TPP tissue culture products help prevent contamination!

Problem: Tissue culture is a vital element of many research laboratories.  Extensive experiments often stem off of cultured cells and provide valuable insight.  Any break in the ability to use cultured cells or newly isolated primary cells impacts many experiments down the line.  Growth of unwanted microbes and mold in tissue culture impairs the ability of a researcher to perform further experiments.  The researcher must then use valuable time and resources to obtain new cells and avoid contamination in the future.  Contamination in tissue culture is a complex and frustrating issue afflicting thousands of researchers every year.   Contamination is not only costly, but results in great productivity loss. 

Solution:  TPP, distributed in the United States by MIDSCI, addresses each contributing issue to contamination with its innovative and unique design. 

               

One of the biggest obstacles to overcoming and avoiding contamination is removing the optimal breeding grounds for microbes and mold spores.  In order for microbes to grow, they need heat, a moist environment, and sustenance.  Incubators provide the perfect breeding ground.  TPP prevents the issues of condensation to build up between dishes, plates, and flasks while stacking.  TPP dishes plates and flasks are manufactured with vented stacking feet (Figure 1).  This not only decreases condensation by even air flow, but also evenly distributes temperature.  Even temperature distribution among multiple samples and treatments is crucial in maintaining reproducibility. 

An additional contributor to microbe and mold growth is the location of the treated surfaces and “deck” seen in some flasks (Figure 2a).  TPP has a flat growth surface directly up to the neck of the flask.  In addition, only the bottom surface of the plates, dishes, and flasks are treated.  When flasks are moved, media sloshes up onto the “angled deck” of some flasks providing an ideal breeding ground for microbes and mold.  The “angled deck” or “dead space” provides yet another disadvantage: In order to add small amounts of liquid to the flask with a pipettor, the shaft must go into the neck of the flask to reach the media (Figure 2b).  Introducing the shaft to the neck of the flask opens the flask for contamination.  TPP flasks, with their lack of dead space, allow for only the sterile pipette tip to enter the flask for small liquid additions .  TPP’s innovative design overcomes the obstacles the “angled deck” presents and prevents yet another contributor to contamination. 

                The vented or filtered cap on a flask is meant to prevent contamination or allow for varying amounts of air to enter.  The caps only provide a barrier to contamination when used and secured properly.  Caps can be easily secured improperly, slightly skewed when turned on the neck of the flask.  Improper cap securing can introduce microbes or mold spores to enter.   The design of TPP filtered caps functionally avoids the issue.  TPP filtered caps have a raised ridge area to help line up to ensure they are closed correctly. TPP vented caps have audible clicks to aid in proper ventilation and the securing of the cap (Figure 3).

               

 

 

       Suspension cells are just as easily affected by contamination as adherent cells.  TPP manufactures a vessel for suspension cells that not only reduces contamination, but saves time and risk of contamination by eliminating the need to transfer to conical tubes before centrifuging .  Gas exchange is often important in the growth of suspension cells and can also be a source of contamination.  TPP bioreactors (Figure 4) have sterile gas exchange through the 0.22µm filter membrane.  In addition, five openings of different size within the cap allow for varying levels of gas permeability, expanding the variety of experim ents that can be performed.   The TPP bioreactor, available in 15, 50, and 600 mLs not only saves the researcher valuable time that would be lost due to contamination, but also time lost in transferring for centrifugation.

 

It is essential that every researcher take extensive steps to ensure their cells and experiments are performed properly and without error.  TPP products help prevent contamination and prevent the costly cleanup and productivity loss. 

Three-dimensional cell culturing by magnetic levitation

N3D hits Nature AGAIN! Contact your MIDSCI representative to try this ground breaking 3D cell culture technology. Recently, biomedical research has moved toward cell culture in three dimensions to better recapitulate native cellular environments. This protocol describes one method for 3D culture, the magnetic levitation method (MLM), in which cells bind with a magnetic nanoparticle assembly overnight to render them magnetic. When resuspended in medium, an external magnetic field levitates and concentrates cells at the air-liquid interface, where they aggregate to form larger 3D cultures. The resulting cultures are dense, can synthesize extracellular matrix (ECM) and can be analyzed similarly to the other culture systems using techniques such as immunohistochemical analysis (IHC), western blotting and other biochemical assays. This protocol details the MLM and other associated techniques (cell culture, imaging and IHC) adapted for the MLM. The MLM requires 45 min of working time over 2 d to create 3D cultures that can be cultured in the long term (>7 d). Read more at: http://www.nature.com/nprot/journal/v8/n10/full/nprot.2013.125.html

