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A proposed greenbelt development that would see a crumbling building converted into a house will be decided by a Government inspector.
Last year Bradford Council refused plans to convert the Gatehouse at Upper Shibden Hall, off Halifax Road on the Calderdale boundary at Catherine Slack, into a residential scheme.
The scheme would have involved an extension to the building, transforming it into a three-bedroom house and increasing the size of the building by 50 per cent. There would also be new car parking spaces created.
Council planning officers said the development was ""disproportionate" and "harmful" to the openness of the greenbelt area.
Applicants The Shibden Estate Company, has now appealed that decision, meaning a Government-appointed inspector will decide whether the development can go ahead.
The appeal says: "The building is not Listed but clearly has historic merit and as a heritage asset it should be protected for the future.
"The restoration of the original building emphasises the historic character of the building and the extension represents a modern addition to bring the building back into economic use.
"Any scheme does require a reasonable amount of living accommodation to be provided in order to make the restoration viable and the proposal is an extension which is designed to do this without looking disproportionate."

subset: commoncrawl


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Download the GardenTags App today and join 100,000+ gardeners getting FREE plant identification, garden management and more!
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🐝🐞🍎🍒🍐🍓🥝🥕 Northwestern USA zone 5b-ish maybe 6 #pnw #oregon #native #plantnatives
Found the trellis now. Hurray! 9.19.18 #passionflower
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Thank you neighbor! @ShelleySnyder
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Glad you got it added Shelley. But, I don't know for sure about your question. Was there a link or any options? @ShelleySnyder
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Gorgeous! Is this on the one you bought a little while ago Laura?
Tiny, broken #passionflower. It's actually a great spot. Might do really well. Interplanting with yellow climbing rose.
Oh my that is mighty tiny!💘😍
Passion flowers are usually quick growers aren't they do it won't look like that for long 😊
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U.S. asked to probe test of unapproved implants on inmates
This Aug. 2, 2018, file photo shows the U.S. Food and Drug Administration building behind FDA logos at a bus stop on the agency's campus in Silver Spring, Md. (AP Photo/Jacquelyn Martin, File)
BATON ROUGE, La. -- The U.S. Department of Food and Drug Administration is being called to investigate a short-lived pilot program that sought to treat inmates with anti-addiction implants never approved by federal regulators.
The Advocate reports advocacy group Public Citizen requested the probe in a letter Wednesday. The pilot by the Louisiana Department of Public Safety and Corrections and California-based creator BioCorRx, Inc. involved a surgical implant that released the opioid and addiction fighting drug naltrexone.
Officials said the chemical would be metabolized over several months. The FDA already approved naltrexone as daily pills and monthly shots.
BioCorRx offered the corrections department 10 free implants for volunteer recipients. For unclear reasons, the pilot stopped after the first implant. Recipient 39-year-old Alvin Dutruch says he has been doing well in recovery since his release.


