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RALEIGH, N.C. -- Startup TechWire (www.startuptechwire.com) has launched to champion entrepreneurs and innovation.

 

The professional news outlet reports on business, innovation, and education for America's vibrant startup community, especially the life sciences.

 

“The number of entrepreneurs across the U.S. is constantly growing, but not everybody is hearing about all the new tech that’s out there. It’s exciting to help build a stronger innovation community by sharing news and information on these startups and the businesses that support them,” David Menzies, editor/publisher, said.

 

Nearly a fifth of working adults in the U.S. – approximately 27 million people – identify as entrepreneurs. Many of these are solopreneurs, running a business alone without employees in order to stay lean and nimble to adapt to change, although this is somewhat limiting to scalability.

 

Many of these solopreneurs, Menzies explained, do not have the resources to get their stories out.

 

“That’s one of the ways Startup TechWire can help, by showcasing their products and ideas as well as covering trends related to their technologies,” he said.

 

Larger entrepreneurial trends in 2016 include innovation in utility apps geared toward “real life” issues such as travel, health and fitness, music, and news; bots and artificial intelligence; and products that promote productivity.

 

Menzies originally began publishing the tech news website as a means to generate extra visibility and web traffic for clients of his PR consultancy, Innovative Public Relations. Peers in the PR field began asking him if they could send their clients’ news items, and the publication grew, with Menzies utilizing his years of experience as a print newspaper editor to cull submissions and put forth a professional publication.

 

Word of mouth generated interest beyond the publication’s initial focus on North and South Carolina, with Startup TechWire now covering the entire U.S.

 

Fresh, original articles and user-submitted news items are updated daily, complimented by feeds from respected media sources.

 

Startup TechWire is an extraordinarily powerful and effective means of reaching a great number of people locally, nationally and globally.

 

For more information on editorial and advertising opportunities, visit www.startuptechwire.com or email This e-mail address is being protected from spambots. You need JavaScript enabled to view it .

 

About Startup TechWire

Startup TechWire (www.startuptechwire.com) reports on business, innovation, and education for America's vibrant startup community. It is a professionally edited online news outlet providing readers with timely information about the life sciences, entrepreneurism, high tech and education. Fresh, original articles and user-submitted news items are updated daily, complimented by feeds from respected media sources. Startup TechWire is published by Innovative Public Relations, Inc., a publicity and branding consultancy helping clients achieve their business development and organizational goals. All content is edited by Editor/Publisher David Menzies 919-274-6862 (www.daviddeanmenzies.com) an award-winning 22-year professional communicator, published author and former print newspaper editor with a rich technology background.

 

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Lumiprobe.com Lumiprobe fluorescent reagents

Inside this newsletter
  • Useful for Manufacturing Oligonucleotide Therapeutics
  • Citations of Interest
  • Scientist's Questions - Increase your knowledge and ideas!
  • Lumiprobe New Years greeting!


Lumiprobe reagents reduce cost, increase productivity and improve quality of research and production. Thanks to all ! Save 5% use discount code: biocrowd

Lumiprobe offers over 80 reagents for peptide, oligonucleotide, amine, alkyne binding - useful for Manufacturing Oligonucleotide Therapeutics!

In 2014, whether researching, creating, or manufacturing new drugs and treatment, Lumiprobe reagents were giving results in drug development research. Click chemistry was used with peptide-based nanoparticles in vivo, comparing the advantages between linear and cyclic peptides for intracellular delivery, process improvements for the economic  production,  peptide characterization , detecting and controlling oligonucleotide impurities, and exploring the development of peptides for diagnostic applications.  Novel oligonucleotide and peptide therapies were also enhanced when Lumiprobe's reagents were included.

Dye NHS esters - amine-reactive cyanine activated esters for the labeling of proteins, peptides, and other molecules

Sulfo NHS esters - water soluble sulfo-Cyanine3 SE activated ester for amino-biomolecule labeling.

Azides - fluorescent biomolecule labeling through Click Chemistry

Alkynes -
alkyne dye,  Useful for both copper-catalyzed, and copper-free Click Chemistry.

Maleimides -
bright and photostable thiol-reactive dye for protein labeling

Amines - fluorophores with free amino group (amino-dye). It can be conjugated with NHS esters, alkynes, carboxy groups (after activation), and epoxides.

Hydrazides -
dyes with a reactivity for carbonyl groups (aldehydes and ketones) activated by acid.  It can be used for the labeling of glycoproteins (i.e. antibodies) after periodate oxidation of sugar moieties.

Citations of Interest !

Would you like your paper featured on Lumiprobe citation webpage?
Over 100 papers sorted by date or reagent are available for your review.
Email: This e-mail address is being protected from spambots. You need JavaScript enabled to view it

In Site-Selective Protein Immobilization by Covalent Modification of GST Fusion Proteins.
Zhou, Y.; Guo, T.; Tang, G.; Wu, H.; Wong, N.-K.; Pan, Z.
Bioconjugate Chemistry, 2014 doi: 10.1021/ bc50 0347b



The immobilization of functional proteins onto solid supports using affinity tags is an attractive approach in recent development of protein microarray technologies. Among the commonly used fusion protein tags, glutathione S-transferase (GST) proteins have been indispensable tools for protein–protein interaction studies and have extensive applications in recombinant protein purification and reversible protein immobilization. Here, by utilizing pyrimidine-based small-molecule probes with a sulfonyl fluoride reactive group, we report a novel and general approach for site-selective immobilization of Schistosoma japonicum GST (sjGST) fusion proteins through irreversible and specific covalent modification of the tyrosine-111 residue of the sjGST tag. As demonstrated by sjGST-tagged eGFP and sjGST-tagged kinase activity assays, this immobilization approach offers the advantages of high immobilization efficiency and excellent retention of protein structure and activity.