Monoclonal Antibody Production – A discussion on the Culturing of Mouse Hybridoma Cells

The production and maintenance of a hybridoma cell begins with the fusion of a specific antibody producing B cell, to a cancer B cell called a myeloma, which does not produce an antibody by itself. Fusion results in an immortalized line called a hybridoma that will faithfully produce a specific antibody against a single epitope called a monoclonal antibody. Once produced, proper maintenance and culturing is required to maximize the performance and continued production of the antibody in question. These were the topics covered in our last Ask the Expert session – Monoclonal antibody production and the culturing of mouse hybridoma cells. Dr. Fawcett is Director of the BioTechnical Institute of Maryland (BTI) a non-profit institute located in Baltimore, Maryland. He is also the Founder and Director of BioSciConcepts, a social venture of BTI that provides hands-on training for professional scientists in cell culture, baculovirus based expression, as well as topics such as molecular biology, PCR and real-time PCR. Dr. Fawcett who has been in biotechnology for over 30 years, provided real hands-on suggestions that readers could benefit from right away. Question topics included: Use of azaguanine Cell viability post cryopreservation Purity of antigen Serum-free culture Problem obtaining hybridomas in spite of strong signal Culture Conditions Rat-mouse hybridomas Read the answers here: http://cellculturedish.com/2013/07/monoclonal-antibody-production-a-discussion-on-the-culturing-of-mouse-hybridoma-cells/

All About CHO – A discussion on everything from the basics to troubleshooting

While CHO cells are a popular choice for biopharmaceutical manufacturing, they can still present a challenge to work with. With so many cell culture scientists regularly searching for ways to improve on product yield, product quality, cost, and a myriad of other manufacturing challenges faced with biopharmaceutical manufacturing, we felt that CHO cell production was a great topic to pursue. Question topics included: Why CHO cells are so popular in biomanufacturing Different types of CHO cells CHO genome sequence Shipping requirements Lactic acid build up in culture Check out the article here: http://cellculturedish.com/2013/06/all-about-cho-a-discussion-on-everything-from-the-basics-to-troubleshooting/

New and Improved Cultureware is Designed with the Scientist in Mind

In the past cultureware has mostly been focused on culturing standard cell lines that have historically been easy to culture. However, with the explosion of stem cell culture and the now common practice of culturing many different cell types, cultureware has needed to evolve as well. See how Techno Plastic Products (TPP) have innovated their products for these changes. http://cellculturedish.com/2013/07/new-and-improved-cultureware-is-designed-with-the-scientist-in-mind/

New Vaccines- Coming Soon to a Doctor's Office Near You...

These new vaccines hold much promise in improving public health and can positively impact many lives.  Check out the new vaccines in this interesting article by The Cell Culture Dish: 

http://cellculturedish.com/2012/07/new-vaccines-coming-soon-to-a-doctors-office-near-you/

Your Questions Answered - "Ask the Expert" on The Cell Culture Dish

Many of the common cell types we use are robust and grow on cell culture treated plastic.  However there are many cell types that have difficulty attaching and/or spreading on cell culture treated plastic.  In other cases, cells may attach but not differentiate unless the attachment surface correct.  To learn more about matrices or if you are using them and have questions, this Ask The Expert topic is your chance to learn more about attachment surfaces and the rationale for using them.

Check out what people are asking:  http://cellculturedish.com

Your Questions Answered - "Ask the Expert" on The Cell Culture Dish

In the past cultureware has mostly been focused on culturing standard cell lines that have historically been easy to culture. However, with the explosion of stem cell culture and the now common practice of culturing many different cell types, cultureware has needed to evolve as well. Over the years, improvements have been developed to increase the success of cultureware and to provide a more hospitable cell-growing environment, including innovations like gas-plasma treatment and biological extracellular matrices. Yet there is still opportunity to improve cultureware and address some of the most common challenges cell culture scientists face.

Check out what people are asking about their cultureware on "Ask the Expert" at The Cell Culture Dish

 

MIDSCI featured in "Ask the Expert" on The Cell Culture Dish

Our own Graziella Mendonsa, PhD is participating in this week's "Ask the Expert" on The Cell Culture Dish. Please visit http://ow.ly/ncxOy to participate!

We'd love to hear from you!