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Leading through Facebook
It fascinates me where leaders decide to connect. We've all heard about Facebook's CEO Mark Zuckerberg announcement on his Facebook page of his gift to the world in honor of his new daughter's arrival. In case you have not heard, he and his wife are giving 99% percent of their Facebook shares while they are alive to charitable purposes.
Recently I was told about the Secretary of Defense Ash Carter's Facebook page. It is worth looking at as an example of a leader demonstrating his humanity, courage and connectedness to communities and our troops. The reason I say courage is leaders have to have a thick skin to open themselves up and be fully transparent. Everyone has an opinion.
I was not sure what a Secretary of Defense does exactly but Wikipedia set me straight. Clearly, a job not for the faint-hearted. Essentially, he is the CEO of the Department of Defense. "The Secretary of Defense is in the chain of command and exercises command and control, subject only to the orders of the President, over all Department of Defense forces (the Army, Navy, Air Force and Marine Corps. Also the United States Coast Guard during any time it is transferred to the Department of Defense) for both operational and administrative purposes."
You get some sense of what Carter does most days — giving speeches, visiting veterans, commenting on current events like the Paris attacks, visiting old haunts like Boston, memories of serving service members abroad on Thanksgiving — and the people he gets to see and make time for. You get to hear about how he views the world and what's important to him. During his visit to Boston, he wrote, "This visit follows recent trips to Silicon Valley as part of my effort to break outside the 'five-sided box' and connect our Department with the important advancements in technology happening all across this country." I was not sure what he meant by the five sided box comment but I presume it is the Pentagon. Breaking out of the box sounds just right to me when it comes to leadership.
This week was a particularly special week because he opened up all jobs in the military to women ("there will be no exceptions"). Comments galore. He wrote:
"When I became Secretary of Defense, I made a commitment to building America's force of the future – the all-volunteer military that will defend our nation for generations to come. Like our outstanding force of today, our force of the future must continue to benefit from the best people America has to offer. In the 21st century, that requires drawing strength from the broadest possible pool of talent. This includes women, because they make up over 50 percent of America's population. To succeed in our mission of national defense, we cannot afford to cut ourselves off from half the country's talents and skills. We have to take full advantage of every individual who can meet our standards.
That is why today I announced my decision to open all military service occupations and positions to women. There will be no exceptions. This means that as long as they qualify and meet the standards, women will now be able to contribute to our mission in ways they never could before. Today, we have the finest fighting force the world has ever known. With this decision, we take another step toward that continued excellence."
It is not easy being a public figure — business, government or otherwise. But if you want to show your human side, listen to others, publicly engage, and connect, Facebook provides that opportunity. We could all dare to listen more to others and round out our reputations.


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In-flight Internet company Gogo announced that three airlines – American Airlines, Delta Air Lines, and Gol Linhas Aéreas – have gone live with the company's Gogo TV live television offering.
Gogo TV delivers live TV programs to passengers' personal electronic devices such as tablets or smartphones and is also integrated into the seatback entertainment system.
American Airlines was Gogo's first customer for in-flight Internet service when it went live in 2007.
Gogo told Frequent Business Traveler that a fourth airline, Japan Airlines, as well as its subsidiary, Japan Transocean Air, will go live with Gogo TV in October.
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Transport and Logistics in Kenya
Transport and Logistics in Kenya (Master in Business in Africa)
Kenya (East Africa) is the largest East African logistics centre
Maritime transport in Kenya
The Port of Mombasa is the largest East-African port
Access to:
Lake Victoria
Kenyan Roads
Lagos-Mombasa Corridor (6,260 kilometres)
Cairo-Gaborone Corridor (8,861 kilometres)
Railways in Kenya
Airports in Kenya
More information: Business in Kenya, at EENI Business School & HA University Training Portal.
Transporte en Kenia Transport au Kenya Transporte no Quénia
(c) EENI Business School & HA University (Africa) . We do not use cookies


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Regardless of how clean you keep your Knoxville home, germs can linger in some unexpected places. In this Knoxville Home How To, we explore the top 5 most germ-infested items in your Knoxville home and how… read more →
Everyone Knoxville homeowner has one room in their home that is small and difficult to decorate. Whether you're in process of getting your Knoxville home ready to sell or just in the midst of redecorating, here… read more →
According to the most recent additions of The 2014 Old Farmer's Almanac, this winter will be one of the worst in recent history, with record low temperatures and above normal snow fall. If the weather in… read more →


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Directions to Landmark at Tanasbourne
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Yasif Ahmad Faysal teaches English at the University of Barisal.


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The 'heart' of bluegrass music, banjo is a popular instrument which is used in traditional Irish and country music genres. It is a guitar-like string instrument which is popularly used for playing cheerful songs. Following is a collection of articles that acquaint you with facts about how to string, tune, and play banjo - 'the happy instrument'.
How to Play the Banjo
Learning to play the banjo is just a process of understanding the basics of the instrument and putting a substantial amount of practicing time. Following is a guide on how to play the banjo for beginners...
How to Tune a Banjo
The process of knowing how to tune a banjo is similar to that of tuning an acoustic guitar. This article will tell you more on how to tune a 5-stringed banjo...
Top 10 Banjo Manufacturers in the World
It was the African-Americans who introduced the banjo in America. Having started off as an instrument that had a gourd body and stick neck, the design of the banjo has come a long way in terms of design, tone, and significance....
How to Buy a Banjo for Beginners
So, you have decided to learn how to play a banjo but, do not know what to look for while buying your first one? Here is an article telling you, how to buy a banjo for beginners.


