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Vinod Gupta is without a doubt one of the successful Indian businesspersons. He has established businesses which have rewarded him so much that he has joined the league of dollar billionaires. However, even with his success, he has never forgotten about his humble backgrounds in India. The small town he was born in played an important factor in making him the person he is today. Vinod Gupta attended school in rural parts of India before he was accepted into Indian Institute of Technology. He later moved to the United States and joined the University of Nebraska for a degree in agricultural engineering.
His first job was at Commodore Corporation. He was hired as a marketing analyst. One of the tasks he was to handle is gathering a list of mobile home dealers around the country. While carrying out this task, he realized that there was no information or a cohesive list to carry out the task. He had to order for phonebooks in print form in order to create his list. Vinod Gupta realized that the list he had compiled would be of much help to businesses around the country.
Vinod decided to creating the database a business idea. He borrowed $100 from a bank and created marketing mailers to send the database information to anyone needing it. Much to the surprise of himself and others, the orders for his database were overwhelming. In 1972, Vinod Gupta made his business official and named it American Business Information. His company expanded quickly as other businesses realized the worth of the business he had put up.
In two decades, the company he started with $100 was valued at s$500 million. He renamed the company InfoUSA and ensured that it had an established framework which would enable its transition into a technology-driven database company. He eventually sold Info USA in 20101 at $680 million. Get Related Information Here.
Over the years, Vinod has given back to the community and is especially committed to education, especially to Women's Education. He acknowledges that its one the most effective tools of fighting poverty and as such has been very determined to grow education in India.
See also: https://www.crunchbase.com/person/vinod-gupta-1a55


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Seek or Shout is a new — so new that it's beta — service from Cision, a company that helps PR professionals by offering databases of media contacts, media news and other support. According to the site:
Seek or Shout is a new community for anyone who creates or promotes content, including bloggers, journalists, freelance writers, public relations pros, marketers and students.
The idea is that bloggers and other writers can ask the community for sources, ideas, etc. and then promote the finished piece among their peers. Like other social networks, its success will not only depend on how well the site is designed, but whether anyone shows up.

subset: commoncrawl


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28,000 Records - 2019 Email Lists
For businesses that want to market their products and services to other businesses, an email marketing database gives you the opportunity to directly contact industry executives. Special Databases provides affordable, high quality email marketing lists so that you can power your business's email marketing campaign with accurate, relevant, up-to-date contact information. Our records are complete and double-verified, and we provide the best quality to cost ratio on the market.
If you are looking for the best value in email marketing lists, you have come to the right place. Special Databases is a premier provider of email marketing lists, and we take pride in honoring our commitment to quality above all else.
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With one of our email marketing lists and databases, you can reach tens of thousands of contacts through email marketing to promote your business, sell your products, and increase sales. Email marketing is a great way to get in touch with the people that will benefit from what your business has to offer, and Special Databases takes out all of the guesswork for you by providing a complete and comprehensive email database and list to get you started. Whether you are an established business or a start-up company, using one of our email marketing lists and databases will help bring your company up to speed with technology and give you direct access to industry insiders. Our email marketing lists are the best value because we offer the highest quality information at the most reasonable price. With our competitive prices and money-back guarantee, Special Databases is the only provider that you will need to find all of the email marketing lists that you can use to promote your business. Order one of our lists today and transform your business's marketing potential, grow your clientele, and drive more sales!


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Sonesta Great Bay Resort & Casino
Sonesta Great Bay Resort & Casino
Sonesta Great Bay Resort & Casino
LocationLittle Bay Road 19
Philipsburg, AN
Description: Sonesta Great Bay Resort & CasinoGreat Bay Beach Resort and Casino is situated on the Dutch side of Saint Maarten, 25 minutes from Princess Juliana's International Airport, and overlooking the Great Bay harbor. It is about a 10-minute walk from the downtown capital of Philipsburg where you can enjoy duty-free shopping.The resort s amenities include a full casino, infinity pool, heated outdoor whirlpool, gift shop, activity center, watersports, floodlit tennis court, panoramic elevator, and almost a mile of sandy beaches. For dining and drinks, the resort houses a lobby bar, swim-up pool bar, living room bar and lounge, Bay View Restaurant (buffet dining), Molasses Caribbean Restaurant (elegant indoor dinner dining) and La Cucina Eatery. The hotel also offers free parking. All rooms feature satellite TV, in-room movies, coffeemakers, hairdryers and individual climate controls.Please note that this property offers both all-inclusive and room only categories. Please choose the appropriate all-inclusive/inclusive room category if you would like for your stay to include meals, beverages and additional offers specific to the hotel. All-inclusive programs vary by hotel. The price listed for room categories not designated as all-inclusive reflects only the room rate, tax and fees and will not include meals or beverages.
Other Local Hotels
Captain OliverOyster Pond
Oyster Pond
Gran Melia Puerto Rico Golf ResortRio Grande
Rio Grande
Grand Case Beach ClubGrand Case
Grand Case
Holland House Philipsburg
Rio Grande Plantation Eco ResorRio Grande
Rio Grande
The Marbella Club VillasPalmas del Mar
Palmas del Mar
The St Regis Bahia Beach ResortRio Grande
Rio Grande


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Trump Wins Indiana GOP Primary, Fox News Projects
WATCH: Trump Connects Cruz's Father to Lee Harvey Oswald
'Are You Canadian?': Cruz Gets Into It With Indiana Trump Supporters
Fox News projects that Donald Trump will win the Indiana GOP primary.
The New York real estate mogul fended off a defeat by Texas Sen. Ted Cruz, who in recent weeks had stepped up his rhetoric against the front-runner.
The victory significantly improves Trump's chances of amassing the 1,237 delegates needed to clinch the Republican presidential nomination this summer in Cleveland.
With just nine states remaining in the nominating process, the stop-Trump wing of the Republican Party may be rapidly running out of options, analysts say.
Update, 7:15 pm ET: "Ted Cruz could not have done worse in this state if he had planned it," John Roberts reported from Cruz's headquarters in Indianapolis minutes after the results were announced.
He noted the senator's coordinated efforts in the days leading up to today's primary, including teaming up with John Kasich, announcing Carly Fiorina as his running mate, and even picking up an endorsement from Indiana Gov. Mike Pence.
Many had seen Indiana as "a firewall" for Cruz, leading his performance in this primary to raise questions on whether there is still a clear path forward, Roberts reported.
Vulgar Children, Parents Scream Curses at Trump Supporters
Jobs and Economy Most Important for Indiana GOP Voters, Exit Polls Show
We 'Don't See You as a Friend': Laid-Off Coal Miner Confronts Hillary in WV


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A glowing skin is something we all dream of. Nothing makes you feel as confident as having glowing, radiant skin. People buy more and more products to make their skin radiant and beautiful, but these products work only for some time. Home remedies can do wonders for the skin. These natural remedies can return your lost charm and glow, that too, instantly. So, don't waste hours in salon, just give a try to these home remedies.
- Mix spoon honey and lime juice. Apply it to your face. After 15 minutes wash it with warm water. Both honey and lime juice work as bleaching agents. As a result your face will start glowing instantly.
- Scrub your face with tomato slice for 15 mins and wash it with cold water. The natural bleaching agents in tomato will bring glow to your face.
- Take small ripe papaya slice and make paste. Mix honey to that paste. Apply this mix to your face and wash it with warm water after 15 minutes.
- Take two spoons of rice flour and mix with milk to make it paste. Scrub your face with this paste for 10 minutes and wash the face with warm water. This scrubbing improves the blood flow and eliminates the black heads. Your skin will become soft and smooth
- Put two spoons of multani in four spoons of water for 10 minutes. Make it as a mixture and apply it to your face as a mask. When the mask dried, wash your face with warm water.
- Mix one spoon sugar, few drops of lemon and oilve oil into a mixture. Scrub your face this mixture. Wash your face with warm water after 10 minutes.
- Two spoons of turmeric and one spoon of lime juice must be made into a mixture. And apply it on your face. Wash your face, after the mask is dried completely. Your skin will be fresh and soft.
- Mix one spoon papaya, avocado and cucumber paste into a paste. Apply it to your face. Wash it after 20 minutes. Your skin will get additional glow.