EGF receptor-targeting peptide conjugate incorporating a near-IR fluorescent dye and a novel 1,4,7-triazacyclononane-based 64Cu(II) chelator assembled via click chemistry.
Viehweger, K.; Barbaro, L.; Garcia, KP; Joshi, T.; Geipel, G.; Steinbach, J.; Stephan, H.; Spiccia, L.; Graham, B. Bioconjugate Chem., 2014, 25(5), 1011–1022. doi: 10.1021/ bc50 01388









A new Boc-protected 1,4,7-triazacyclononane (TACN)-based pro-chelator compound featuring a “clickable” azidomethylpyridine pendant has been developed as a building block for the construction of multimodal imaging agents. Conjugation to a model alkyne (propargyl alcohol), followed by deprotection, generates a pentadentate ligand, as confirmed by X-ray crystallographic analysis of the corresponding distorted square-pyramidal Cu(II) complex. The ligand exhibits rapid 64Cu(II)-binding kinetics (>95% radiochemical yield in <5 min) and a high resistance to demetalation. It may thus prove suitable for use in 64Cu(II)-based in vivo positron emission tomography (PET). The new chelating building block has been applied to the construction of a bimodal (PET/fluorescence) peptide-based imaging probe targeting the epidermal growth factor (EGF) receptor, which is highly overexpressed on the surface of several types of cancer cells. The probe consists of a hexapeptide sequence, Leu-Ala-Arg-Leu-Leu-Thr (designated “D4”), followed by a Cys-β-Ala-β-Ala spacer, then a β-homopropargylglycine residue with the TACN-based chelator “clicked” to its side chain. A sulfonated near-infrared (NIR) fluorescent cyanine dye (sulfo-Cy5) was introduced at the N-terminus to study the EGF receptor-binding ability of the probe by laser-fluorescence spectroscopy. Binding was also confirmed by coimmunoprecipitation methods, and an apparent dissociation constant (Kd) of ca. 10 nM was determined from radioactivity-based measurements of probe binding to two EGF receptor-expressing cell lines (FaDu and A431). The probe is shown to be a biased or partial allosteric agonist of the EGF receptor, inducing phosphorylation of Thr669 and Tyr992, but not the Tyr845, Tyr998, Tyr1045, Tyr1068, or Tyr1148 residues of the receptor, in the absence of the orthosteric EGF ligand. Additionally, the probe was found to suppress the EGF-stimulated autophosphorylation of these latter residues, indicating that it is also a noncompetitive antagonist.

Size- matched alkyne-conjugated cyanine fluorophores to identify differences in protein glycosylation.
Burnham- Marusich, A.R.; Plechaty, A.M.; Berninsone, P.M.
ELECTROPHORESIS, 2014, 35, 2621–2625. doi: 10.1002/elps.201400241

Currently, there are few methods to detect differences in posttranslational modifications (PTMs) in a specific manner from complex mixtures. Thus, we developed an approach that combines the sensitivity and specificity of click chemistry with the resolution capabilities of 2D-DIGE. In “Click-DIGE”, posttranslationally modified proteins are metabolically labeled with azido-substrate analogs, then size- and charge-matched alkyne-Cy3 or alkyne-Cy5 dyes are covalently attached to the azide of the PTM by click chemistry. The fluorescently-tagged protein samples are then multiplexed for 2DE analysis. Whereas standard DIGE labels all proteins, Click-DIGE focuses the analysis of protein differences to a targeted subset of posttranslationally modified proteins within a complex sample (i.e. specific labeling and analysis of azido glycoproteins within a cell lysate). Our data indicate that (i) Click-DIGE specifically labels azido proteins, (ii) the resulting Cy-protein conjugates are spectrally distinct, and (iii) the conjugates are size- and charge-matched at the level of 2DE. We demonstrate the utility of this approach by detecting multiple differentially expressed glycoproteins between a mutant cell line defective in UDP-galactose transport and the parental cell line. We anticipate that the diversity of azido substrates already available will enable Click-DIGE to be compatible with analysis of a wide range of PTMs.

 

Tech Support: Lumiprobe offers FREE tech support before or after you order.
Questions - Increase your knowledge and ideas!

I have few questions about the click-chemistry labeling and need suggestions. In the protocol for Click-Chemistry Labeling of Oligonucleotides and DNA,
1.  Triethylammonium acetate buffer (final concentration 0.2 M, pH 7.0) was used. Does it matter that types of salts and pH of buffer effect the labeling efficiency? Can PB buffer be used?
2.  Azide was dissolved in DMSO and 1.5 * (DNA concentration), is it OK that azide was dissolved in water? In my experiment, azide was connected to peptide and had a good water solubility. Is it 50% DMSO necessary for the performance of click-chemistry labeling?
3.  Degass steps were performed during the procedure. Does it sensitive to oxygen?
4.  Precipitating the conjugate with acetone or ethanol was preformed to remove excess small molecules. Can Amicon ultra 3K device (Millipore) for buffer changing get the same result?
5.  Can reaction yield get >90%? Will +4 Centigrade lower the yield?

Thank you for your email!

1. In practice, Click Chemisty displays little dependence on the composition of buffer. It should not contain free thiols (i.e. no DTT or mercaptoethanol). The reaction can be carried out at pH of at least 4 to 10. I believe phosphate will work fine.
2. You can dissolve your azide in water without any issues. The protocol uses DMSO because it is optimized for non water-soluble azides.
3. Yes, the reaction is sensitive to oxygen. Alkyne dimers can form upon oxidation. Moreover, hydroxyl radicals are formed upon oxidation of Cu(I) catalyst, and they can damage biomolecules, especially DNA/RNA. We recommend to degass the solution.
4. Any purification will work, precipitation is the easiest for oligo labeling. You can desalt the reaction.
5. Yes, yields may exceed 90%. Lowering temperature to +4 Centigrade will not harm, but probably will extend the required conjugation time.

Please do not hesitate to contact Lumiprobe if you have more questions.

Actually, I do have one more question about NHS-ester labeling. The optimal pH value for modification is 8.3-8.5. In most cases I saw, the pH value suggested is 7.4. For example, SDA (pierce, product 26167) I used for proteins labeling is used in PBS. Is there any difference between DNA labeling and protein labeling? The pKa value of primary amines is the key factor?

Optimal pH for NHS ester labeling is indeed 8.3-8.5. pH value of 7.4 can be used if you need to label preferably N-terminus of a protein rather than side chains of lysine. The terminal amino-group is less basic than lysine amine, and it is less protonated at this pH than that of lysine. Therefore, it reacts preferably with the NHS ester. At higher pH ranges, both amino groups are non-protonated, and since lysine amines are more reactive, they react first.

If you need to label protein lysines or aminolink-DNA, use pH 8.3-8.5.

Do you provide the service which conjugate peptide to cy5 NHS ester?