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My main focus as a photographer is to capture YOU, exactly the way you are today, in this moment. Whether you are short, tall, blonde, brunette, married, single or anything in between, be that. My goal is not to make you become someone else. My goal is to have you trust that being YOU is what makes you beautiful.
The main focus of my life is to positively influence those around me. The perfect version of me would treat my family like Jack Pearson treats his, yet have a little bit of Beth's sass (This is Us anyone?). Donuts and discounts are my Kryptonite. I love traveling, with or without my family, but always with my camera in hand. I dance in the car. And around the house. I sing at the top of my lungs at least once a day, typically made up songs as if my life was a musical. I tell myself my family loves when it happens. I grew up in Wisconsin and will always love the Packers. Sundays are for church, but every day of the week is for Jesus (I hope we can still be friends even if you don't feel the same way. We are all humans after all right?). God has given me an amazing life. Some days it seems perfect, and other days I want to crawl into a dark corner of my house and never emerge. I'm getting better at sharing the harder days because nobody wants to watch a perfect life on social media anyway (and, duh, nobody's life is perfect so stop comparing yours to lives on the Internet. We all cry, have bad days, struggle, laugh, and try again. It's up to you which of those you choose to see in others and see in yourself.)
Whether we're hanging out, you've hired me to photograph your life, or you want me to speak at your next event, I promise that we will laugh together, cry a little (both tears of joy and out of love), and if you are lucky we might even dance.
Above all I hope I can show you just how beautiful your life is with you in it, just the way you are.