Using the iPhone/iPod to Automate Cytotoxicity Testing with the BiO Ass

Have you ever used your iPhone/iPod with your research? What about with your 3D cell culture work?

N3D Biosciences is launching their BiO Assay to do just that! Check it out!

http://www.n3dbio.com/wp-content/uploads/2013/05/Using-the-iPhone-to-Automate-Cytotoxicity-Testing-with-the-BiO-Assay.pdf

Using the iPhone/iPod to Automate Cytotoxicity Testing with the BiO Assay

Have you thought about using your iPhone/iPod for your research? Look at the new product N3D Biosciences is launching to do just that. Great stuff! http://www.n3dbio.com/wp-content/uploads/2013/05/Using-the-iPhone-to-Automate-Cytotoxicity-Testing-with-the-BiO-Assay.pdf

New and Improved Cultureware is Designed with the Scientist in Mind

Check out our guest blog on The Cell Culture Dish written by our own Graziella Mendonsa, Ph.D.

A Guest Blog by Graziella Mendonsa, Ph.D., Product Manager at MIDSCI

http://cellculturedish.com/2013/07/new-and-improved-cultureware-is-designed-with-the-scientist-in-mind/

Introduction

In the past cultureware has mostly been focused on culturing standard cell lines that have historically been easy to culture. However, with the explosion of stem cell culture and the now common practice of culturing many different cell types, cultureware has needed to evolve as well. Over the years, improvements have been developed to increase the success of cultureware and to provide a more hospitable cell-growing environment, including innovations like gas-plasma treatment and biological extracellular matrices. Yet there is still opportunity to improve cultureware and address some of the most common challenges cell culture scientists face. Techno Plastic Products (TPP) is a Premium Swiss manufactured line of cell cultureware that is designed by scientists for scientists. Virgin plastics of highest purity and quality are poured with perfection into uniquely engineered molds to deliver the perfect flask, dish or plate for adherent cells to attach to. Suspension cells also get an updated housing with a conical Bioreactor that significantly enhances distribution of gas exchange and nutrition. Quality control is the number one priority for TPP, which includes the testing of each and every piece manufactured at their site.

Cultureware Challenges

Maintaining Standard Temperatures

Cell culture in general is treated as a great in vitro model system to assay signal transduction, cellular cross talk and responses to externally applied gene activators or inhibitors. Since cells in culture are sensitive to environmental stimuli and react accordingly for adaptation, it is essential to maintain their necessary vitals such as temperature and CO2 stable and non-fluctuating. Almost every lab with multiple scientists performing cell culture usually stacks their flasks, dishes and plates to save on space in their incubator. However, a key question to ask is “Are the vessels in the middle of the stack the same temperature as that on the top or bottom of the stack?”. The answer is a resounding “Probably Not!.” Therefore TPP has engineered a raised growth surface on their flasks, dishes and plates to avoid insulation of the adjoining vessels and has designed the vessels to have a stable footing for enhanced security in stacking. Maintaining Optimal Nutrition, Gas Exchange and Gene Expression The foundation of cell culture at the time of seeding in a flask or dish, undoubtedly sets up for consequent cellular viability and propagation during passaging and for consistent downstream gene reporting assays. Therefore how cells are initially thawed from frozen and plated during the first 2-3 days has a dramatic impact on cellular behavior downstream. Therefore controlling the flatness of the surface upon which cells adhere to reigns important to obtain consistent cell monolayers. Cells that are growing in layers are not only deprived of nutrition and the optimal gas exchange, but are also now transcribing uneven gene expression. TPP manufacturers one of the flattest growth surfaces in the market.

Evaporation of Media

Evaporation of media in 96-well plates appears to be one of the most common complaints from cell culture scientists. This is due to the overheating of the outer wells and a consequent temperature gradient across the well plate and across the stack of well plates in the incubator. This uneven heating can also lead to condensation build up which are breeding grounds for bacteria. The temporary solution generically recommended is to fill the outer wells with media alone or PBS. However the wastage of unused wells for cell growth and experimentation amounts to almost 40%. TPP plates eliminate this concern with its raised growth surface for optimal stackability and consequent even heating from all sides. Ultimately, a 96-well plate is used for multiple replicates of control and experimental treatments and with TPP plates, one can achieve that goal because surrounding environmental conditions such as temperature are no longer a contributor to variation.