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I would like to present my hand painted oil on canvas painting (recreation) of the artwork BURNING FLIGHT . I made this piece with the same amount of soul and emotion just like the first original painting. This piece is created with oil paint on artistic canvas using my unique technique of a palette knife. The artwork has texture, you can feel the strokes by touching the painting. By purchasing on this site, you are buying directly from the artist Leonid Afremov. I guarantee your satisfaction and the best customer experience.
The certificate of authenticity will include the name of the owner who purchased the piece of artwork. The certificate is signed by Leonid Afremov. If you buying this painting as a gift, please provide us the name of the gift recipient for the certificate. We can also ship to the address of the recipient of your gift.
about this painting:
Have you ever thought how difficult it is to convey the movement in the picture? Paintings art that represents still-life scenes is one thing, but making a motionless canvas full of action and drive is a different story. Only truly talented artists manage to catch this elusive moment when the movement is most expressive and paint it in a natural way.
If you are into such ambivalent art, this piece of art seems to be a perfect option for you!
Leonid Afremov's style is famous for its expressiveness. This oil painting is one of the best examples to study its peculiar traits:
Many works in this paintings art gallery poetize the beauty of autumn parks beloved by the artist, but this one shows us another side of the artist's talent:
All in all, what message does this picture get across? We think it's the proof how wrong people are when they say that photos and videos can tell more than any picture. Yes, catching a movement is much easier with a camera than with a brush and paint… but is "easy" really better than "original" and "unique"?
We accept paypal or any credit/ debit card payment. We guarantee the safety of your information. The information is encrypted and not stored anywhere after the payment was completed. We will not share your information with any other site nor company. Your personal information is strictly confidential.
We use shipping for shipping around the world.
You will receive an email from the shipping manager with all the details about your purchase within 2 days after your payment.
Read more in shipping policy


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Date: March 8, 2018
Time: 11:30 am to 1:30 pm
Topic: "A Day in the Life of a Concrete Contractor"
The NVCAC is not equipped to accept credit cards onsite. Please plan to pay in advance of the meeting if you choose to pay by this method.
Please note:
A 3% convenience charge will be applied to all credit card payments.


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Ralph J. Roberts: A Life in Photos
Jun 19, 2015
Ralph Roberts played a key role in the development of the cable television industry.
But perhaps most importantly, he was a kind and humble human being who loved his family deeply and unconditionally. Take a look back at his extraordinary life.


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MEMORIAL DAY SALE STARTS NOW! 20% OFF SITE WIDE - USE CODE REMEMBERDM20 - excludes sales and clearance items only. FREE Shipping on US Orders $75+
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Stop the show in our Kori floral dress! This dress features a single snap closure at the base of a wrapped v-neck, an elastic waist, ruffle short sleeves, and a flowing wrap-hem ruffle skirt. A beautiful floral design that ensures a perfect and flattering fit.


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The Additive Manufacturing Optimisation and Simulation (AMOS) project is developing a number of additive materials and processes for the repair of aircraft parts.
An ageing aircraft fleet and the recent global recession have contributed to the increasing cost of aircraft maintenance, repair and overhaul. An aircraft's wings, engines and fuselage are susceptible to damage both on ground (by service and maintenance equipment and fixtures) and in the air (by foreign objects, birds, lightning strikes etc). Although scheduled maintenance checks are performed throughout the lifecycle of an aircraft, defects can occur at any time and affect the performance of the aircraft. Consequently, unscheduled maintenance becomes necessary to replace defective components and ensure safety, reliability and airworthiness, and as each defect is different, unique solutions are required for each one.
Direct energy deposition (DED) systems are a very flexible type of additive manufacture (AM). They show great potential for the cost-effective and efficient repair / re-manufacture of aerospace components such as turbine blades and landing gears. Their use will allow damaged components can be repaired (on-demand) and material lost in service to be re-deposited to restore the component to its original shape. This would reduce repair lead times, costs, and material waste and extend the service life of damaged or worn components as damaged components could be quickly repaired rather than scrapped.
The Additive Manufacturing Optimisation and Simulation (AMOS) project is working with a number of different AM processes and material so as to assess their use for repair. The objectives of the project are to:
AMOS results were presented at the launch of the Nuclear AMRC Midlands in Derby.
Around 150 regional manufacturers and stakeholders attended the launch event at the iHub on Wednesday 13 February, to experience the state of the art in manufacturing R&D in the Nuclear AMRC's new workshops, and jo...
Read more
3DPRISM has released an open-access online training course for additive manufacture (AM). The course includes information on AM systems, materials, and process parameters as well as giving training on CAD modelling for AM, basic maintenance and health and safety. 3DPRISM was a 27 month project funded ...
Xiaoyi Guan from McGill University in Montreal spent 10 days in France at the ECN campus in June for AMOS, looking at the additive machine which will be used to deposit Titanium in the project
AMOS is linked to the Canadian MANU705 submission as per the rules of call MG1.9 International cooperation in aeronautics with Canada.
2017 © All Rights Reserved.


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Venue Named Among Best in Oregon
With 300 acres of scenic filbert orchards and parkland, Dorris Ranch in Springfield is one romantic setting for weddings.
Brides and grooms can choose from two locations that pile on the charm: the turn-of-the-century Tomseth House and the rustic barn.
The park and its offerings received national recognition this month, when the bridal planning website Wedding Spot named it one of the Best of Oregon for 2017.
"Every year, we recognize the top venues across the nation through our coveted Wedding Spot Award," the company said. "Out of 13,000 wedding venues on Wedding Spot, you were voted this year's best of the best!"
Recent bride Megan Felton said her wedding at Dorris Ranch "turned out beautifully."
"It was amazing, and more than I expected it was going to be," Felton said. "If you want something that's romantic and peaceful, Dorris Ranch is definitely the best."


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Microgaming's Environment & Energy Efficient New HQ Open for Business (VIDEO)
Exciting times lie ahead for the Isle of Man based gaming developer Microgaming, one of the world's leading producer and supplier of online casino software and services.
Proud to be a socially responsible member of the community over at the Isle of Man, Microgaming has moved on to reaffirm the status by opening new 'environment and energy efficient' headquarters called 'Sixty Two'.
Microgaming's new offices are developed adjacent to the existing building. The company's new HQ started construction in April 2015 and was completed on 31 July 2017, but have just now been officially open for business.
New HQ Building
The five-storey building houses over 100 employees and features a unique sky bridge joining the new and the old building office, thus creating a unique business campus for the company.
Microgaming CFO John Coleman said the new headquarters have been build based on the understanding of employees' needs and requirements, adding that the new building was specifically designed with their ideas in mind.
Staff and suppliers all had a say in the concept of the new building which is officially put to use after a number of years of hard work and commitment to see this project successfully through.
Microgaming further explained their new building had been built to meet the latest environmental requirements and energy efficiencies. It will include functions such as smart lighting which automatically dims to reduce energy consumption, whereas the air quality, temperature monitoring and centralised recycled facilities on each floor have also been incorporated.
From only six members of the staff back in 2001 when Microgaming first set up its office on the Isle of Man, the company evolved into a two-building campus in the heart of Douglas and with a sky bridge that was the company CEO's special wish.
"Sky Bridge was a must for me!", Roger Raatgever said.


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Venezuela's close ally China said it believed voting in the country's Constituent Assembly election was generally held smoothly, brushing off widespread condemnation from the United States, Europe and others and evidence of voting irregularities.
The U.S. government slapped sanctions on Venezuelan President Nicolas Maduro earlier on Monday in response to Sunday's election, which it called a sham.
Governments from Spain to Canada to Argentina and Peru joined Washington in denouncing the vote, which was boycotted by the opposition and widely seen as an affront to democracy.
Venezuela inflated the turnout figures for its constituent assembly election by at least 1 million votes, the company that provides the country's voting machines said on Wednesday, an accusation the government quickly dismissed as irresponsible.
But in a statement late on Wednesday, China's Foreign Ministry said it had noted that the elections were generally held smoothly, though it also noted the reaction from all relevant sides.
China does not believe in interfering in the internal affairs of other countries, it added.
We sincerely hope that all sides in Venezuela can orderly resolve the relevant issue with peaceful dialogue within a legal framework, and protect the country's stability and socio-economic development, the ministry said.
China believes that the Venezuelan government and people have the ability to properly handle their internal affairs. A stable developing Venezuela accords with all sides interests.
China and oil-rich Venezuela have a close diplomatic and business relationship, especially in energy.
Smoothly!!!?? Which planet are they on? It was rigged and violent from the outset.Aug 03rd, 2017 - 08:39 am +1
I think China owns Venezuela so they would kinda support it.Aug 03rd, 2017 - 11:19 am +1
Díme a quién apoyas y te diré quién eres.
Venezuela and China. And North Korea. And Cuba. Birds of a feather.
Commenting for this story is now closed.
If you have a Facebook account, become a fan and comment on our Facebook Page!