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12 Powerful Medical Documentaries You Should Watch
As a medical professional, it's essential to stay up to ...
Call Now 1300 764 482
Digital and mobile technologies are bringing huge benefits to the healthcare sector. By embracing these new technologies, healthcare organisations can improve patient health outcomes, be cost-effective, and provide timely care. Patients will also be able to better manage and control their health and medical information. This shows that technology can help ensure a brighter, healthier future for everyone.
So what are the latest trends in healthcare technology you should watch out for this year? Read on.
Artificial Intelligence (AI) and Machine Learning (MI) speed up the processing of healthcare data much faster than a human can and they do it more efficiently too. AI and ML algorithms are also used to identify and diagnose health problems like gut disorders and blindness or vision impairment and to interpret test results. Additionally, AI is used to analyse information from a patient's Electronic Health Record (EHR) in order to more accurately diagnose their health problem and come up with various ways to treat it. What's more, AI and ML can be applied in pathology and radiology to detect irregularities in MRI scans and X-rays.
The Internet of Medical Things (IoMT) is an infrastructure consisting of connected medical devices, sensors, software applications, and healthcare IT systems that focus on medical testing, monitoring, and diagnostics.
IoT-enabled medical devices include sticking plasters that monitor your body temperature, blood pressure and heart rate, and wearable devices like smartwatches and smartphones with health-related apps that track whether you're sleeping, healthy or taking your pills, and sends alerts to your doctor when there are problems. Portable biosensors are used to analyse patient data for diagnosing health problems, while ingested or implanted sensors can discreetly measure temperature and transmit this data to your doctor. IoMT can contribute to better health outcomes.
Augmented Reality (AR) doesn't lose touch with our realness. When you combine AR and AI together, healthcare apps can be extremely beneficial to both doctors and patients. You can turn your smartphone camera on your body and check the location of your digestive organs, skeleton, cardiovascular and respiratory systems, and so on.
In medical practice, AR can be used to help improve safety, administer injections, and assist in the performance of major operations. AR can also be used to make certain tasks more efficient, such as superimposing patient records and vital signs in real time while a doctor is assessing a patient.
Like AR, Virtual Reality (VR) can also help with training clinicians through simulation, educating patients, and aiding with treatment. For instance, Cool! VR Pain Relief uses a virtual world of landscapes and changing seasons to distract a patient from their pain and offer relief.
An Electronic Health Record (EHR) allows the patient and their doctor to have permanent access to patient data, eg diagnostics, chronic diseases, cardiac waveforms, and previous prescriptions. This can help improve the interaction between the doctor and patient for more productive delivery of care. An EHR can also reduce the number of papers you have to bring with you when visiting your doctor. Moreover, various healthcare providers can now access a patient's medical information through a shared EHR, which is supported by a mobile app.
The huge volume of health data generated from wearables and sensors has led to a rise in new challenges like interoperability, data integrity, security, and privacy. Blockchain can help solve these challenges by placing the patient in the middle of the healthcare ecosystem and, in turn, this will help the patient to gain more control over their health data. When it comes to EHR's, blockchain can help prevent patient data from being changed or stolen by using a singular secure protocol.
Health-tracking apps can track blood pressure, heart rate, sleep period, distance covered, and number of steps taken with accuracy. They can also let you see the data in a readable form, save the data, do a statistical analysis of the data, compare your results with standard results, and provide advice for improving your health results.
Therapeutic apps, also known as digiceuticals, help to improve health and wellness. They've been tested for efficacy by regulatory bodies like the Therapeutic Goods Administration (TGA). Therapeutic apps are prescribed by medical practitioners together with normal methods of treatment and are clinically validated for their purpose. They can be used to help people with chronic pain, asthma, or attention-deficit hyperactivity disorder (ADHD).
Telehealth puts the online doctor in your pocket. With a smartphone and app, you can search for doctors online and compare their stats, skills, qualifications, and patient reviews before you choose a doctor to consult with. Once you've found the right doctor, you can consult with them using chat or video call. Telehealth is all about virtual care and it can be very useful for rural and remote patients who can't partake in a face-to-face consultation with a doctor or specialist.
A smart hospital relies on a connected infrastructure of smart medical devices for the purpose of improving existing patient care procedures and introducing new processes. The aim of a smart hospital is to transform patient data into insight and then act on that insight. For example, it'll collect data, use AI and ML to analyse the data, and then make these insights available to clinicians and other stakeholders through several devices like desktops, tablets, and smartphones.
Robotics are used in hospitals and aged care facilities to perform a number of tasks, such as:
Robotics also has the potential to help in other areas of healthcare and to diagnose various health conditions.
As you can see, these technology trends are set to revolutionise healthcare this year and beyond. You can expect superior, safer, and quality healthcare services to be provided to patients, and a patient's own health and wellbeing will be more closely accessible to them. You may even see autonomous vehicles like drones delivering medication or self-driving ambulances bringing patients to a hospital in the near future. Whatever developments are made, technology certainly has the power to transform medicine and health for the better.
SyberScribe's state-of-the-art medical transcription service is made possible by technology. You can use a digital recorder and an Internet-connected PC to access our hosted dictation and transcription platform and our transcriptionists will then turn your reports around in a timely manner and file them securely back into your management system.
So if you're ready to make the most of technology in your healthcare organisation, contact us to find out more about our medical transcription services.


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Incense holders resin with a statue depicting The buddha in meditation, the zen-guaranteed !
A door incense nicely decorated, and efficient to pick up easily the ash after burning a stick.
Material : resin. Handcrafted in Bali (Indonesia). Colour random (gold or silver).
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I've never travelled. Locked up in that idea I went halfway around the world, I saw a single landscape, I visited only one country, I met only three men, a dog, a pig and a monkey, just one woman and dozens of animals and trees. How many trees have there been in your life?, I use to ask myself when I lose hope. Trees don't like us that much. I don't think to them as friends but they help me to get hope back. I mean, who wants friends with such enemies!
I've never studied. Damned for a sin from which I could neither learn anything nor ask for forgiveness, I approved everything until God stopped believing me. Later, science ceased to understand me, money stopped to needing me, democracy stopped to listening to me, nature stopped to seeking me and I do not entertain culture anymore. Despite that, I never stopped believing enough to get to write my own traps:
Trap 1. If you think it's a game, you'll learn how to obey. Trap 2. If you do not believe, you'll write it. Trap 3. If you believe, you'll never hear yes again.
Reading books is the promise of paradise. To read books is to steal to the wounded child you were the opportunity to explain himself. To read books is the grown up form of being humiliate in order to be happy. If you refuse to read books you end up having the trees as your enemies which is the most obvious way to reach the earth. Education is sacred and that is why it shouldn't be in the hands of teachers or priests. No priest or professor who does not go to the gym has any credibility anymore. The proof of that is that they're still murderers. Read, travel, activities of priests, teachers and of all those who do accuse and seek for a shelter in that trench over there, far away from the eternal struggle.
I've never worked. Same as priests and teachers, I've never worked either and I've choosen to do what I've felt me least able to do, which is, to venerate the Father. To work is to earn the right to disobey the Father which means a resignation that is within the reach of few people. So far, I've just reached the right to not dress the habit which is the right to do not win.
I've never had a project. Too extremist to be violent, the future didn't give me any chance. In the meanwhile, I keep writing never or always to compel my past from forgetting me.
I've never been here before. I know because I do not have rights and because I can see it in the eyes of cats, children and the dead. Men and women abandoned us. Where are the human beings that promised us with? Cats, children and the dead do not seem to care too much, why should I be worried? While I'm looking for my place in the society of the waiting cats and children, paradise is widening, leaving the dead without any place.