Unfortunately, Lumiprobe does not make labeled peptides and does not perform conjugation. However, Lumiprobe can supply you or service company of your choice with our fluorescent reagents. The labeling protocol is simple enough to be performed by you. If you have peptide purification facilities such as HPLC -  if you work with peptides, you likely have it). If you prefer labeling as a service, there are local companies who do it - Lumiprobe can supply our reagent to this company. This will help save money on reagents.

Why use Lumiprobe's fluorescent dyes such as Cyanine5 NHS ester?

http://www.lumiprobe.com/p/bodipy-fl-nhs-ester

Cyanine5 fluorophore emission has maximum in red region, where many CCD detectors have maximum sensitivity, and biological objects have low background. Dye color is very intense, therefore quantity as small as 1 nanomol can be detected in gel electrophoresis by naked eye.

* high quality,  ideal for very cost-efficient labeling
* works well in organic solvents for small molecule labeling of soluble proteins,
peptides and oligonucleotides
* Water-soluble sulfo-Cyanine 5 NHS ester for sophisticated cellular & protein targets
* Compatible with various instrumentation including fluorescent microscopes,
imagers, scanners, and fluorescence readers.

 

Send your tech questions ? Any comments on the newsletter? What information would you like in the next issue?

Lumiprobe.com

Lumiprobe New Year's greeting!
Restock your lab in January and save 5%.
Use discount code: biocrowd

Thank you for integrating Lumiprobe fluorescent probes into your work in 2014!
Restock your lab or begin a new project - Lumiprobe offers you a new year's email discount of 5%. Use discount code: biocrowd

If you need something not found in Lumiprobe's catalog - ask!

Lumiprobe offers to help with your research, and create a custom probe for you at no extra charge! Reagents can often be customized for your needs at price schedule similar to Lumiprobe's other reagents. Ask Lumiprobe's tech support - you'll receive attention to your research, and ideas on what or how to do the click chemistry reaction. Lumiprobe's website offers instantly downloadable Certificate of Analysis (CoA) detailing meticulous quality control, real NMR, UV and mass spectra, and HPLC chromatograms.

Lumiprobe |    Products/Technical Info |    Protocols |    Ordering |    Contact
Lumiprobe.com

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Single-use technologies have revolutionised the biopharmaceutical industries in more ways than you might think. It is indeed most common nowadays to see single use applications cascading along the many upstream and downstream processes which channel through the sector.

But what about the people who work tirelessly along the bioprocess supply chain? Well, it would seem Starbucks Japan has woken up and smelt the coffee when it comes to single use. The company’s outlets across East Asia are keeping  biopharmaceutical workers - along with the public - topped up on coffee which is brewed with a twist.

The art of great coffee


Starbucks Origami Personal Drip coffee is the first Starbucks single use drip coffee product offered in the world. Pick from one of three roasts, open the sealed bag, fan out the origami filter, affix to a mug and pour hot water over the ground beans.

Then sit back and enjoy the perfect blend.

Personal Drip, so says Starbucks, invokes the “artistry and hand-crafted nature of Japan’s ancient art of origami and this innovative product allows customers to brew a single cup of Starbucks coffee at home that does not require any special equipment”.

We’d have to agree. After all, at ALLpaQ we like to think of ourselves as coffee connoisseurs. Even so, this is by far the most creative way we’ve seen of brewing coffee.

Right then, time to put the kettle on. Anybody want one?

 

Related reading:

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b2ap3_thumbnail_PD-PCR-GIF.gifb2ap3_thumbnail_PD-PCR-GIF.gif

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As a young man, mother would always tell me to “clean your room, son”. Fastforward a decade – or two or three – and the occasional pot or pan still lurks under the bed. Mother, I apologise.

 

When it comes to manufacturing and scientific research though, cleanliness is a must.

Keeping your cleanroom “clean” is imperative – the nostalgic teenage kick of sweeping things under the carpet has no place here.

Your cleanroom, after all, must only be occupied by a controlled level of contamination.

Ensuring any cleanroom facility is equipped with the right tools is therefore a must.

Bioprocessing cleanroom particle containment

The ALLpaQ Cleanroom 1000 bioprocess container is a bit of a clean freak. Obsessively compulsive about keeping unwanted particles at bay, the container delivers greater hygiene, cost effectiveness, functionality and versatility day in, day out.

Unlike steel equivalents, the ALLpaQ Cleanroom range is:

Got a question about cleaning your room?

ALLpaQ Cleanroom bioprocess containers have been developed for specific use within a cleanroom environment.


Find out more

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If you’re wondering whether plastic bioprocess containment is right for you, flick through the bioprocessing guide which explains how the ALLpaQ solution enhances the sustainability and profitability of your business.

It's a real page turner


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Hi there. Thanks for checking out my dating profile. I’m an ALLpaQ plastic bioprocess container. What am I looking for?

Well, the answer is pretty easy. I’m searching for that special someone to share my days with.

About my perfect match

I’m a real glutton for single use bags, especially those whose share my passion for protecting precious biopharmaceutical media.

As a bioprocess container, I’m often away on business, travelling via road, sea and air. A life and travel companion is therefore a must. Love is, after all, a journey which starts at forever and ends at never.

Of course, I like the little things in life as well. So if you’re the type who enjoys cuddling up in cold room and warehouse environments, drop me a line.

Let’s do it: let’s fall in love

Set up a first date to find out more about how ALLpaQ bioprocess containers can optimise your pharmaceutical storage and shipment supply chain.

Tell me more

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Growing up, Willy Wonka’s chocolate factory was the place I wanted to be. Sure, the guy had eaten too much sugar. But the lure of this mad inventor’s edible landscapes more than made up for the fact that he was a little ramped up.

From within the sugary subtext of Roald Dahl’s masterpiece an interesting subtext can be pulled – many of his delights were package-free. Like a ripe tomato, Wonka products could be plucked then directly consumed.

This was somewhat forward thinking of Dahl, portending to today’s food packaging zeitgeist. Some manufacturers, for instance, believe they have discovered the golden ticket to making package-free products a reality in our supermarkets.

But would you purchase a product contained in edible packing? There’s a number of innovative development companies out there who believe, eventually, you will. They say the answer to reducing excess food packaging resides within how nature creates its own biodegradable packaging.

Let’s take the bite sized snack protectors – they are all the talk right now. These products aim to redefine the way we package food with durable, water-resistant edible membranes made from natural food particles. The portion of food is safeguarded within its casing much like the skins of fruits.

Interesting, right?