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Alexander McQueen
Light Blue Cotton Dress
This gorgeous dress from Alexander McQueen is crafted from a quality blend of cotton with an elegant embroidered flower design. This sleeveless masterpiece features a round neck, a fitted waist, a flared style and a short length.
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Keri Systems, Inc.
Since 1990, Keri Systems, Inc. has become one of the world's most popular providers of card access control and integrated security systems. Keri offers a full range of systems from simple products for a single door at one location to systems with thousands of doors spread over multiple locations, integrating photo badging, video, biometric, graphic maps and telephone entry. Their products are cost-effective, easy to install and use and can be customized and expanded as needed. They are also extremely reliable, with durable hardware products with an unconditional 2-year warranty as well as a lifetime warranty on proximity reader technology.
- Doors.NET™ software
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LOS ANGELES, CA (KTVI) – Todd "Too Short" Shaw, 46, was booked on DUI and drug possession charges early Wednesday.
L.A. police suspected the rapper of driving under the influence according to the NY Daily News. In addition to attempting to run away from police on foot, the he also allegedly attempted to ditch narcotics in a squad car.
Too Short is a veteran in the rap industry. He's been one of the most visible west coast rappers of the last two decades. Shaw's influence on the current generation is apparent as other artist have paid homage to his music. Rapper 2 Chainz told MTV that Too Short was a 'Father Figure' to him in growing up during an interview.
More Details: The New York Daily News


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The John Kenneth Gott Library is pleased to announce our collaboration with the Northern Piedmont Community Foundation for the upcoming Give Local event on May 5th.
This year marks the second annual Give Local Piedmont with 133 local non profit organizations participating. Help us raise $1,000,000 in one day for local community organizations spanning Fauquier, Culpeper, Madison, and Rappahannock counties.
Join us in brightening our future and supporting our community!
Read more about the Northern Piedmont Community Foundation here:
Learn more about the John Kenneth Gott Library and Fauquier Heritage and Preservation Foundation here:


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A smirking paedophile who sent sick explicit pictures of himself to girls as young as 12 has been jailed .
Joshua Manton preyed on four youngsters aged 12 and 13 from across the country using social media.
The 19-year-old encouraged the children to exchange the indecent images , the Chronicle Live reported .
A court was told Manton, from Burnley, knew the girls were underage but had still tried to meet up with one.
His crimes only came to light when the mum of one of his victims spotted the incriminating photographs on their phone.
Manton had lived in Newcastle while studying psychology and criminology at Northumbria University.
He was a guitarist for Cult Lust, who described themselves as a "scuzz-rock mosh pit punk" band.
His former bandmates have rushed to distance themselves from him.
The band are going to change their name and re-record the material Manton had a hand in creating.
Manton was traced back to his home address on Westwood Road, Burnley, in January 2015.
Police seized his mobile phone before trawling through thousands of messages and finding pictures and videos of his genitals.
Sam says he only met Manton earlier this year at a gig.
He added: "He has always been touchy people having access to his phone and his computer. He would never let us touch anything to do with his social media, and that makes sense now."
Defending, Michaela Williams told the court her client was immature and understood the consequences of his actions.
At Northampton Crown Court Manton was sentenced to two years and four months in a young offenders' institution, but will only spend half of that behind bars before being released on licence.
The judge also ordered him to sign the sex offenders' register for at least 10 years.


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Total Views: 414
ISSN 2354-4147: International Standard Journal Number (ISJN) e-ISJN: A4372-2604
Pollution Monitoring and Control in Rubber Industry in Nigeria
Article Number: DRJA392601501
O. Momodu1*, A.O. Ekundayo2, I.S. Ogiehor2, A.F. Iyayi1, S. Ohaga1 and D. Momoh1
1Rubber Research Institute of Nigeria (RRIN), Iyanomo, Benin City, Edo State, Nigeria.
2Department of Microbiology, Ambrose Alli University (AAU), Ekpoma, Edo State, Nigeria.
*Corresponding author E-mail: Â email@example.com.
Received: May 7, 2017 Accepted: June 16, 2017 Published: July 28, 2017
The production activities of the rubber industry in Nigeria generate a large quantity of rubber (wastewaters) effluent most of which are often discharged untreated into nearby surroundings. A study carried out in 1994-1997 on the numbers and types of rubber processing factories and effluent generated and discharged in Nigeria and their modes of disposal/ control revealed that 41 rubber processing factories were located in 10 states of the country of which only 27 were still functional. Of these functional factories, 18.5% produced ribbed smoked sheets (RSS), 77.8% technically specified rubber (TSR) and 3.7% latex concentrate. Seventy-four percent (74%) had serious pollution problems while 18.5% had mild pollution problems. Fifty-five percent (55%) had received complaints from the residents living around the factories, of odour and pollution of their source of domestic water. Only 7.4% had good effluent treatment facilities in place while 11.1% had simple effluent treatment systems and 81.1% did not treat their effluent at all. While 66.6% of factories discharge their effluents into water bodies (rivers/streams), 33.4% send theirs to nearby waste/farmlands. However, recent investigation has shown that the number of the functional factories have reduced tremendously though some new ones have emerged, the overall pollution situation by the rubber industry is still bad. To improve the pollution levels caused by these processing factories, some efficient treatment systems in form of physical, chemical and biological processes can be used to reduce the pollution load of the effluent to tolerable limits.
Keywords: Rubber effluent, Pollution, Effluent treatment, Monitoring.


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I started out not liking this book because the style was different for me. The sidebar parenthesis's were annoying and distracting. It felt like a chore to keep up.
Once they were gone, or less frequent, I started to get into the story and the characters.
I want to continue with the series because I want to see how things play out with Brandy, Peggy Sue and Vivian. Especially since Vivian said she wanted to tell Brandy something about Peggy Sue and never did.


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With the elections almost upon us politicians up and down the country are growing ever more frenetic in their efforts to convince you that "I AM THE ONE"! The last minute attempts to ensure all bases are covered has more than a few in a dervish-like trance-state of near exhaustion. The whole thing is a great game, dirty tackles and all! Not that you can expect any dirty play from your Green Party candidates. We are not ones for petty off-the-ball harrying or name-calling even if we do not shy away from going in with the shoulder! We leave gratuitous mud-slinging to others. I am listening to a panel on Sean O'Rourke's radio show (|RTE 1) discussing the good, the bad and the ugly of local democracy and the doing away with our Town Councils. Will we miss them? The general feeling is that their days were numbered way back in the 1970s when much of their power was transferred to County Councils. One thing is certain: with Town Councils gone we will make a big saving on unjustifiable expenses and junkets. What chance we might see reform to present abuses in this area at County Council level? Now, more than ever, people want to know that it is not simply a case of business as usual in the Council. They want public representatives who will do what is right rather than doing what they are told. There is a palpable sense of anger towards politicians of all persuasions, a sense that a promise made is a promise waiting to be broken, and that, for the majority of politicians, party will always come before the common good. No wonder people find it hard to trust politicians. Let us hope that our incoming Councillors can learn to work together, for the good of his constituents, in a responsible and transparent way. Councillors need to have one overriding aim, one which starts with the question: "What can I do to make Wexford a better place to live, shop and work?". Councillors cannot wave a magic wand and make everything alright. What they can do is lay the groundwork to allow us build a better Wexford whether through better policy development and more careful allocation of scarce resources. They can, and must, ensure that tax/rate payers get a bigger bang for their buck by working more closely with local business and the community. Such an approach can deliver better services, better roads, safer streets, and a Wexford that is open for business.
Recent Posts
- Birdwatch Ireland and the Heritage Bill
- 200 million euro capital programme for Wexford
- Hinkley Point C Nuclear Plant
- Same Old, Same Old!
- Climate Change: the government still doesn't get it
- Waterford to host Convention 2017
- Challenges facing Rural Ireland
- South East Economy Continues to Stagnate
- No Minister for Environment?
- Green Party welcomes Government u-turn on introducing pay-by-weight charges
- Green TD attacks parties for putting politics ahead of country
- 'This election is make or break for the Green Party'
- Next politician that calls to your door ask him/her to outline their party's environmental policy
- Same as it ever was!