Eliminating Dead Space

Focusing in on the flasks, it is strikingly obvious that TPP flasks are shorter in length. That was purposely designed to encompass the entire bottom of the flask as the treated growth surface. Therefore, with no wasted “dead space” in flask, the cell to media ratio is now optimally maximized. All TPP flasks come with an angled neck; this small design tweak serves as the best method to eliminate media accumulation in the neck and thus the consequent high risk of contamination. TPP also has filtered caps for their flasks that are designed to be hydrophobic in nature, so that incase media does slosh against the cap when the flask is either laid at the microscope or put back in the incubator, the risk of contamination is minimized greatly. The angled neck also provides full access to the internal corners of the flask for a serological pipette or scraper during trypsinization or cell harvestation respectively. Summary TPP dishes have ingenuity spelled all over it. From the unique grip ring on the edge of the dish that prevents accidental dropping to the internal clock orientation for visual quadrants during microscopy, TPP continues to invent upgrades to the cell culture dish that take cell cultureware from standard to ergonomic. TPP cultureware, designed by scientists for scientist, provide you with unique and innovative upgrades that translate to more consistent seeding, handling, treatment and harvestation of cells. Peace of mind included.

And the JUNE BACKPACK WINNER IS...

Our JUNE BACKPACK WINNER is... Benjamin Goldschmidt from University of MO - Columbia!! Congrats!! Your rep, Nick B, will personally deliver it to you. Thanks everyone for participating! For your chance to win, fill out our survey at http://ow.ly/mxIF8. We draw winners every month. :)

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Researchers at KU Medical Center identify a new target for treating pain

June 14, 2013

By David Martin

A hormone that has been used to control blood pressure may hold the key to a new approach for treating pain, according to research at the University of Kansas Medical Center. The hormone, angiotensin II, constricts blood vessels and releases a substance that causes the body to retain salt and water. KU neuroscientist Peter Smith, Ph.D., and his colleagues have found that blocking angiotensin II receptors prevents the increase in sensitivity that normally accompanies inflammation. The mechanics of the angiotensin II system's ability to regulate pain pathways was not appreciated before, Smith says.

The study has been published online in The Journal of Pain. 

The finding is based on a previous study of estrogen that Smith, professor of molecular and integrative physiology and director of the Institute for Neurological Discoveries at KU, published in 2008. "We know that the hormone estrogen makes women more prone to many painful conditions, and we found that estrogen actually caused some pain-sensing nerve cells to grow," he says, adding that many chronically painful conditions are accompanied by an increase in pain-sensing nerves.

In looking at what genes were affected by estrogen, Smith and his colleagues were surprised to learn that nerve cells started making more of a protein that allows cells to respond to angiotensin II. This led the researchers to speculate that the protein, angiotensin II receptor type 2, or AT2, might be important to pain.

Tests on rodents confirmed the relationship. In the experiment, rats' hind paws were injected with an irritant. Exposure to touch and heat revealed the paws' sensitivity.

Some of the rats received a chemical known to inhibit AT2. In these rats, angiotensin II did not cause pain-sensing nerves to sprout, and paw sensitivity was the same as in rats that did not receive the irritant. In short, the AT2 blocker uncoupled the link between inflammation and the nerve response.

"It appears that AT2 is critical for the sensations of pain, and that AT2 receptor inhibitors are about as effective as morphine - but without all of the dangerous side effects," says senior scientist Anuradha Chakrabarty, Ph.D., the paper's first author. Zhaohui Liao, M.D., also contributed to the study.

The study, Smith says, confirms that the proliferation of pain-sensing nerves associated with chronic pain syndromes is meaningful. "More sprouting means more pain," he says.

The study is the first to shed light on the mechanism of overgrowth of pain-sensing nerves, Smith says. Inhibiting AT2, he adds, "represents one of the few drug therapies that actually is known to be directed at the underlying biological mechanisms that may be responsible for some aspects of chronic pain."

Chronic pain has been estimated to cost the U.S. between $560 and $635 billion in health care costs and lost productivity. Nonsteroidal anti-inflammatory drugs may not work for all patients and have the potential to damage the tissue of the gastrointestinal tract, kidney and liver. Opiates pose the risk of addiction and overdose and are unsuitable for long-term treatment of chronic pain.

An Australian pharmaceutical company is currently testing an AT2 receptor blocker in a clinical trial with patients with postherpetic neuralgia, nerve pain caused by shingles.

The KU study was supported by the National Institutes of Health grant RO1HD049615.

Article found at http://ow.ly/mcdSL

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