Perhaps what is more interesting is how the debate taps into the need for sustainable packaging across all industries. In the pharmaceutical sector where we work, for example, sustainability is at the forefront of our packaging design.

ALLpaQ was, in fact, born out of a desire to rethink the traditional packaging model pharma and life science companies had become accustomed to when transporting valuable fluid media.

Packaging sustainability

Bioprocess containment isn’t going away anytime soon.This is a reality. It remains a part of daily life for most businesses and organisations. Even so, we all have a duty to store and transport media responsibly.

At ALLpaQ then, rather than aiming to eradicate containment packaging, we fabricate designs which streamline their operational capabilities. They are stackable and collapsible, for instance.

This means companies can save space in shipment and massively reduce transport journeys via air, sea and land. It’s a win win for both profits and environmental sustainability.

The bottom line is that, as creators of packaging, we all need to think outside the proverbial box.

Like Willy Wonka with his sugary delights, we need to ask ourselves: “Is there a better way we can do this?” And, if there is, let’s do something about it. Actions always speak louder than words.

Related reading:

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World-Class Provider of Fingerprint Capture Technology Participating in Biometrics 2014, IACP 2014 Conferences

SPARTANBURG, S.C. – Integrated Biometrics, developers of the world’s smallest and lightest FBI compliant fingerprint scanners, is sharing its technological expertise and leadership at two major industry events this month.

Business Development Executive Thomas Buss is presenting at the Biometrics 2014 conference in London, England, and the company will also be participating in the 121st Annual IACP Conference and Exposition in Orlando, Florida.

“Government agencies, law enforcement, and military organizations rely on our fingerprint scanning technology to identify both friendly and unfriendly people while providing protection for their citizens,” said Steve Thies, CEO, adding, “We’re happy to contribute to their  efforts solving the problems of size, speed, accuracy and durability in our ever increasing mobile world.”

Buss, a 38-year industry veteran, is presenting, “Next generation Mobile Fingerprint Sensor Technology:  LES is More” at Biometrics 2014. His talk includes information on how the new light emitting sensor (LES) technology from Integrated Biometrics represents a paradigm shift for the industry, showing how the technology has been recently adapted for use in military and public safety applications.

The International Association of Chiefs of Police (IACP) conference provides over 13,000 attendees with access to the latest law enforcement technology via live demonstrations, hands-on exhibits and participatory events.

“Our success in developing products which are more ideal for  mobile environments than traditional silicon or optical sensors is due in large part to our technical team developing game changing  FBI Certified sensors with our unique and patented film; it’s very important to us to promote our solutions to government  markets worldwide,” Thies explained.

Integrated Biometrics’ world-class fingerprint solutions work in direct sunlight on dry or moist fingers, resist abrasion, and are 90-95% smaller and lighter than optical scanners. For more information visit www.integratedbiometrics.com.

About Integrated Biometrics
Integrated Biometrics provides enrollment and verification fingerprint sensors to hardware and software integrators serving government agencies and commercial markets worldwide. More ideal for mobile environments than traditional silicon or optical sensors, Integrated Biometrics FBI-certified fingerprint sensors utilize our durable, patented light emitting sensor (LES) film and work in direct sunlight on dry or moist fingers, resist abrasion, and are 90-95% smaller and lighter than optical scanners. Integrated Biometrics offers the only Appendix F FBI-certified sensor that meets mobility requirements demanded by end users, solving the major problems of size, speed, accuracy and durability. Find out more online at www.integratedbiometrics.com.

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Ladies and gentlemen, welcome aboard ALLpaQ AIR. We have some pre-flight information about how this bioprocess container optimises the airfreight of media held under temperature controlled conditions in an environmental chamber.

Before lift-off, please place a single-use bag inside a bioprocess container, fill your media into the bag and, to maintain product temperatures, load the bioprocess container inside a Envirotainer RAP e2 container.

Now take count of how many passengers fit inside. One? Two? Three? Four? Owing to the dimensions of standard bioprocess containers, healthcare companies have previously only been able to ship two units per Envirotainer RAP e2 container.

Does this match your calculations? If you can only fit two bioprocess units inside this environmental container then surely you’d like to fit four? After all, the more media your company ships per flight the more you save – it’s not a case of less is more but rather more is less.

AIR’s modified footprint doubles the payload, allowing each container to be filled to the brim with four bioprocess containers. Filling each Envirotainer RAP e2 container full of AIR gives lift-off to:

Like one Russian Doll seamlessly slotting into another, AIR is dimensionally designed to slot snuggly inside of a Envirotainer RAP e2 container. This means that when your media is cruising at 35,000 feet, the financial forecast outside will always be sunny en route to your destination.

Getting ALLpaQ AIR off the ground


AIR took flight when a client in the healthcare sector approached ALLpaQ with a need to ship 600+ litres of biopharma media between mainland Europe and the USA.

The problem: Dimensionally the company was restricted to transporting a maximum of two units per Envirotainer RAP e2 container.

The solution: A bioprocess containment modification project to ensure that the company maximised every inch of space within each environmental chamber.

Key objectives included:

The outcome: AIR, which has been independently validated, allows for four units to be held within each Envirotainer RAP e2 container. ALLpaQ calculates that AIR will generate savings of circa €1 million in transportation costs to the client over the lifecycle of the project.

Thankyou for flying ALLpaQ AIR

As we start our descent, please make sure your tray tables are in their full upright position. On behalf of our crew, thank you for choosing ALLpaQ AIR – the only bioprocess container committed to giving your media a first class journey at economy costs.

If you have any questions about today’s flight and why ALLpaQ AIR is the most cost-effective solution of its type when it comes to air cargo, please ask one of our fluid handling attendants.

Make contact to book your flight plan

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Lumiprobe Newsletter - Bodipy analogs - Fluorescent probes!
Lumiprobe adds
BDP FL azides * BDP FL NHS ester * BDP FL maleimide
BDP FL - borondipyrromethene dye is an excellent dye for fluorescein (FAM) channel.

Thiol reactive BDP FL maleimide is a reactive dye for protein labeling, which has identical structure with BODIPY ®   FL maleimide.

BDP FL is a borondipyrromethene dye which has absorption and fluorescence spectra similar to fluorescein (FAM). However, this dye exhibits very high photostability. It is non-charged, and has low molecular weight. Its brightness is similar to fluorescein, R110 and xanthene dye derivatives like Alexa Fluor® 488.