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Agencies: New York Police Department NYPD
Last updated: 11 months ago
NYPD officers from the 75th Precinct Anti-Crime Unit shot and killed a 17-year-old boy during a shootout in East New York Thursday.
Police say Kadeem Torres was fleeing plainclothes officers near Fountain and Blake avenues around 4 p.m., when he drew a gun and fired.
Police chased him to 890 Belmont Ave., returned fire and killed him in the backyard.


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Over the course of European trading the Euro attempted to advance on the Pound in response to positive economic reports for the Eurozone.
While the Pound softened following the release of the UK's consumer price index, the Euro was supported by domestic data and managed to trim recent declines.
Data compiled by Eurostat showed that the Eurozone's trade surplus widened in February on an annual basis thanks climbing export levels and stagnating import levels.
Eurozone exports increased by 3 per cent on the year – resulting in a foreign trade surplus of 13.6 billion Dollars in February.
In January the trade surplus stood at a revised 0.8 billion Euros.
Meanwhile, Germany's ZEW current situation survey surged from 51.3 last month to 59.5 in April. A reading of 51.4 had been expected.
But it wasn't all good news.
The German ZEW economic sentiment survey slipped slightly from 61.5 in March to 61.2 in April while the nation's expectations measure dropped from 46.6 to 43.2 – considerably less than the reading of 45.0 expected.
The decline in investor confidence in the Eurozone's largest economy was the fourth consecutive monthly drop and reminds investors that German economic stability is at least partially reliant on external factors.
According to senior economist Christian Schulz; 'The crisis in Ukraine continues to sow uncertainty. Fortunately, strong Germany is in the best position to deal with the fallout from the Ukraine crisis. That may act as a shield for the Eurozone recovery.'
Yesterday's comments from European Central Bank President Mario Draghi regarding the possibility of a strong Euro forcing the introduction of additional stimulus also limited Euro gains.
Meanwhile, the British Consumer Price Index showed that UK inflation slowed to a four and a half year low in March, adding to the case for the Bank of England holding interest rates and triggering modest declines in the Pound.
Tomorrow's Eurozone inflation reports are bound to have a notable impact on the Euro exchange rate.
The final data are expected to show that the Eurozone's CPI advanced by 1.0 per cent in March month-on-month following a gain of 0.3 per cent in February. On the year the consumer price index is projected to show an increase of 0.5 per cent, with core CPI coming in at 0.8 per cent.
The Euro to Pound (EUR/GBP) exchange rate is also likely to be affected by UK employment data while, US manufacturing production figures could drive EUR/USD movement.
Investors interested in the EUR/CAD exchange rate meanwhile will be focusing on the Bank of Canada's rate decision, taking place at 15:00 GMT on Wednesday.
Euro (EUR) Exchange Rates
[table width="100%" colwidth="50|50|50|50|50″ colalign="left|left|left|left|left"]
Currency, ,Currency,Rate ,
Euro,,US Dollar,1.3807 ,
Euro,,British Pound,0.8255 ,
Euro,,Australian Dollar,1.4708,
Euro,,New Zealand Dollar,1.5978,
Euro,,Canadian Dollar,1.5173,


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American High School
- Director:
- Sean Patrick Cannon
- Release Date:
- 07 Apr 2009
- Genre:
- Comedy
- Country:
- Runtime:
- 86 min
"American High School," a romantic comedy, is centered on two people who marry young and contend with the consequences of "bad" actions. Moreover, it is a story about a young, frightened girl, Gwen Adams who tries to break away from her father, find peace with her husband, and find peace within. In a world that expects everything from her, being herself was the last thing on her mind. When pushed to the breaking point, she discovers her own self-worth. But is it too late?
Download available
- Runtime:
- 86 minutes
- Language:
- English
- Subtitles:
- No
- Resolution:
- 480x272 px
- Quality:
- Normal
- Price:
- $0.35
Actors movie: American High School
- writer
- Gwen Adams
- Holden Adams
- Hilary Weiss
- Principal Mann
- Miss Apple (as Nikki Ziering)
- Jonny Awesome
- Mr Seuss


text:
The Lifeskills Program can be developed and managed in stages for a single class, a whole cohort, stages of learning or whole of school. It can be implemented within curriculum, across curriculum or as a special area of learning, for example, Gifted and Talented students. The program also complements other enterprise learning tools that are available, for example the Real Game series.
The Lifeskills Program enables young people to develop enterprising skills, attitudes and creative approaches to learning right across the curriculum. This "learning by doing" program satisfies the delivery of outcomes of the Commonwealth initiatives of Enterprise Education and of Consumer and Financial Literacy as well as many of the key learning outcomes of the NSW K-6 syllabuses. Setting up and running a company is something that children and young people of all ages will benefit from. But it isn't just about setting up in business. It is about giving young people the chance to experience real business and work-related learning through entrepreneurial enterprise activities.
Please click here for further information about the Lifeskills Program.
The student led program facilitates the delivery of outcomes of the Commonwealth initiatives of Enterprise Education and Consumer and Financial Literacy as well as many of the learning outcomes of the NSW DET Key Learning Areas.
The Program has a core element (Float) and three dimensions (Earn, Spend and Underwrite).
Please click on the red highlighted text to view each section of the program which has been linked to the some NSW Primary Curriculum areas,
Quality Teaching elements and Financial Literacy Dimensions.


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Summer Resorts 2003: Mammoth Mountain, California
Every summer, 1.5 million visitors pass through the gates of Yosemite National Park. Unless you get to one of those gates at sunup, admission can mean idling on an access road for hours. Why battle Yosemite's traffic when you can go to Mammoth Mountain-located on the eastern edge of the national park-and bike, hike, fish, golf or relax without wasting time behind the wheel?
Mild temperatures and California sunshine provide the perfect weather; Mammoth provides the singletrack (80 miles of it). The fit can brave the climb to the 11,053-foot summit, breaking for refreshment at canopied water stops, while the less ambitious can board the gondola for a scenic seven-minute ride to the top.
Anglers will want to pack their permits and tackle. Fishing bliss abounds in the crystal blue mountain lakes and streams of the High Sierra. Beginners needn't worry: The Orvis Flyfishing School will show you how to outwit the region's legendary golden trout. If golf is your game, let the pros at the Sierra Star-California's loftiest 18-hole course-guide you to your best game.
Come evening, unwind with a stroll through the new 20-acre pedestrian Village at Mammoth. Or kick back with a pale ale at Hennessey's Tavern or Dublin's in Mammoth Lakes, and toast your cleverness. A national park-caliber vacation, minus the crowds: brilliant.
•DON'T MISS Ride round-trip on the YARTS bus ($40; 800-626-6684) from Mammoth into Yosemite Valley.
•BEST DEAL A package including one night's lodging and either a round of golf or guided flyfishing starts at $123 per person, double occupancy. 800-626-6684; mammothmountain.com


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Australia's Ross Smith-Glenn Warfe caused a huge upset in the Yonex-OCBC US Open Grand Prix Gold on Wednesday when they stunned the top seeds in the men's doubles competition.
The duo, who are ranked No. 69 in the world, took out Taipei's Fang Chieh Min-Lee Sheng Mu in the second round of the US$120,000 tournament with some splendid attacking play.
Read full news detail on BWF