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Get in touch today! (+65) 8300 7300 [via Call/WhatsApp]
A Quick Glance:
■ Spacious studio apartment with balcony - expect an unblocked city view!
■ In a pristine condition and comes fully furnished.
■ Tan Tock Seng Hospital and Novena Square are both in the vicinity
■ Commute effortlessly to Orchard Road or the business hub via Novena MRT station.
Soleil @ Sinaran is a 99 year leasehold apartment located at 2 Sinaran Drive, Singapore 307467, in the prestigious District 11. It is located in the heart of Novena district. It is completed in 2012 by the reputable Fraser Centrepoint Homes with a total units of 417.
Soleil is a french word "sun", as the name implies this condominium will brighten up your everyday life. Soleil's twin heights with a graceful S-shaped curve made of highly reflective glass enhances the shining beauty of this splendid luxurious architecture. Rising out of Sinaran, the twin heights of Soleil stand as testaments to the beauty and power of our sun. Prepare for a living experience that is above and beyond the norm. Living in Soleil, means living life in a new glow.
Nothing spells elegance like Soleil @ Sinaran. Lull your soul and calm your mind here. Experience a new level of indulgences here with many of its fabulous facilities for you to recharge, rejuvenate and refresh you mind, body and soul. Here you can ease away the tiredness accumulated and seek inner serenity. The wide range of facilities available include: lap pool, spa pool, gym, fitness station, tennis courts, function room, and playground. Private spa pavilions allow you to relax and enjoy the spa facilities from your very own home. There are 7 entertainment pavilions each with its own private spa pools to allow you to host your very own private pool parties. There is also a pub at the sky terrace where you can unwind after a hard day at work. The myriad of in house facilities exclusively created for residents will pamper you to the max.


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At our place, we're here for you. We provide customized personal training and unique group classes to get you fit and healthy in ways you've never imagined.
Our gym is designed to challenge you and our trainers are designed to push you. We want you to feel your body change. We want you to be able to reach your goals. We want you to get that peach booty you've always dreamed of.
If you put in the time, we will make sure you see results. We pride ourselves in helping all people change for the better -- on the inside (mentally) and the outside (with fitness & food).
Please be kind and arrive a few minutes early to your appointments so you're all set to go at your scheduled start time (if you are late, that time will not be made up at the end).
Please refrain from arriving more than 10 minutes early as you may disrupt another client's session or Fresh may not be open yet.
Please do not bring guests with you who have not signed up for classes.
Children are NOT permitted in the facility for safety & liability reasons.
Please be courteous of others.
Please refrain from wearing wet or muddy shoes into the gym. We have plenty of boot & coat space in the front room :)
Please respect the property and equipment.
Please respect your trainers time and provide 2 or more hours notice if you must cancel or wish to reschedule an appointment. If you do not provide us with said notice, your session will be forfeited and will not be rescheduled for free.
Please respect the class attendees and provide 2 or more hours notice if you must cancel your class. If you do not cancel your class prior to 2 hours, 1 class will still be deducted from your package.
All payments are final.
FYI: There are no restrooms in our gym. The facilities are located outside our gym in the Native Pride building. Please note that this is a truck stop and these are public restrooms.
Fresh Fitness & Food (business & staff), Native Pride and/or J.C. Seneca are not responsible for lost, stolen or abused items in or outside of Fresh Fitness & Food.