This fluorophore is ideal for fluorescent microscopy and many other applications. The fluorophore can substitute fluorescein for almost any application, and it is compatible with any FAM-capable fluorescent instrumentation.

 

BDP FL azide is an analog of BODIPY ® FL azide, a Click-chemistry capable bright and photostable dye for FAM channel.

This green-emitting fluorophore is compatible with all types of fluorescence measuring instruments for FAM (fluorescein) and dyes like Alexa® Fluor 488.

The fluorophore is a representative of borondipyrromethene class of fluorescent dyes, which possess high quantum yields in aqueous environments, and high stability towards photobleaching.

BDP FL NHS ester is an advanced dye for 488 nm channel, a replacement for fluorescein, a molecule identical to BODIPY FL ® NHS ester. An amino-reactive dye for the labeling of proteins and peptides.

While the absorbance and emission spectra of this molecule stay within FAM excitation and emission channels, this dye provides much better photostability, and outstanding brightness. The fluorescence spectrum of BODIPY-FL is narrower than that of FAM. This provides a better brightness for monochromator based instruments, when emission wavelength can be tuned to dye maximum.

The dye is neutral, possesses low molecular weight, and retain high quantum yield in conjugates.

The dye is a good replacement for fluorescein (FAM), BODIPY-FL, Alexa Fluor 488, DyLight 488, Cy2, and other 488 nm dyes.

Citations of interest - Take a look!
Would you like your paper featured on Lumiprobe citation page? Email: This e-mail address is being protected from spambots. You need JavaScript enabled to view it

Ultrasensitive fluorescence-based methods for nucleic acid detection: towards amplification-free genetic analysis


Ranasinghe, T.; Brown, T.
Chem. Commun., 2011,47, 3717-3735
DOI: 10.1039/C0CC04215C

Real time PCR is the mainstay of current nucleic acid assays, underpinning applications in forensic science, point-of-care diagnostics and detection of bioterrorism agents. Despite its broad utility, the search for new tests continues, inspired by second and third generation DNA sequencing technologies and fuelled by progress in single molecule fluorescence spectroscopy, nanotechnology and microfabrication. These new methods promise the direct detection of nucleic acids without the need for enzymatic amplification. In this feature article, we provide a chemist's perspective on this multidisciplinary area, introducing the concepts of single molecule detection then focussing on the selection of labels and probe chemistry suitable for generating a signal detectable by ultrasensitive fluorescence spectroscopy. Finally, we discuss the further developments that are required to incorporate these detection platforms into integrated ‘sample-in-answer-out’ instruments, capable of detecting many target sequences in a matter of minutes.

Tech Support: Lumiprobe offers FREE tech support before or after you order.
A few questions:

I just ordered the BDP FL-azide dye that I want to click onto my peptides which have an alkyne. Do you have a protocol for running this click reaction in solution? The one which you provide on the website is for oligonucleotides and I was wondering if you have a protocol more specific to peptides?

Lumiprobe recommends you to start with our oligonucleotide protocol. Compared to oligos, peptides have a couple of peculiarities. First, they may contain thiol groups which adversely affect Cu-catalyzed reaction. If this is your case and conjugation does not perform well, try increasing Cu catalyst concentration, and do your best to exclude oxygen from the reaction. If there are no thiols, conjugation is normally non- problematic.

Second, purification of the peptides is different - for example, acetone precipitation may not be appropriate. Use your favorite purification method, such as HPLC.

The protocol for oligonucleotides is appropriate for peptides in all other aspects.

I am looking for a probe to use in a polarization study of protein association-dissociation. The protein is a dimer of 100 kD that dissociates into two equal 50 kD monomers, so the change in polarization is very small using fluorophores with lifetimes like fluorescein or rhodamine, i.e. few nsec. The Kd for the protein dissociation is low, 50 nM, so the fluorophore needs to be pretty bright also. So, I'm looking for the usually incompatible combination of : a) high extinction coefficient, b) high quantum yield, and c) longer lifetime. It would be nice if the absorbance max is longer wavelength than fluorescein, but in any case must be longer than 350 nm. I would prefer NHS or maleimide chemistry, but can work with other options also.

Thank you for your email! It is quite a difficult task to find a dye fitting all of the desired properties. There are no dyes of Lumiprobe range that fulfill all of them, and I do not know about such products among competing product ranges.

BDP FL is a fluorescein-like dye which has similar brightness (i.e. high extinction coefficient and quantum yield), and reported to have fluorescence lifetime about 50% higher than fluorescein.

Pyrene is a polyaromatic hydrocarbon dye which is reported to have much longer fluorescence lifetime (about 25-fold of fluorescein). However, extinction coefficient is not high (about 1/2 of fluorescein), and it is a blue emitter (emission max around 450 nm). Lumiprobe offers Pyrene azide 1 and Pyrene azide 2 and also BDP FL NHS esters. The pyrene azide dyes also exhibits effect of excimer formation (you can consider this effect to study association, too).

Among Lumiprobe's cyanine dyes, there are red and NIR fluorophores, but they have lower quantum yields and fluorescence lifetimes similar to fluorescein, although their extinction coefficients are much higher.

So although I cannot suggest the product that fits all requirements, hopefully you can tailor some available products to your task!

I’d like to order azide dye for click reaction and like to know if you have green colour as I see only cy3 and cy5 in your website. If you do have green colour azide, could you please also send the quotation?

Lumiprobe can offer FAM azides - FAM azide 6-isomer and FAM azide 5-isomer , the two isomers have very similar properties and also BDP FL azide (an analog of BODIPY FL). Both are for FAM green channel.

Thanks a lot for your information. I am wondering if this FAM azide will be suitable for flow cytometry application as I have a plan to do both microscopy and FACS.

FAM is suitable for flow cytometry and FACS if your instrument supports this channel (almost all of them do).

BDP FL is also another dye for this channel, it is more photostable.

I'm searching for a protocol that would allow me to attach a fluorescent protein to a non-fluorescent one, possibly using the click chemistry. I was wondering if this is at all possible and if so, how many modifications/protein there are on average. I'm dealing with an enzyme and I care about preserving its catalytic activity hence I was wondering if it's possible to control the extent of its modification in order to limit it as much as possible. Preferably I'd like to have just few alkyne groups/protein.

It is possible to make conjugates between different proteins via Click Chemistry. You can label one with azide NHS ester, and another with alkyne activated ester , and then click them together.