text:
Saturday, November 8, 2014
A Flower for You
14 comments:
Ooooh, this is beautiful!!!ReplyDelete
Gorgeous card & love all the bling!ReplyDelete
Thanks so much for playing along with us @ Love to Scrap! & hope to see you back again for our *Say It In Color* challenge!!!!!
lovely card with such lovely colours thanks for joining us on our first challenge Jane Dt xReplyDelete
So beautiful Linda, I love that frame. Thank you for sharing with us at A Creative Need this month!ReplyDelete
This is so pretty, love the colors. Thanks for playing with us over at Loves Rubberstamps!!!ReplyDelete
Oh Linda this is so gorgeous - love the rich colours and the black to contrast and enhance them.ReplyDelete
Fantastic project. Thanks for playing with us at Party Time Tuesdays!ReplyDelete
This is so beautiful and love all of your fantastic details. Thanks so much for joining our challenge over at Loves Rubberstamps!ReplyDelete
So pretty! Love the card shape and the way you used the pearls. Thanks for joining us at Aud SentimentsReplyDelete
Good Luck
Julie T
- AnonymousNovember 11, 2014 at 1:23 AM
Hello, A lovely project, I love the image and the colour choice. A lovely style. Thank you for joining us this month with our first challenge at A Crafty Mess Good luck. We would love to see you again next month.ReplyDelete
Teighan DT
What a beautiful card, I love the black die and the addition of the pearls really makes it stand out. I love the flower and the tag with the sentiment is perfect for our challenge at Aud Sentiments, so glad you joined us so I could see your lovely work.ReplyDelete
Wow, this is beautiful. Love the die you used and how you filled it with pearls. We're delighted that you joined our Challenge last week at Word Art Wednesday. We hope to see you again this week. Leaving you with God's Word in Hebrews 12:2, "Let us fix our eyes on Jesus, the author and perfecter of our faith, who for the joy set before him endured the cross, scorning its shame, and sat down at the right hand of the throne of God." Have a blessed day.ReplyDelete
This is gorgeous!!! And I was right I thought it was a Spellbinder's die!!! Love the Penny Black stamp you used too!!! TFS!!!ReplyDelete
so pretty thanks for joining us at sisteract and good luck in the drawReplyDelete
hugs Tonnie DT
Thank you for your comment, I read and appreciate each one - you make my day! All comments are moderated and spam deleted.
Those leaving a comment on any post on this blog do so in the knowledge that their name and blog link are visible to all who visit this blog and by providing the comment, they have published their own personal details on this blog.{{hugs}}}


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Our Plant
Located in the city of Jiangmen in Guangdong province, we are strategically located near Xinhui International Port. Not only we strategically located to deliver our lubricants to entire China, we can also export lubricants to anywhere in the world. The plant, covering a 55,000 square meters land, can produce up to 50,000 tons per year and can be expanded to 100,000 tons per year.
Lubritech Limited blending plant uses new state-of-the-art equipments, and it is one of the most advanced independent lubricants blending plants operating in China. The blending plant was fully designed by a well known Germany Consultant Company, and it was completed in 2010 with an investment of USD20million.
Managed and controlled by a computerized operational system, our plant is equipped with the latest lubrication technologies such as simultaneous metering blending (SMB), automatic batch blender (ABB), drum decanting unit (DDU), and full pigging systems. In order to ensure the highest quality and cleanest of the blended oil, our pigging systems are designed through the whole operational pipelines; from DDU to any ABB, from ABB & SMB to any finished tanks, and from any finished tanks to all the filling / packaging lines.
Our plant is certified in ISO/TS16949:2009 with a very stringent quality process in every facet of its operation. Besides, we are also certified in ISO14001/2004 and ISO9001:2008, with its quality and environmental management systems are meeting or exceeding international and local requirements. Being one of the few lubricants plants certified by ISO/TS 16949:2009 and ISO 14001/2004, we are also one of the few lubricants plants that have been granted its export license and we are allowed to export lubricants to overseas.


text:
Roger J. Morneau
Roger Morneau was a retired salesman who has dedicated his life to an intercessory prayer ministry. He wrote A Trip Into the Supernatural, Incredible Answers to Prayer, More Incredible Answers to Prayer, When You Need Incredible Answers to Prayer, and The Incredible Power of Prayer.
AmazingFlawless! An unimaginable timeline events of Mr. Morneau. Absolutely a must have book! One that can change ones entire perspectives of life as we know it. The only one thing I would like to point out is the grammar error or missed spelled words throughout the chapters. I purchased the Ebook version and I'm not sure if the hard copy has the same grammatical errors.


text:
On Wall Street, the prices of shares in technology companies have been bouncing around even more than most other industry sectors. Tech stocks were battered on Tuesday, driving down the overall market.
The picture is mixed and uncertain. But based on the early readings before quarterly earnings begin in earnest soon. I.B.M., surrounded by rumors of weakness, announced its third-quarter results in advance last week, beating analysts' estimates. Oracle, too, says it is holding up fine. But SAP, the enterprise software maker, and Rackable Systems, which supplies server computers to big Web companies and many smaller businesses, recently lowered guidance for the quarter.
Gartner this week presented its information technology spending projections for 2009, and it sees the souring economy taking a toll. Its worst-case projection (increasingly likely, it seems) places worldwide growth in technology spending at 2.3 percent for 2009, down from 5.8 percent previously. Stronger Asian markets like China are propping up the overall numbers. In North America, Gartner sees spending growth at half a percent in 2009, down from 5.3 percent previously.
In an interview, Peter Sondergaard, Gartner's senior vice president for global research, explained that information technology was now so integral a part of business operations that spending was no longer going to be much higher or lower than the broad economy. "This isn't like the 2000 to 2002 period, when much of the world lost confidence in I.T.," he said.
But 2009, Mr. Sondergaard suggests, will be a year of battening down for turbulent times. "There is no big thing in I.T. in 2009," he said. "There is cost reduction, risk management and compliance."
Horizons will shorten, and projects that can wait likely will. "C.I.O.'s have to learn to think like C.F.O.'s," he explained, using the common shorthand for chief information officers and chief financial officers.
At a Gartner conference in Orlando on Monday, Mr. Sondergaard offered a "top 10″ list for squeezing more out of tight budget dollars. In the spirit of the new austerity, we'll just give you the top five (the bottom five don't add much):
- Reduce headcount and freeze hiring.
- Curtail data center expansions and "virtualize" servers, putting more software loads on fewer machines.
- Renegotiate with technology and services suppliers.
- Consolidate functions and systems to achieve greater economies of scale.
- Outsource commodity services.
It's enough to warm any C.F.O.'s heart.


text:
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Shoe Styles - Boots, Etc.
The following images are examples of shoes we've made for other clients. We offer them as suggestions or jumping-off points for your shoes, rather than as a selection of styles to choose from. You may find a picture of exactly the shoe you're looking for here. More likely, you'll get an idea that you can combine with others to design your own unique shoes.
Click on any image to view a larger version.
Recently added images:
Previously added images:
|Cuffed boot with bells
|Street boot
|Dog Paws
|Fringed Western Wingtip
|X-High Lace-up
|Dalmation Dog Paws
|Western Wingtip with Appliqué
|Western Wingtip
|Pointy Cuffed Stove-pipe
|Pointy Spat Look
|Fringed Western
|Fringed Western
|Low Buckle
|Low Buckle
|Mini Lollipop Spat Look
|Mini Lollipop Spat Look
|Mini Lollipop Spat Look
|Mini Lollipop Stove-pipe
|Square-toed Spat Look
|Square-toed Spat Look
|Lollipop Fire Boot
|Lollipop Fire Boot
|Lollipop Fire Boot
|Western Boot
|Western Boot
|Western Boot Spats
|Western Boot
|Full-Length Spat Look
|X-Wide Cuffed Street Pirate Boot
|Pointy Spat Look
© Copyright 2007, all rights reserved
Spear Specialty Shoes • 12 Orlando Street • Springfield, MA 01108-2412 • (413) 732-7184 • firstname.lastname@example.com


text:
What a talent, yes indeed. 'Serenade' was beautifully penned and delivered with sincerity. Not content with melting the audience this skilful music man added rhythm and beatboxing to the end of his set, much to the thrill of on-lookers.
The Newquay Voice
©2018 Aaron Douglas Music // All Rights Reserved // Faultless Design // The King Of Pixels


text:
For many people Somalia is synonymous with violence, warlordism, famine, terrorism, jihadism and piracy. But Somalia is not an entirely lawless and ungoverned land. Somalis have used their own resources and traditions of conflict resolution to re-establish security and governance in many communities. Accord 21 seeks to improve understanding and links between Somalis and international policy and practice. Read more about this publication here.
Using a Kindle?:
Using Kobo eReader, Blackberry Playbook, Apple's iBooks, Google Books, Firefox or Adobe?:


