Formation of Mesenchymal Tissues in Alvetex® Scaffold

The Alvetex®Scaffold is a novel substrate that enables a solution for simple and routine 3D culture. It is composed of a highly porous polystyrene scaffold that has been engineered into a 200 micron thick membrane to enable entry of cells and efficient exchange of gases and solutes. Cells enter the fabric of the scaffold, retain their natural 3D structure, and form close 3D interactions with adjacent cells. Unlike conventional 2D culture, cells in Alvetex®Scaffold do not grow as monolayers and do not undergo the flattened shape transition that can result in aberrant changes to gene and protein expression and consequently cellular function.

Mesenchymal stem cells (MSCs) are adherent multipotent cells derived from tissue such as bone marrow and which possess the ability to differentiate in vitro into a number of tissue types including bone, cartilage and muscle [1].

In this post, we demonstrate that MSCs extracted from the bone marrow of adult rats can be successfully cultured in 3D in Alvetex®Scaffold and induced to differentiate into osteogenic and adipogenic derivatives more efficiently than their 2D counterparts. We also report bone formation and the production of extracellular matrix by MG63 cells which represent an established cell line derived from a human osteosarcoma. The data generated here is supported by peer-reviewed literature [2,3] and clearly shows that Alvetex®Scaffold promotes enhanced in vitro differentiation. [Read more…]

PBMCs & drug development

Use of human samples including peripheral blood mononuclear cells (PBMCs) in drug discovery is critical for increasing the chances of success for a small molecule screen hit. A review based on recent published data!

PBMCs DD 1Several theories try to explain why many drug discovery and development projects have a moderate success rate. Even with the advances made in methods (HTS, HCA, library managment, etc), there are many hurdles for a small molecule on the long road to FDA or EMA approval.

One way that may improve predicted efficacy and toxicity of drug leads is to use human samples, such as blood, early on in drug discovery programs. Peripheral blood mononuclear cells  (PBMCs) can be easily used in a variety of ways during the drug discovery process to gain a better understanding of the effects of a small molecule. [Read more…]

Case study: 3D Cancer Cell Cytotoxicity in vitro Assessment

Nowadays in Cancer research, there is a strong need for three-dimensional (3D) in vitro experimental approaches that mimick as much as possible the in vivo tumour ecosystem. In addition to solving issues related to animal models, these in vitro 3D models allow precise tuning of experimental design when investigating cancer cell biology but also, compound screening and disease modelling. In this post, we’ll take a look at the growth of the popular breast cancer cell line, MCF-7, using Alvetex®Scaffold technology (Reinnervate – a ReproCell company) to create a 3D structure that closely resembles the structure of tumour tissues grown in vivo.

[Read more…]

Cell counting made easy!

After you’ve read this post, I hope that you’ll agree that what the title of the post announces is the right assumption. How? Read on!

I invite you to take a look at Cyto-X, which is a ready-for-use cell viability reagent for proliferation and cytotoxicity assays. When added to cell culture, a highly water-soluble tetrazolium salt is reduced by dehydrogenases in live cells to form an orange colored formazan product that is soluble in culture medium. The amount of formazan is directly proportional to the enzymatic activity of dehydrogenases and the number of living cells.

With its high sensitivity and low cytotoxicity, Cyto-X outperforms traditional assays using other tetrazolium salts such as MTT, XTT, MTS or WST-1.

Here are 3 good reasons to try Cyto-X Colorimetric cell counting reagent…

1. It’s more sensitive

 

Cyto-X 1

 

Left: Human Dermal Fibroblasts (HDF) were seeded at different concentrations. Cells were counted the next day using the Cyto-X (blue) and MTT (orange) assays.
Right: Bovine Aortic Endothelial Cells (BAOEC) were treated with staurosporine for 16 hours. Cell number was determined by Cyto-X and MTT assays.

 

2. It’s less toxic

Cyto-X 2

Left: Cyto-X was added to Human Umbilical Vein Endothelial Cells (HUVEC) for 24 hours. Following the Cyto-X cell counting assay, HUVEC are still proliferating and capable of uptaking DiL-Ac-LDL, demonstrating HUVEC are metabolically active, and that downstream applications are possible.
Right: By contrast, toxic formazan is formed after the MTT assay and HUVEC are dead.

 

3. It’s user friendly: fewer steps & less waiting time

Cyto-X 3

So, doesn’t this make cell counting easy? I could also claim “Cell counting made easy and inexpensive!”. Not only is this protocol quicker, it also uses less reagents vs. MTT (no phenol-red medium, no extraction solution, no wash solution, etc) and is as cheap as 0.36 € per test point.

Convinced? Leave your comments or get in touch below!

The easy way to find your bioactive small molecules for stem cell biology

In addition to a plethora of cytokines and growth factors which can be used in stem cell biology to stimulate differentiation, proliferation, and maintenance of different cell types, there are a number of small molecules which are known to act in similar ways on stem cells. The crucial point, is that these molecules have to be produced and formulated in a way which keeps them bio-active. [Read more…]

Tips for primary cell culture

Functionality and viability of primary cells can be impaired by incorrect thawing procedures, storage or culture conditions. It’s generally admitted that applying the same protocols as for cell lines leads will lead to bad cell quality. Well, based on our experience with primary cells, here are a few tips you can follow to ensure you get the best performance.