However, there are more straightforward ways to achieve labeling:

If you need to label a protein, you can just use one of Lumiprobe's dye activated esters . For example, BDP-FL label will fit GFP fluorescent channel, other fluorescent proteins can be covered by other dyes, too. This will more likely retain your catalytic activity, because fluorescent dyes are small, and likely will label somewhere away from your catalytic site.

If you definitely need to label with fluorescent protein, it is probably better to use genetic engineering to produce a chimeric protein.

Please do not hesitate to contact Lumiprobe if you have more questions!

Do you would suggest to label the enzyme directly with a dye without using the click chemistry. Do you know how many dyes per enzyme one usually gets using this approach? Does your company have an experience of labelling enzymes? Thank you!

It is possible to make different loadings of dye per protein molecule with this approach.

Lumiprobe has experience in labeling proteins, but enzymes should not be very different if they tolerate pH around 8. We recommend you to use sulfonated dyes like sulfo-Cyanine3 NHS ester and sulfo-Cyanine5 NHS ester to label enzymes specifically in very mild conditions. Although the reaction looks simple, nevertheless, all substrates are different. In some reactions, it is possible that there is some chemical reason why labeling does not take place.

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Any comments on the newsletter? What information would you like in the next issue?
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Perl As Programming Language of Choice for Biologists!! - adidarwinian

The author of this paper explains Perl As Programming Language of Choice for Biologists, Bioinformaticists, Bioinformaticians, Programmers, and non-biologists. Advantages of Perl for biologists are explained in detail. Read full research paper at http://adidarwinian.com/perl-as-programming-language-of-choice-for-biologists/

 

Perl As Programming Language of Choice for Biologists by adidarwinian

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    The author of this research paper has lucidly explained why Perl is the Programming Language of Choice for Biologists, Bioinforma

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Lumiprobe Newsletter

When was the last time you visited Lumiprobe's updated website?

Take a look ! Lumiprobe manufactures and sells over 80 reagents offered in 1mg, 5mg, 25 mg, 50 mg and 100 mg sizes plus bulk and larger sizes available! Each page has pricing, availablility, information on structure, absorption and emission spectra, downloadable MSDS, and general properties.

Want a quick guide to what binds with what? amine reactive, azides, thiol mono, NHS esters, Azdes, maleimides, ROX, carboxylic, water soluble, BDF FL or Pyrene and more!
Lumiprobe's catalog by alphabetic listing

Custom Synthesis
Want something not in the catalog? Need a large volume? Lumiprobe has created dyes since 2006. Often a research lab requests a custom probe, that is added to the Lumiprobe catalog. What do you need to use in your research? Talk with Lumiprobe tech support and something can be created for you, at no additional development charges.

Downloadable Lumiprobe Catalog: http://lumiprobe.com/pdf/catalog.pdf

Lumiprobe supports all size research labs!

Featured Reagent : Delve into Dye NHS esters (Reactive dyes)
ROX NHS ester, pure 6- isomer | BDP FL NHS ester | Sulfo-Cyanine3 NHS ester
Sulfo-Cyanine 5 NHS ester | Sulfo-Cyanine7 NHS ester | Cyanine3 NHS ester
Cyanine3.5 NHS ester | Cyanine5 NHS ester | Cyanine5.5 NHS ester | Cyanine7 NHS ester | Cyanine7.5 NHS ester
Limelight on Cyanine5 NHS ester
During the last few years, Cyanine5 flurophore (analog of Cy5) has become an incredibly popular label in life science research and diagnostics. Fluorophore emission has maximum in red region, where many CCD detectors have maximum sensitivity, and biological objects have low background. Dye color is very intense, therefore quantity as small as 1 nanomol can be detected in gel electrophoresis by naked eye.

This Cyanine5 NHS ester (analog to Cy5 NHS ester) is a reactive dye for the labeling of amino-groups in peptides, proteins, and oligonucleotides. This dye requires small amount of organic co-solvent (such as DMF or DMSO) to be used in labeling reaction (please see our recommended protocol for more details). This reagent is ideal for very cost-efficient labeling of soluble proteins, as well as all kinds of peptides and oligonucleotides. This reagent also works well in organic solvents for small molecule labeling. For more sophisticated targets such as easily degradable proteins, when use of DMF or DMSO is undesirable, consider using water-soluble sulfo-Cyanine 5 NHS ester which does not require co-solvent, and has very similar fluorescent properties.

Protocols http://www.lumiprobe.com/protocols

Citations of interest - Take a look!
Would you like your paper featured on Lumiprobe citation page? Email

Use of Lumiprobe Cyanine dyes for diagnostic tool and drug development:
1) Liu, Z.; Miller, S.J.; Joshi, B.P.; Wang, T.D.
In vivo targeting of colonic dysplasia on fluorescence endoscopy with near-infrared octapeptide.
Gut, 013, 62, 395-403. doi:10.1136/gutjnl-2011-301913
Peptides were labeled with peptides were labelled with Cyanine5.5, a NIR dye, and characterised by mass spectra. Research demonstrated the created near-infrared (NIR) peptide is highly specific for colonic adenomas on fluorescence endoscopy in vivo and sub-cellular resolution images were achieved
Full paper: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3563943

2)Xie, M.; Shi, H.; Ma, K.; Li, B.; Shen, S. Wang, X.; Jin, Y
Hybrid nanoparticles for drug delivery and bioimaging: Mesoporous silica nanoparticles functionalized with carboxyl groups and a near-infrared fluorescent dye.

J Colloid and Interface Sci., 2013, 395, 306-314. doi: 10.1016/j.jcis.2013.01.001

The development of a drug delivery system with fluorescent biolabels is important in anti-cancer drug delivery application due to the potential for simultaneous diagnosis and treatment of diseases. Here, we reported the synthesis and multiple functionalization of mesoporous silica nanoparticle (MSN) for bioimaging and controlled drug release. After the functionalization with carboxyl group, the nanoparticles exhibited much better dispersity and stability in aqueous solution than MSN. Furthermore, a substantial doxorubicin (DOX) loading level was achieved and DOX loaded nanoparticles exhibited noticeable pH-sensitive behavior with accelerated release of DOX in acidic environment. Compared with native DOX MSN, DOX MSN/COOH Cy5 exhibited enhanced intracellular uptake efficacy and stronger effect on killing tumor cells. Meanwhile, it was observed that the MSN/COOH Cy5 was able to locate in the cytoplasm of MCF 7 cells and could accumulate in tumor tissues for a long period of time. Overall, the functional nanoparticle could potentially be used for simultaneous controlled drug release and near-infrared fluorescent bioimaging