text:
Singapore in a nutshell
On June 27, 2011 by nizamrahman With 0 Comments - Uncategorized
Life at the speed of light. More people going forwards than backwards. Space is at a premium, but construction still goes on. And the roadways are pretty difficult to navigate at first.
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text:
September 01, 2011
Borrowers' uptake of fixed interest rate home loans rose by less than one percent of approvals over August despite lenders introducing large reductions to the cost of their fixed term loans, according to Australia's largest independently-owned mortgage broker.
Fixed rates accounted for 14.1% of Mortgage Choice's home loan approvals last month, up only slightly from 13.3% in July (though rising for the third consecutive month), while the popularity of ongoing discount loans continued its steady increase, up from 38.6% to 41.5% of approvals.
Company spokesperson Kristy Sheppard said, "New borrowers' appetite for ongoing discount home loans has steamed ahead for 10 consecutive months now. We have reached a point where demand for such mortgages is more than double that for any other, at 41.5% of all our August approvals."
"This loan type, where the interest rate is discounted over the loan term usually in return for an annual fee, overtook standard variable as the favourite in April and hasn't looked back. The trend speaks volumes about new borrowers' and refinancers' mindset around interest rate rises and the value they place - or rather, don't place - on locking in their rate at the moment.
"I expected the take-up of fixed rate home loans to grow noticeably in August due to the well publicised reductions many lenders have been applying to their fixed term pricing. Despite our lender panel's average three-year fixed rate falling half a percent in the past four weeks alone, fewer than one in seven new mortgage holders fixed part or all of their rate last month.
"Borrowers' reticence to fix may also be influenced by memories of the break costs many people faced when considering switching out of their fixed terms during Spring 2008 to Autumn 2009 when the cash rate fell from 7.25% to 3.00% and home loan interest rates followed."
Demand for standard variable and basic variable home loans fell in August, to 19.2% and 18.2% of approvals, as did that for line of credit and introductory rate loans, to 4.7% and 2.4%.
Note: Mortgage Choice currently writes one in 25 new home loans in Australia, equating to approx. $10 billion in approvals per year, hence it provides a clear insight into borrower preferences. The 19 year old mortgage broker has a loan book of over $42 billion.
For further information or to arrange an interview, please contact:
Belinda Williamson
Mortgage Choice
(02) 8907 0472 or 0407 416 124


