats

2019 ◽  
Vol 22 (4) ◽  
pp. 350-357 ◽  
Author(s):  
Diane Addie ◽  
Lene Houe ◽  
Kirsty Maitland ◽  
Giuseppe Passantino ◽  
Nicola Decaro
Keyword(s):  
Cell Culture ◽  
Virus Infection ◽  
Real Life ◽  
In Vitro Study ◽  
Oral Route ◽  
Virus Shedding ◽  
Virus Load ◽  
Fuller’S Earth ◽  
Feline Coronavirus

Objectives Feline infectious peritonitis (FIP) is caused by infection with feline coronavirus (FCoV). FCoV is incredibly contagious and transmission is via the faecal–oral route. FCoV infection, and therefore FIP, is most common in breeder and rescue catteries, where many cats are kept indoors, using litter trays. Whether it is possible to break the cycle of FCoV infection and reinfection using cat litters has never been investigated. The aim of the study was to examine the effect of cat litters on FCoV infectivity and virus load in multi-cat households, and transmission frequency. Methods Fifteen cat litters were mixed and incubated with FCoV, centrifuged and the supernatants tested in vitro for the ability to prevent virus infection of cell culture. To test applicability of in vitro results to real life, virus load was measured in two households in a double crossover study of four Fuller’s earth-based cat litters by testing rectal swabs using FCoV reverse transcriptase quantitative PCR. Results Four litters abrogated FCoV infection of cell culture, nine reduced it to a greater or lesser extent and two had no effect. One brand had different virus inhibitory properties depending on where it was manufactured. Fuller’s earth-based litters performed best, presumably by adsorbing virus. In the field study, there appeared to be less virus shedding on one Fuller’s earth-based cat litter. Conclusions and relevance The in vitro study successfully identified cat litters that inactivate FCoV; such litters exist so do not need to be developed. Fuller’s earth-based litters best prevented infection of cell culture, but did not completely abrogate FCoV transmission in two multi-cat households. A dust-free clumping Fuller’s earth litter appeared to fare best, but virus shedding also varied on the control litters, complicating interpretation. Sawdust-based cat litters are not useful in FCoV-endemic households because they track badly and have a poor effect on virus infection.

Cat Interferon inhibits Feline Leukaemia Virus Infection in Cell Culture

1972 ◽  
Vol 237 (78) ◽  
pp. 270-271 ◽  
Author(s):  
RICHARD RODGERS ◽  
THOMAS C. MERIGAN ◽  
WILLIAM D. HARDY ◽  
LLOYD J. OLD ◽  
ROBERT KASSEL
Keyword(s):  
Cell Culture ◽  
Virus Infection ◽  
Leukaemia Virus ◽  
Feline Leukaemia Virus

Comparison of a Real-Time PCR Method with a Culture Method for the Detection of Salmonella enterica Serotype Enteritidis in Naturally Contaminated Environmental Samples from Integrated Poultry Houses

2012 ◽  
Vol 75 (4) ◽  
pp. 743-747 ◽  
Author(s):  
BWALYA LUNGU ◽  
W. DOUGLAS WALTMAN ◽  
ROY D. BERGHAUS ◽  
CHARLES L. HOFACRE
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Salmonella Enteritidis ◽  
Culture Method ◽  
Pcr Assay ◽  
Culture Methods ◽  
The Real ◽  
Selective Enrichment ◽  
Conventional Culture ◽  
Field Samples

Conventional culture methods have traditionally been considered the “gold standard” for the isolation and identification of foodborne bacterial pathogens. However, culture methods are labor-intensive and time-consuming. A Salmonella enterica serotype Enteritidis–specific real-time PCR assay that recently received interim approval by the National Poultry Improvement Plan for the detection of Salmonella Enteritidis was evaluated against a culture method that had also received interim National Poultry Improvement Plan approval for the analysis of environmental samples from integrated poultry houses. The method was validated with 422 field samples collected by either the boot sock or drag swab method. The samples were cultured by selective enrichment in tetrathionate broth followed by transfer onto a modified semisolid Rappaport-Vassiliadis medium and then plating onto brilliant green with novobiocin and xylose lysine brilliant Tergitol 4 plates. One-milliliter aliquots of the selective enrichment broths from each sample were collected for DNA extraction by the commercial PrepSEQ nucleic acid extraction assay and analysis by the Salmonella Enteritidis–specific real-time PCR assay. The real-time PCR assay detected no significant differences between the boot sock and drag swab samples. In contrast, the culture method detected a significantly higher number of positive samples from boot socks. The diagnostic sensitivity of the real-time PCR assay for the field samples was significantly higher than that of the culture method. The kappa value obtained was 0.46, indicating moderate agreement between the real-time PCR assay and the culture method. In addition, the real-time PCR method had a turnaround time of 2 days compared with 4 to 8 days for the culture method. The higher sensitivity as well as the reduction in time and labor makes this real-time PCR assay an excellent alternative to conventional culture methods for diagnostic purposes, surveillance, and research studies to improve food safety.

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