Tech Support: Lumiprobe offers FREE tech support before or after you order.
A few questions:
Could you compare sulfonated vs non-sulonated cyanine dyes?
Read the Lumiprobe paper comparing sulfonated and nonsulfonated cyanine dyes http://www.lumiprobe.com/tech/cyanine-dyes

I'm looking at using Cyanine5 as a background dye in some PCR experiments to check for loading amounts of liquids into these experiments (add and scan on first cycle). Thus I'm looking for a non-reactive version of the dye to add in that will not interfere with proteins (such as polymerases) or DNA. Can Cy5-azide that is used for click chemistry be used for this application. Are there other possible Cyanine5 types that would be possible for this application.
Sulfo-Cyanine5 azide (http://www.lumiprobe.com/p/sulfo-cy5-azide) should be suitable for your application. While regular Cyanine5 azide has low water solubility, sulfo-Cyanine5 is very soluble in water. It does not seem to interact with normal biomolecules, and it has same fluorescence as other Cyanine5-containing probes.

Could you send me a quotation and delivery time for 25mg of 43020 Cyanine 5 NHS ester and its protocol?
Lumiprobe thanks you for your enquiry and sends formal quotes whenever requested. Each reagent on the Lumiprobe website
has its own webpage with information. Pricing, availability and information about Cyanine5 NHS ester is at: http://www.lumiprobe.com/p/cy5-nhs-ester
Under General properties there is downloadable MSDS. The Cyanine5 NHS ester has chemical structure, absorption and emissions spectra. Protocol for NHS ester labeling of amino biomolecules and other protocols are at: http://www.lumiprobe.com/protocols

What Cy5 polimethyne dyes can be used in hematology analizer with flow citometry, to differential of leukocites in 5 parts? The light source used in the system is a semiconductor laser emitting red light at 633 nm
Yes, it is possible to use this cyanine dye with a flow cytometer. If your instrument has Cyanine5 channel then it is compatible. Cyanine5 and sulfo-Cyanine5 are suitable dyes for your instrument - you can use them with your cytometer/sorter.

Do you ever have a sulfhydryl reactive version of a sulfonated Cy3 or Cy5?
Lumiprobe offers two sulfhydryl reactive water soluble cyanine dyes. sulfo cyanine3 maleimide and sulfo cyanine5 maleimide

Send your tech questions
Any comments on the newsletter? What information would you like in the next issue?

Visit Lumiprobe.com

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North Carolina Center of Innovation Network Accelerating Ideas, Driving Business, Establishing A Culture for Growth

RESEARCH TRIANGLE PARK & GREENSBORO, N.C. -- NC COIN (North Carolina Center of Innovation Network) has launched its new website, NCCOIN.org, to provide access to educational and networking resources and showcase its new look designed to highlight the organization’s mission to accelerate ideas, drive business, and establish a culture for growth.

Formerly the North Carolina Center of Innovation for Nanobiotechnology, NC COIN has changed its name to reflect its true mission, promoting innovative technologies in the areas of biotech, nanobio, agbio, advanced materials and medical devices, all of which use Nanobiological techniques and tools.

“Momentum has been steadily building for more proactive support in bringing together the community of active leaders in biotech, agriculture, and nanotechnology throughout North Carolina,” Joe Magno, COIN executive director said.

The new website debuts COIN’s newly redesigned logo reflecting the organization’s “Start Here” theme in the form of a start button. An energetic color scheme permeates the site to convey an uplifting sense of innovation and possibility.

Fresh content on NCCOIN.org communicates the many efforts of the organization to connect and unite human, intellectual, and financial resources to help members grow their businesses globally through a targeted focus on networking events, K-12 programs, educational conferences and meaningful seminars. Much of this is done through COIN events including Show-and-Tells, 90 minute events designed to promote networking and cooperation among companies, universities and researchers; Seminars to keep members informed on timely legal issues and relevant topics in accounting, insurance and capital formation; Half and Full Day Conferences where influential speakers cover a wide range of topics such as advanced manufacturing, intellectual property and big data; Conversations with subject matter experts to discuss life sciences and technology topics of interest; and Community Projects such as Centers of Gravity and K-12 after school programs.

“What emerges from our organized conversations isn’t just a set of best practices or updates on the latest advances in the field.  Our goals are more concrete -- we aim to provide resources and opportunities that will drive innovation and move ideas forward,” Magno explained.

“Our new website will also showcase our Founding Members, who are all major players in the ecosystems we are focused on growing” said Karen Shank, COIN’s Director of Business Development adding, “we are a membership supported orgainzation and all of our events and programs center around our member’s needs”.

For more information on membership contact Karen Shank, Director of Business Development at This e-mail address is being protected from spambots. You need JavaScript enabled to view it .

About NC COIN
The North Carolina Center of Innovation Network (COIN) is where conversations take place and connections are made and leveraged to accelerate ideas, drive business, and establish a culture for growth. We engage a diverse community of active leaders in biotech, agriculture, and nanotechnology through sought-after events, corporate connections, and innovative community programming to facilitate the global movement and exchange of ideas, the most powerful catalyst to enabling innovative economic growth in the 21st century. At the center of building momentum @ www.nccoin.org.

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Reintroducing Lumiprobe to new users for this academic year! Lumiprobe Fluorescent dyes, click chemistry reagents, Life Science Solutions -
For 5% Discount use code: biocrowd

Thank you for last year!  Visit Lumiprobe.com updated website. Perhaps you are unaware of all that Lumiprobe has done in the last year:

Over 80 reagents! Quick summary of Fluorescent and clickchem reagents:
http://www.lumiprobe.com/catalog/alphabetical

Each reagent page has information, sizes, absorption and emission spectra, MSDS, and general properties

Cyanine dyes explained:
http://www.lumiprobe.com/tech/cyanine-dyes

Tech support
ask questions, before or after you design your research. Lumiprobe will help you decide what to use. Also Lumiprobe can make custom probes for you

Protocols -   how to do click chemistry
http://www.lumiprobe.com/protocols
- Click-chemistry labeling of oligonucleotides and DNA
- NHS ester labeling of amino biomolecules
- Maleimide labeling of proteins and other thiolated biomolecules
- SYBR Green I staining of DNA in gels
- qPCR with SYBR Green I

Scientific citations
Over 100 papers offering ideas and info
http://www.lumiprobe.com/citations

Storage conditions - Most reagents good for 24 months!
http://www.lumiprobe.com/tech/reagent-storage-conditions

I'd like to read your papers. Send them to
This e-mail address is being protected from spambots. You need JavaScript enabled to view it

thanks!
Traz

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The hedgehogs are known for their act of turning into a spiny ball when in danger. The Hedgehog – the Spiny Ball!! - adidarwinian discusses biology of hedgehogs along with enchanting poetry on these beautiful spiny mammals!