text:
- Articles
- Open Access
Kinetic Analysis of Glycoprotein–Lectin Interactions by Label-Free Internal Reflection Ellipsometry
© Humana Press 2008
- Received: 29 January 2008
- Accepted: 9 May 2008
- Published: 18 July 2008
Glycoproteomics is undergoing rapid development, largely as a result of advances in technologies for isolating glycoproteins and analyzing glycan structures. However, given the number and diversity of glycans, there is need for new technologies that can more rapidly provide differential carbohydrate–protein structural information on a large scale. We describe a new microarray platform based on a label-free imaging ellipsometry technique, which permits simultaneous detection of multiple glycoprotein–lectin interactions without the need for reporter labels, while still providing high throughput kinetic information at much lower cost. Our results demonstrate the utility of LFIRE™ (Label-Free Internal Reflection Ellipsometry) for the rapid kinetic screening of carbohydrate–lectin recognition. The technology was also used to evaluate the benefits of the lectin immobilization format using multi-lectin affinity chromatography (M-LAC) to capture glycoproteins (with enhanced binding strength or avidity) from biological samples. Using a printed panel of lectins, singly or in combination, we examined the binding characteristics of standard glycoproteins.
Results and Discussion
Using kinetic measurements, it was observed that the binding strength of lectins to carbohydrates is enhanced using a multi-lectin strategy, suggesting that improved selectivity and specificity can lead to increased functional avidity. The data presented confirm that this label-free technology can be used to effectively screen single or combinations of lectins. Furthermore, the combination of LFIRE™ and M-LAC may permit more rapid and sensitive identification of novel biomarkers based on carbohydrate changes in glycoproteins, and lead to a better understanding of the connections of glycan function in cellular mechanisms of health and disease.
- Lectins
- Glycoproteome
- Imaging ellipsometry
- Label-free
- Glycoproteins
- Internal reflection ellipsometry
- Multiplexing
- Kinetics
Glycosylation of proteins is a nearly ubiquitous post-translational modification observed in eukaryotic organisms. It is estimated that roughly half of the mammalian proteome consists of glycoproteins [1]. Glycosylation plays a fundamental role in a diverse set of biological processes such as immune response, cellular regulation, and cell signaling [2]. Alterations in glycosylation patterns have been linked to the progression of several diseases, most notably cancer [3, 4].
In recognition of the importance of glycosylation, many proteomics researchers have turned their attention to the glycoproteome in search of markers for disease diagnosis and treatment. These include candidate biomarkers that change in either the abundance of the glycoprotein or the glycan structure itself, or both. However, the study of glycosylation is a challenging undertaking; this can partly be attributed to the complexity of glycan structures. Glycans can exist in highly heterogeneous forms, with a variety of possible structural configurations and linkage types; they are present as N- or O-linked and can occupy several sites on a glycoprotein. Several methodologies are available to characterize glycostructures, such as those based on anion exchange separation of fluorescently tagged glycans [5], mass spectrometry using nano-LC coupled to a linear ion trap Fourier transform mass spectrometer (nano-LC-LTQ/FTMS) [6], and MALDI-TOF [7]. In addition, carbohydrate–protein interactions have typically been studied using kinetic measurements by surface plasmon resonance (SPR) [8, 9] and frontal affinity chromatography using lectins [10]. Previously, methods focused on carbohydrate arrays, where the low-affinity lectin–carbohydrate interactions are much simpler and easier to model but do not allow researchers to directly examine changes in glycosylation of proteins [11]. In the past few years, lectin microarrays have become increasingly popular. Kuno et. al. reported using a new microarray procedure based on evanescent-field-excited fluorescence detection of lectin microarrays as a strategy for glycan profiling to obtain kinetic information in a multiplexed fashion [12]. Technologies capable of analyzing multiple lectin–glycoprotein interactions in parallel in a single experiment and in a high-throughput manner offer increased utility, as reported previously [8, 11, 12]. Such technologies can be utilized to efficiently study disease-associated glycosylation changes on a proteome scale to further accelerate the quest for "glyco-markers" of clinical value.
Lectins comprise a special family of carbohydrate-binding proteins distributed widely in the plant and animal kingdom. After a boom in lectin discovery and characterization in the 1970s, many lectins became commercially available for research purposes [13]. Lectins have been essential for the understanding of carbohydrate-based biological recognition. The use of lectin affinity chromatography has gained popularity in glycoproteomics as a way to simplify the sample before analysis, to enrich specific glycoprotein content in blood samples. In addition, the differential specificity of lectins allows specific populations of glycoproteins to be targeted and may permit changes in glycosylation to be detected. The wide variety of lectins available commercially enables many different subsets of glycans to be investigated. Technologies such as the multi-lectin affinity chromatography (M-LAC) [14] or serial lectin affinity chromatography (S-LAC) [15] have been both used for interrogating the glycoproteomes of human plasma, serum, urine, and cellular lysates. Alternatively, lectin or carbohydrate arrays have also been introduced to study changes in the glycoproteome. A significant benefit of array technologies is that they require very little sample, and a wide spectrum of ligand interactions may be screened rapidly in parallel format.
We (W.H., M.H., M.K.) developed the concept of M-LAC or multi-lectin affinity chromatography as a way of improving the specificity/selectivity of glycoproteins isolated from complex biological specimens. We have demonstrated previously [16, 17] that the M-LAC column has excellent enrichment capabilities for glycoproteins from human plasma or serum. This technology provides an approach for detecting up or down regulation of glycosylated and non-glycosylated proteins from the same sample. The advantage of M-LAC over single or serial lectin capture may lie in its more complete single-step capture and enrichment of low level glycoproteins from complex samples, such as serum or plasma. We also have demonstrated that M-LAC can be used to monitor changes in protein glycosylation. This is achieved by step-wise elution of glycoproteins using three different sugar displacers specific for each lectin in the M-LAC column. For example, it has been demonstrated that the treatment of transferrin with neuraminidase to remove sialic acid causes a shift in distribution of transferrin isoforms between fractions eluted with three different displacers [18].
The combination of specificities of the lectins as found in the M-LAC column potentially can lead to a functional advantage due to multi-valency and an increase in binding strength over single lectins. It would be advantageous, however, to have a procedure for efficiently optimizing the type and ratio of lectins used in a specific sample set. In this report, we used a prototype imaging instrument (LFIRE™) based on the principles of ellipsometry, a widely used and versatile optical technique for measuring the thickness and optical parameters of ultra-thin films, which can be calibrated directly to the amount of material bound on a wide range of surfaces. The theory of ellipsometry is discussed extensively [19, 20] and will not be addressed in detail here. Although the underlying theory of ellipsometry has been understood for over a century [21], it was not until the 1960s that ellipsometers were used routinely in the early computer industry to characterize the growth of thin metallic and semi-conductor films. In the life sciences, there were a few pioneering works using ellipsometry for immunological studies and for measuring the kinetics of protein adsorption. A review of these works from 1972 [22] demonstrated the use of ellipsometry in only a small number of applications. Over a decade later, some of the first quantitative comparisons, between ellipsometry and radiolabeling techniques, of the surface mass density of adsorbed layers of proteins were demonstrated [23]. During the 1990s, imaging ellipsometry and imaging SPR had been developed and used to study the film thickness of biological layers [24–28]. Whereas imaging SPR focuses on reflected light intensity, LFIRE™ is characterized by the rapid time-resolved detection of polarization changes of a reflected monochromatic light beam due to interactions with biomolecules near a glass–liquid interface. The resultant measured phase shift between s-polarized and p-polarized light is more sensitive to adsorbed protein layers than the reflected light intensity signal from imaging SPR. In addition, SPR imaging requires a metallic layer such as gold, which limits the spatial resolution of images due to the spatial extent of the excited surface plasmons [29]. The LFIRE™ measurements are performed in situ with high spatial resolution, so that the changes in thickness and refractive index due to specific binding events within the array may be measured. Data is acquired several times per minute so kinetics may be determined for a high number of targets in parallel in both fluid cell and well-plate formats.
The LFIRE™ technology was used as a proof of concept to investigate the interaction and kinetics of standard glycoproteins with single and combinations of lectins typically used in the M-LAC format. It was determined that this approach could efficiently measure the binding kinetics of both single and combinations of lectins. The ability to perform multiplexed real-time measurement of binding kinetics adds a new dimension to the field of glycomics by allowing the analyst to probe multiple samples and combinations of lectins to interrogate carbohydrate–lectin interactions of clinical relevance. Post-translational changes in the carbohydrate structure resulting in either trimming or addition of sugars will affect the binding kinetics, a measurement that can be further used to identify and characterize new biomarkers for clinical applications. This will have broad impact in the study of diseases such as cancer, where surface, transport, and secreted proteins all show significant changes in glycosylation.
In this work, we describe the development of a new technology platform (LFIRE™) based on the principles of ellipsometry. We applied it to lectin microarrays and demonstrate that this technology can be a useful tool for quantitative and qualitative analysis of complex kinetic interactions between various combinations of lectins and different carbohydrate motifs. Furthermore, the combination of M-LAC and LFIRE™ technologies can be very powerful for high throughput ligand selection and/or characterization of glycoprotein biomarkers that can be isolated from patients with defined diseases.
Materials and Methods
Lectin Array Construction
Lectins concanavalin A (ConA), wheat germ agglutinin (WGA), and jacalin supplied by Vector Labs (Burlingame, CA, USA) were reconstituted in M-LAC binding buffer (25 mM Tris, pH 7.4, 0.5 M sodium chloride, 1 mM MnCl2, 1 mM CaCl2, and 0.05% sodium azide) for storage. To remove Tris, the samples were dialyzed into 50 mM 4-2-hydroxyethyl-1-piperazineethanesulfonic acid-buffered saline, 0.005% Tween 20 (HBST), supplemented with 1 mM manganese chloride and calcium chloride immediately before printing samples. All lectins were diluted to 0.667 mg protein per milliliter in phosphate-buffered saline (PBS), pH 7.5, for printing on high-sensitivity LFIRE™ optical slide substrates, which include a proprietary optical coating (Maven Biotechnologies) under a custom polyethylene glycol (PEG)-based hydrogel N-hydroxysuccinimide ester (NHS)-terminated surface chemistry (Accelr8 Technology Corporation, Denver, CO, USA). Mono-amine-terminated 5-kDa PEG was obtained from Nektar Therapeutics, Huntsville, AL, USA. Porcine thyroglobulin, hen egg ovalbumin, bovine fetuin, and asialofetuin glycoproteins were supplied by Sigma-Aldrich, St. Louis, MO. With a printing program developed by Maven, the SpotBot MicroArrayer, and a single Stealth946 pin (Telechem Inc., Sunnyvale, CA, USA), each slide was printed with two identical arrays of 20 materials including lectins and reference spots, four replicates each, 250 μm center-to-center, at 50–60% relative humidity, 25–27°C, yielding ∼100-μm-diameter spots. A single pin loading for all slides in each batch reduced variation and allowed maximum flexibility in spot location. Microarrays were immediately packaged in moisture-proof bags with silica gel desiccant and stored at 4°C until use.
LFIRE™ Detection System
Experimental Setup
Just before use, lectin microarray slides were washed with an aggressive spray of PBS-T (0.05 M PBS with 0.05% Tween 20) to remove all loosely bound material, rinsed briefly with distilled water, and dried with a dry nitrogen stream. Slides were attached to LFIRE™ disposable twin flow cells and microfluidic interconnect assembly (Fig. 1), optically mated to a BK7 prism with index-matching fluid, enclosed in the LFIRE™ TEC environmental control fixture, and inserted in the LFIRE system. The flow cells were filled with M-LAC binding buffer, temperature set to 25 ± 0.05°C, and equilibration was obtained before data acquisition. The signal stabilized rapidly in 15 min due to hydrolysis and quenching of NHS moieties on the surface by the buffer.
To test the surface coating supplier's claim that the surfaces may not require blocking, 50 μg/ml human serum albumin (HSA) was injected; flow was stopped and allowed to incubate for 30 min. HSA did not bind significantly (<1% of a monolayer) to the substrate, lectins, or controls (data not shown). Measuring molecular binding at the sensitivity limits of the instrument requires blocking, normalization to negative control spots such as HSA, or both for this particular surface given that, even within the small panel of glycoproteins, nonspecific binding was observed to the bare substrate but not to the negative control spots.
Lectin/Glycoprotein Kinetic Assay
The measurement area was set to encompass both arrays, each in its own flow cell, for simultaneous analysis of two glycoproteins. The running buffer for all experiments was M-LAC binding buffer flowing at 25 ul/min. All data was captured as 16-bit TIFFs. After 3 min of lead-in time to establish the baseline, 10 μM glycoprotein samples in the same buffer were injected at a flow rate of 25 μl/min, for a net incubation (dwell) time of 4 min. The arrays were rinsed continuously with running buffer only for 15 min to determine off-rates. The final measurements constituted the arbitrary "endpoints" for this study, although most samples still demonstrated some dissociation. All of the glycoproteins except ovalbumin were eluted from the arrays by a 30-min exposure to 20 mM Tris, 0.5 M NaCl, 0.17 M methyl-a-d-mannopyranoside, 0.17 M N-acetyl-glucosamine, and 0.27 M galactose, pH 7.4. Analysis of the images was done using ImageJ v1.34 s (NIH/public domain), and IgorPro (WaveMetrics, Inc, Lake Oswego, OR, USA) was used for data analysis.
Binding of Thyroglobulin on ConA and M-LAC Columns
Lectins were immobilized onto styrene–divinylbenzene co-polymers coated with a cross-linked polyhydroxylated polymer (POROS™). HPLC support using the Schiff base method for covalent immobilization (aldehyde activated media, Applied Biosystems) was used according to the instructions provided by the manufacturer. HPLC columns were packed with either the immobilized Con A or with a mixture of the three individual lectins (ConA, WGA, and JAC) combined in a 1:1:1 (23:8.0:11 mg) molar equivalent ratio to make the M-LAC column. The amount of Con A in the single lectin affinity column was the same as that of the M-LAC support.
Thyroglobulin protein was prepared by dissolving 20 mg in 2 ml of M-LAC binding buffer for a final concentration of 10 mg/ml; 5 mg of the thyroglobulin solution was injected onto each of the ConA and M-LAC columns. To increase the residency time, the protein was loaded slowly (0.05 ml/min). The flow rate was then increased to 2.5 ml/min. Unbound protein was washed with 10 column volumes (CV) of binding buffer, and the bound protein was eluted with 11 CV of 100 mM acetic acid at pH 4.0. The lectin columns were neutralized with 11 CV of neutralization buffer (0.5 M Tris, 1 M sodium chloride, and 0.05% sodium azide, pH 7.4) and equilibrated with 10 CV of binding buffer.
Results and Discussion
Lectin microarray printing map, with each lectin or reference spot printed in quadruplicate
Column 1
Column 2
Human serum albumin
Protein A/G
5 kDa amino-PEG
ConA/WGA/Jac 0.11:0.22:0.33
ConA/WGA/Jac 0.23:0.29:0:0.17
ConA/WGA/Jac 0.34:0.12:0.17
HSA/ConA/WGA/Jac 0.23:0.29:0:0.17
HAS/ConA/WGA/Jac 6:1:1:1
Human serum albumin
The kinetic measurements are shown in Fig. 7d and were performed under identical experimental conditions as the data shown in Fig. 5. The binding affinity of thyroglobulin for the mixture of lectins \(\left( {K_a = 2.1 \times 10^6 {\text{M}}^{{\text{ - 1}}} } \right)\) was found to be about six and 15 times higher over ConA (3.4 × 105 M−1) and WGA (1.4 × 105 M−1), respectively. Although it is not the scope of this paper to define carbohydrate–lectin multivalent biochemistry, kinetic analysis using multiple and single lectins can greatly aid in the selection of lectin combinations to enrich low-level glycoprotein markers in complex clinical samples such as plasma or serum. In ongoing further studies, we are using LFIRE™ imaging ellipsometry as a complementary measurement tool to multi-lectin affinity chromatography for glycoprotein analyses. The combination of both technologies can be used to reveal subtle differences in glycan-binding specificities of closely related lectins or lectin combinations. For this purpose, the microarray format is particularly suitable for studying a single probe's interaction with a large number of targets. Hence, this methodology permits detection of the binding of a specific glycoprotein to a panel of lectins in a multiplex format to monitor subtle changes in glycosylation associated with disease. This can be very relevant in the field of clinical glycoproteomics for the identification and/or validation of "glyco-markers" in diseases such as cancer, which are typically associated with alterations in the amount and type of glycan content. Future experiments are planned to study the structure–function relationships of carbohydrate moieties especially in the discovery of novel clinically relevant biomarkers.
In conclusion, the data suggests that LFIRE™ technology can be a useful tool for quantitative and qualitative analysis of complex kinetic interactions between various combinations of lectins and glycoproteins. Moreover, by obviating the need for labels, analyses can be performed more economically and in parallel using lectin microarrays. For discovery of potentially significant new biomarkers, the powerful combination of M-LAC and LFIRE™ offers the possibility of analyzing lectin–glycoprotein "fingerprints" or patterns associated with disease occurrence, stage, and/or response to drugs.
Some of the authors (Kullolli, Hancock, and Hincapie) wish to acknowledge support from the National Cancer Institute for grant U01 CA128427-01, "Glycan Markers for the Early Detection of Breast Cancer."
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By Mya Gosling
THE PLOT SO FAR: Cesario (Viola) has just left Olivia, whom he (she) was wooing on behalf of Orsino. Little does he (she) realize that his (her) proxy wooing has been more effective than he (she) thought…
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Mastering 802.11n: Implementation tips and answers to frequently-asked WLAN questions
802.11n promises to let us build bigger, faster wireless networks that deliver better reliability and capacity. But tapping this technology's enormous potential will require new products, careful planning -- and a bit of ingenuity. In this podcast, we discuss innovative approaches that you can use to overcome the most common 802.11n deployment barriers, from the pinch of Power over Ethernet to bandwidth bottlenecks. In this podcast, speaker Lisa Phifer shares her advice on how to avoid and troubleshoot common WLAN issues.
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802.11n promises to let us build bigger, faster wireless networks that deliver better reliability and capacity. But tapping this technology's enormous potential will require new products, careful planning -- and a bit of ingenuity. In this podcast, we discuss innovative approaches that you can use to overcome the most common 802.11n deployment barriers, from the pinch of Power over Ethernet to bandwidth bottlenecks.
In the podcast, speaker Lisa Phifer answers the following questions and shares her advice on how to avoid common 802.11n wireless LAN (WLAN) issues. Find more advice from Lisa Phifer and other wireless networking experts by navigating to the links below:
1) Enterprise adoption of 802.11n has been impeded by interoperability concerns. How can companies avoid running into interoperability problems?[Answer is 1:00 minute in podcast.]
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Few of us have the luxury of deploying a new 802.11n WLAN from scratch. Most will need to coexist with legacy 802.11a/g devices -- from existing APs and clients to devices owned by neighbors and guests. To accomplish this, consider these questions...
2) A big concern that many companies have about 802.11n is increased power consumption. Can 802.11n be deployed without costly Power-over-Ethernet upgrades? [Answer is 3:20 minutes in podcast.]
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Before deploying 802.11n Wi-Fi technology, organizations must determine whether their vendor of choice has developed access points that can run on electricity supplied by Power over Ethernet (PoE).
We're planning to deploy 802.11n later this year. I've heard there will be power supply problems. Can you explain what the issue is and how we can solve it?
3) Some businesses are worried about overloading their networks with high-throughput wireless traffic. Will 802.11n really create bandwidth bottlenecks and how can they be avoided? [Answer is 6:03 minutes in podcast.]
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New draft 802.11n access points (APs) use multiple antennas to greatly increase network footprint, available bandwidth and resilience to problems that crippled older 802.11a/b/g APs.
I have to create a wireless network in a large open building. What would you do to boost signal strength? I found antennas that improve range, but I don't see how they can be connected as my router seems to have a fixed antenna that cannot be removed. What products work with these better antennas?
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4) Most companies that will deploy 802.11n already have 802.11b/g wireless. How can administrators minimize impact on legacy clients during an 802.11n upgrade? [Answer is 8:22 minutes in podcast.]
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What is the impact of having both the 802.11g and 802.11n access points in the same environment? Will this cause conflicts for the clients in this environment?
5) Airwave congestion and co-channel interference are big challenges for today's b/g networks; 802.11n will add even more traffic to those crowded airwaves. Do you have any tips for overcoming channel competition? [Answer is 10:06 minutes in podcast.]
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Learn how Wi-Fi Multi Media (also known as Wireless Multimedia Extensions), a subset of IEEE 802.11e MAC Enhancements for Quality of Service (QoS), can be used to extend traffic prioritization to the wireless edge.
Every AP is limited by number and type of radios and channels used. Channels are shared resources, so the sum of all traffic sent and received on that frequency cannot exceed that channel's capacity. Aggregate application throughput is further diminished by 802.11 protocol overhead, client competition, and loss/retransmission due to distance and interference.
6) During any major network upgrade, unexpected problems and costs are always a concern. What's the best way to avoid surprises when deploying 802.11n? [Answer is 12:12 minutes in podcast.]
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The question now facing most companies is not whether to migrate to 802.11n, but where, how and when to do so. This checklist will point out critical factors that should be considered when planning your 802.11n upgrade in order to maximize benefit and minimize pain.
WLAN protocol analyzers can easily identify common WLAN problems like co-channel interference and show the impact that non-802.11 transmissions have on Wi-Fi associations and applications. But if you need to find out what's causing non-802.11 interference, you'll need to use a spectrum analyzer.
About the speaker:
Lisa Phifer owns Core Competence Inc., a consulting firm specializing in leading-edge network technology. She has been involved in the design, implementation and evaluation of networking and security products for over 25 years. Before joining Core Competence, Phifer was a member of the technical staff at Bell Communications Research, where she won a president's award for her work on ATM network management. Phifer teaches about wireless LANs, mobile security, NAC, and VPNs at many industry conferences and webinars. She has written extensively about network infrastructure and security technologies for numerous publications.