[Read more…]

Characterization & Applications of Human Cardiomyocytes

ReproCardio 2 Human iPSC-derived cardiomyocytes have been extensively characterized for their functional responsiveness to the known cardiotoxic compounds. In all our tests, including assays of cardiotoxic compounds not correctly identified by the hERG assay, ReproCardio has generated results in complete agreement with clinical findings.

In this post, we’ll highlight the characterisation of ReproCardio 2 cardiomyocytes and illustrate the applications they are used for. [Read more…]

Mouse Embryonic Fibroblasts Troubleshooting Guide

Mouse Embryonic Fibroblasts (MEFs) are critical biological reagents in stem cell research. Used as “feeder cells” to maintain Embryonic Stem (ES) cells and induced Pluripotent Stem (iPS), the quality of MEFs strongly influences Stem cell culture conditions, and may react differently depending on culture conditions. In parallel, MEFs can also be reprogrammed into a variety of more mature differentiated functional cell types (cardiomyocytes, neurons…). During daily discussions with Stem cell biologists, I have been able to collect Frequently Asked Questions and technical tips related to MEFs.

[Read more…]

5 most popular posts in Stem Cells in 2014!

Following on from the 5 most popular posts in Cell Sourcing, today let’s look back at the 5 most visited blog posts in 2014 in the field of stem cells.

images#1 – Induced pluripotent neural stem cells in screening assay

Introducing Human Induced Pluripotent Stem Cells as a promising option for researchers creating in vitro models for the study of neurodegenerative diseases. We also highlight the benefits of using human iPS-derived neural cells vs. rodent primary neural cells.

 

4-Maintains Normal Karyotype after 20 Passages#2 – Cost-effective serum replacer for stem cell culture!

FBS is an inherently variable component of the cell culture system, so consistent performance demands extensive screening of manufactured lots. To circumvent this problem, companies began offering “stem cell qualified” serum… discover more in this post!

 

Comparison of standard reprogramming methodologies#3 – mRNA reprogramming system: fastest & safest!

Early reprogramming methods utilized retroviral (including lentiviral) vectors to introduce reprogramming-associated transcription factors to the target cells. While these pioneering experiments provided a fundamental proof-of-concept, these vectors required the integration of heritable viral DNA sequence into the host cell’s genome… Discover the benefits of a new mRNA reprogramming system!

 

EGF StemBeads vs soluble EGF in IF staining of E14 and E18 Cortical cells tebu-bio

#4 – Recent advances in Stem Cell culture: Encapsulated Stembeads growth factors

How can I optimise my media to cultivate Stem cells with stable and cost-effective Stem Cell Factors? Find all the answers in this post!

 

c0b67d286c-DopaCell_Spec_Picture3

#5 -Isogenic Knockout iPSC Lines for Modeling of CNS Disorders

Knockout iPSC lines that mimic loss of function of genes associated with various CNS disorders as novel tools for controlled experiments to decipher functionality of each disease‐associated gene, and thus the mechanism underlying the disease.

 

We hope you found these posts of interest, stay up to date in 2015 by following our future articles!

5 most popular Cell Sourcing blog posts in 2014!

Today, I’d like to invite you to take a look at the 5 blogs that saw the most visits on our blog in 2014, in the field of Cell Sourcing & Cell Culture.

blood cells#1 – 6 good reasons (in short!) to use our blood derived cells!

At the end of 2014, we signed a distribution agreement with HemaCare, a world-leader of blood-derived cells. We took a look at just a few of the many benefits for European researchers.

 

FribrOut - prevent overgrowth of fibroblastic and contaminated cells

#2 – How to get rid of fibroblastic and contaminating cells?

Very often primary cell cultures are jeopardized by the outgrowth of contaminating fibroblasts.  The population of the target primary cells is directly affected, since fibroblasts usually grow at much faster rates than other cell types. So what are the solutions? Just read this blog!

 

tebu-bio primary cell sourcing Human Primary Sebocytes Zenbio#3 – Primary cell sourcing: Cryopreserved Human Adult Sebocytes

Sebocytes are emerging cellular models to study skin biology but also to design new compounds to regulate acne, seborrhoea and other sebaceous gland-related diseases. Easy access to cryopreserved and well-qualified sources of Human Adult Sebocytes will boost skin research and compound testing projects.

 

Cardiomyocytes grown on the surface of HyStem with fibronectin#4 – Tips for Optimizing Cell Culture Conditions with Hydrogels

In order to engineer a custom culture system with HyStem, simple pilot studies or optimization assays can be tested to determine the appropriate parameters for each experiment. Tips and guidelines for optimizing basic variables that can have a significant impact on the hydrogel properties and overall cell culture conditions are covered in this blog.

 

OptiTDS - OptiTDS™, is a mixture of tissue dissociation enzymes/reagents #5 – Struggling with tissue dissociation?

Successful culture of primary cells requires obtaining a maximum yield of functionally viable single cells from whole tissues… but this is easier to say than to achieve! Here are some solutions!

Stay tuned in 2015 for more topics, not only in the field of Cell Sourcing and Cell Culture, but many others!