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    Hedgehogs Demystified along with Poems on Hedgehog for Animals and Pets Lovers!!

Nonprofit Life Sciences Organization Strengthens Team of Industry Leaders as it Completes Start Up and Early Stage Legal Series

RESEARCH TRIANGLE PARK, N.C. -- NC COIN (North Carolina Center of Innovation for Nanobiotechnology) is continuing to strengthen its team of industry leaders looking to accelerate ideas, drive business, and establish a culture for growth by welcoming Dr. Sharlini Sankaran and Dr. Steve Greenbaum to the organization’s Advisory Council.

The additions come as COIN marks the completion of its popular, informative five-week Start Up and Early Stage Legal Series presented in concert with Shanahan Law Group and the NC Biotechnology Center.

“Expanding access to resources and opportunities like the Legal Series while gaining insight from respected professionals such as Drs. Sankaran and Greenbaum helps COIN open the gateway to faster development and commercialization of ideas, develop innovative K-12 community programming, and help drive new growth in North Carolina,” Joe Magno, COIN executive director, said.

Sankaran and Greenbaum join fellow industry, academic, research, and professional services leaders from across the state meeting on a quarterly basis to provide guidance and advisement to the COIN Board of Directors and staff regarding programs and events.

Dr. Sankaran is an experienced leader with public policy, economic development, and data management experience. She is Executive Director of REACH NC, a comprehensive statewide web portal to information on research expertise and capabilities at North Carolina’s universities and research institutions. Dr. Sankaran is an active and dedicated volunteer in K-12 and higher Science and Math education, having been involved in the Durham Women and Math mentoring program; serving as judge in the annual North Carolina FIRST Robotics tournament; and speaking on career development to graduate students and postdocs in the sciences and engineering. 

Dr. Greenbaum runs the Homeland Security Division of BAI, a government services company, leading diverse teams of experienced scientists and engineers planning, monitoring, and ensuring compliance for government-sponsored R&D programs. One of his roles at BAI is to build and maintain related workforce development initiatives including internships, consulting projects, and recruiting engagements with graduate programs in relevant scientific and technical areas.

Other COIN Advisory Council members include Dr. Amanda Elam, Galaxy Diagnostics; Dr. Sandra Merkel DeJames, Novozymes; Neal Fowler, Liquidia Technologies; Jinan Glasgow, Neopatents; Dr. Ginger Rothrock, RTI International; and Preston Linn, Nirvana Sciences.

Running from June through the end of July, the Start Up and Early Stage Legal Series was led by Henry Kopf III of Shanahan Law Group and consisted of informative, hands-on sessions designed for professionals involved in startups as well as established businesses and nonprofit organizations. Topics included Pre-Formation and Formation Issues, Hiring and Doing Business, Protecting Intellectual Property, Fundraising and Exits, and International Business Transactions.

In addition to gaining insights from Kopf -- the holder of four U.S. patents who concentrates his practice in the areas of business litigation, business advisement, constitutional takings, administrative law, and technology law -- attendees learned from guest speakers such as David Swintosky of Dunning Capital, an investment banking and financial consulting services group; John Hollenbach, former CEO & President of Doe & Ingalls Management (which was bought by Thermo Fisher); and General David L. Grange, a 30-year U.S. Army veteran and current president of Osprey Global Solutions.

This is the second time COIN has presented the Start Up and Early Stage Legal Series to provide attendees with a solid baseline of knowledge to enhance and improve their business ventures. Attendees were presented with the tools needed to build a proper foundation for their business to prevent future business problems and minimize damage and distractions such problems can cause in addition to learning how to better prepare to discuss issues with their legal team when required.

COIN connects influential business, technology, and educational professionals; provides forums for sharing information on new discoveries and important trends; and promotes innovative technologies to leverage our collective capacity to build a more vibrant economy. Founding members include Novozymes, Kymanox, Danis, CSC Leasing Company, Duke Energy, the North Carolina Biotechnology Center, Applied DNA Sciences, Triad Growth Partners, Total Facility Solutions, Shanahan Law Group, HP, McDonnell Boehnen Hulbert & Berghoff, Jones Lang LaSalle, Liquidia Technologies, Xanofi, RTI International, BAI and Bayer.

For more information on membership contact Karen Shank, Director of Business Development at This e-mail address is being protected from spambots. You need JavaScript enabled to view it .

About NC COIN
The North Carolina Center of Innovation for Nanobiotechnology (NC COIN) was organized to promote the economic development and application of nanobiotechnology and related technologies across the State of North Carolina by promoting innovation and commercialization in the fields of nanobiotechnology and nanomedicine. For information visit www.nccoin.org.

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For many biopharmaceutical companies, plastic is fast becoming the new gold. This is because ALLpaQ’s fleet of plastic bioprocess containers offer a range of ROI-generating benefits that their steel equivalents simply can’t.

Fabricating in plastic, for instance, means that the resultant bioprocess container can be stacked, saving valuable space in warehousing and shipment. Likewise, units can be folded and side-walls replaced to enhance operational longevity.

At ALLpaQ, plastic is indeed typically accompanied by the word fantastic. So it is that we’ve put together 5 interesting facts about our brethren material.

1. Formerly a Greek God

The word “plastic” derives from the Greek word “plastikos” which means mouldable. There are two groups of plastic – thermoplastics and thermosets.

Like wax, thermoplastics can be moulded and re-moulded many times under the right temperature. Thermosets, on the other hand, are akin to concrete and can only be moulded once.

2. Silly putty ain’t so silly

It was back in the 1940s when silicone-based plastic and boric acid were mixed together. From this fiery brew emerged a rather unusual compound which bounced 25% higher than rubber, never rotted and could be stretched every which way but loose.

This, ladies and gentlemen, became known as Silly Putty.


Get 3 more plastic facts at ALLpaQ

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