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While most celebrate the day after Thanksgiving by trampling over each other for the best deals on this shopping holiday; "Black Friday," two of hip-hop's biggest names decided to surprise their fans (and the internet as a whole) with free music. Kendrick Lamar rapped over J. Cole's "A Tale of 2 Citiez" and J. Cole rapped over Kendrick Lamar's "Alright" to further add to the hype of a possible collaborative project by the two MCs.
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Kohl's Corp.'s e-commerce growth fell below its expectations, but was still in double digits, CEO Kevin Mansell and other company executives told Wall Street analysts last week on the Kohl's first quarter 2014 earnings call. The company did not give figures on its e-commerce sales.
Total sales in Q1 2014 were $4.070 billion, a decrease of 3.1% from $4.199 billion in the first quarter a year ago.
The company says it plans to invest significantly in mobile and in-store technology, including its loyalty program and push notifications for consumers in store.
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Another e-commerce investment is in the retailer's ship-from-store program, which it plans to expand to 800 stores from 200 now. The company expects that to happen in the third quarter. Kohl's buy online, pick up in store test will expand to select markets in the third quarter as well, including Chicago, Milwaukee and San Diego.
For the first quarter, the company didn't disclose e-commerce sales but did report:
Kohl's is No. 23 in the Internet Retailer 2014 Top 500 Guide.