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Buy Kitchen Range Hoods In Baltimore, USA
With over 30 years of experience in creating quality, hand-made cast stone kitchen range hoods, Omega Mantels and Mouldings Inc. continues to be a recognized name in Baltimore as we are the premier choice of homeowners, architects and interior designers when it comes to artisan, elegant cast stone hoods. Combining aesthetics and precision, our cast stone products are made to be the breathtaking centerpiece of every kitchen. Visit: https://www.omegakitchenhoods.com/baltimore.php
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Dunno where this goes, so I'll use this thread. Mods can break it out as appropriate.
Mariano Rivera, Roy Halladay, Edgar Martinez, Mike Mussina Elected To Hall Of Fame Rivera is first ever unanimous entry.
For anyone wondering how the vote broke down, here it is below. Ex-Phils, even for a brief period of time (one season or less), in bold.
Here are the full voting results, out of 425 total votes:
Mariano Rivera – 425 (100%) Roy Halladay – 363 (85.4%)Edgar Martinez – 363 (85.4%)Mike Mussina – 326 (76.7%)Curt Schilling– 259 (60.9%)Roger Clemens – 253 (59.5%)Barry Bonds– 251 (59.1%)Larry Walker – 232 (54.6%)Omar Vizquel – 182 (42.8%)Fred McGriff – 169 (39.8%)Manny Ramirez – 97 (22.8%)Jeff Kent – 77 (18.1%)Scott Rolen –73 (17.2%)Billy Wagner – 71 (16.7%)Todd Helton – 70 (16.5%)Gary Sheffield – 58 (13.6%)Andy Pettitte – 42 (9.9%)Sammy Sosa – 36 (8.5%)Andruw Jones – 32 (7.5%)Michael Young – 9 (2.1%)Lance Berkman – 5 (1.2%)Miguel Tejada –5 (1.2%)Roy Oswalt – 4 (0.9%)Placido Polanco –2 (0.5%)Received zero votes: Rick Ankiel; Jason Bay; Freddy Garcia; Jon Garland; Travis Hafner; Ted Lilly; Derek Lowe; Darren Oliver; Juan Pierre; Vernon Wells; Kevin Youkilis
Think Halladay goes in as a Phillie or Blue Jay?
And who decides that? I would probably lean Blue Jays. 148 wins vs 55. Really only had 2 good seasons with Philly even if they were arguably his 2 best years.
Looks like next year is setting up for Schilling.
I hate to say it, but I hope Roy goes in as a Blue Jay. Yes he had a great couple of seasons in Philly but the balk of his career was spent as a Blue Jay.
Since Bob Carpenter, when he purchased the Phillies wanted to re-name the club to be the Blue Jays, maybe we can share?
Sammy Sosa falls off the ballot (under 10%) as does Andy Pettite, surprisingly. I thought the Yankee hype machine would help him more
For better or for worse. Dick Allen's next chance comes in the winter of 2020-21.
I think you stay on the ballot with at least five percent.
I don't think being a Yankee helps one's chances. Mussina barely made it, and it took him several tries. It took Gossage 15 tries. Mattingly and Bernie Williams couldn't come close.
This wasn't exactly rocket science...
Why are people afraid to start new threads for obvious topics?
Funnily enough, Lawman already had! (But not this year I guess.)
I hated him because he was a Brave when they were winning all those division titles, but I can't understand why there is so little voting for Andruw Jones. The guy was an incredible CF, won 10 gold gloves. Hit 432 HR. I mean, I know glove guys rarely make the hall, but he it 400+ homers. Gets 7.5 percent of the vote in his 2nd year on the ballot? Luckily he made the absolute last spot to make the cut to stay on the ballot, but hard to imagine he'll ever get elected, unless by the veteran's committee many years from now.
Agreed.Roy should go in as a Blue Jay. Still, he was great for us.
I look at it this way, Chase, if he makes it, isn't going in as a Dodger (albeit his situation is different from Doc's as he didn't have great years with LA).
It looks like Roy explicitly stated he wanted to go in as a Blue Jay. Nice article on the Athletic about Halladay:
"In 2016, he said that if the Hall of Fame happened to call, he would tell them to fit him with a Blue Jays' cap when they forged his bronze plaque."
Agreed on Halladay going in as a Blue Jay. I think the more intriguing argument is whether Mussina goes in as a Yankee or Oriole. 18 year career, established himself as a star during his first 10 years with Baltimore, then followed it up with 8 good years as a NYY and had post season success as well.
I think I give the nod to Baltimore though.
Although he is a Yankee, it is nice to see there was finally a "unanimous" choice. There should have been 10 but there was always some sports writer that thought he/she would be a smart ass and stop it from being unanimous. The people that did this should have lost their voting rights when they said things like "he deserves to go in, I just didnt want to give him the vote".
Rivera deserves it and deserves to be unanimous. So did Schmidt, Morgan, Nolan Ryan, etc.
Mussina is a guy I would not of thought of really, but when you look at his career stats, he deserves it. He was always more of a steady very good performer than a great player, but honestly so was Hank Aaron and no one would argue with him going in.
Any 5 time all star that has 18 solid seasons will probably get enough stats together to be considered.
Roy is going in with a blank cap.
Blue Jays fans are understandably upset. I think he probably should have gone in as a Jay, regardless of what he'd said in public. I wonder if this is because his kids feel closer to the Phillies than Toronto.
I would be upset if I were a Blue Jay fan. I could understand Mussina not wanting to choose between 2 teams as I believe I heard he spent 10 years in Baltimore before spending the last 8 in NY.


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Health Beat Learning Series - Understanding Stress and Sleep
Community:Northumberland County
Location:Community Health Centres of Northumberland - 99 Toronto Road, Port Hope
Address:99 Toronto Road Port Hope Map
Start date:October 1, 2019
Time:01:00 PM – 02:30 PM
Website:Visit website
Health BEAT Learning Series - NEW
Health education workshops on a variety of topics aimed
at helping you to better manage your health. Check out our
current calendar of events for monthly topics. Please contact us for more information regarding registration.


text: Claiming the distinction of being the first published anthology of modern Indian short fiction, these stories, in spite of a certain provinciality, bring Indian writing more or less into the twentieth century. Gone are the lush myths and religious tales of ancient, pre-Anglicized India. The writers have been influenced not only by the realism of the great nineteenth century writers, but also by two of its most pervasive thinkers--Marx and Freud. The stories are, in today's sense, extra-literary, in that they have been written to teach and correct--to ""make one think."" The tales combine conventional Western realism with the traditional Indian didacticism and tendency towards parable. They seem old-fashioned to the sophisticate accustomed to the comtemporary emphasis on irony and experiment and the current cult of idiosyncrasy. They may seem cliched or even innocuous in spite of their social commitment and concern with injustice. But though the style may seem backward, and the moralizing may strike one as naive, the authors are in general storytellers of much charm and sincerity. Noteworthy are Raja Rao's tale of the plight of the Indian woman, Chandar's moving war story, Bhongi's sometimes mawkish but arresting story of the untouchable, and Santha Rama Rau's story of the educated Indian youth--perhaps, in our terms, the ""hippest"" story of them all. Earlier writers, such as the revered Tagore, tell simpler fables, while the younger, living writers seem to be moving towards a modern complexity. The greatest virtue of this collection is that it will stir a lively interest in India and her problems--the sense of India is this book's most important truth.


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The model of local church staff members having other forms of employment is a trending topic in my life these days.
A View from the Pew: Bivocational Church Staff
A View from the Pew: What to Get Your Church Staff for Christmas
While the rest of the world started Christmas preparations on November 1, many Christian churches in the U.S. generally wait until after Thanksgiving to turn their focus of worship to the celebration of Christ's birth. Whether or not your church has a "Hanging of the Green" service or observes the season of Advent, parishioners often turn their thoughts to showing their appreciation to their church staff with holiday gifts.
Michael Ruffin to join staff of Smyth & Helwys Publishing
Smyth & Helwys Publishing is pleased to announce that Michael L. Ruffin will be joining its staff as a Curriculum Editor on May 1.
A View from the Pew: Making the Most of a Ministerial Transition
An unexpected resignation of a church staff member can feel like a punch to the gut. When a trusted staff member resigns, it's not unusual to feel an acute sense of loss and even betrayal.


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New items in our Stella Mar Store! See our latest news
Size guide
Wednesday February 27, 2019
Cineplex Eglinton Town Center
22 Lebovic Avenue
Toronto Ontario Canada
$15 - Adults, $10 - Youth
These socks are St. Francis inspired. Have you heard the story of St. Francis and Brother Wolf? These socks have cushioned bottoms and are very comfortable.One size95% Polyester 5% Spandex
If you want Where There Is Darkness to play in your area but don't want to host a screening, you can sponsor a showing.
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The cost of $2,500 includes the theatre rental, the screening license, and 20 free tickets to the movie for you to use and give away.
When you buy this package, just let us know which date and area you'd like the screening to be in, and we'll take care of the rest!
*If you want to host your own screening, click here to see our Screening Packages.
This wolf pack is made of a durable oxford fabric. It has two slip interior pockets and one laptop sleeve in the main compartment, a front zipper pocket, two side pockets.
Can you find St. Francis? Put the pieces together to build 'Rocky Landscape With St. Francis,' an engraving by Netherlandish artist, Aegidius Sadeler II.
Our men's 100% soft cotton tank top features the Stella Mar Films logo.
Let your baby eat in fashion with this 100% Polyester fleece bib which features original artwork by our very own Cimela Kidonakis. This Brother Wolf design is inspired by the classic St. Francis legend.
Let your baby eat in fashion with this 100% Polyester fleece bib which features unique artwork by filmmaker and friend Cimela Kidonakis. The tiny hands repeated on this bib represent the informal sign language symbol for "love" which Fr. Rene was frequently caught making in photos.
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This pouch is Fr. Rene inspired. Fr. Rene was frequently seen making this hand symbol which means "love." Original design by filmmaker Cimela Kidonakis. This 100% polyester high quality accessory pouch can be used as a pencil bag, a makeup bag, or even a travel bag - very useful.
Spread the word about our small film company while relaxing in this comfortable unisex zip-up hoodie.


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Here, we present a protocol to detect discrete metal oxygen clusters, polyoxometalates (POMs), at the single molecule limit using a biological nanopore-based electronic platform. The method provides a complementary approach to traditional analytical chemistry tools used in the study of these molecules.
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Ettedgui, J., Forstater, J., Robertson, J. W., Kasianowicz, J. J. High Resolution Physical Characterization of Single Metallic Nanoparticles. J. Vis. Exp. (148), e58257, doi:10.3791/58257 (2019).
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Individual molecules can be detected and characterized by measuring the degree by which they reduce the ionic current flowing through a single nanometer-scale pore. The signal is characteristic of the molecule's physicochemical properties and its interactions with the pore. We demonstrate that the nanopore formed by the bacterial protein exotoxin Staphylococcus aureus alpha hemolysin (αHL) can detect polyoxometalates (POMs, anionic metal oxygen clusters), at the single molecule limit. Moreover, multiple degradation products of 12-phosphotungstic acid POM (PTA, H3PW12O40) in solution are simultaneously measured. The single molecule sensitivity of the nanopore method allows for POMs to be characterized at significantly lower concentrations than required for nuclear magnetic resonance (NMR) spectroscopy. This technique could serve as a new tool for chemists to study the molecular properties of polyoxometalates or other metallic clusters, to better understand POM synthetic processes, and possibly improve their yield. Hypothetically, the location of a given atom, or the rotation of a fragment in the molecule, and the metal oxidation state could be investigated with this method. In addition, this new technique has the advantage of allowing the real-time monitoring of molecules in solution.
Detecting biomolecular analytes at the single molecule level can be performed by using nanopores and measuring ionic current modulations. Typically, nanopores are divided into two categories based on their fabrication: biological (self-assembled from protein or DNA origami)1,2,3, or solid-state (e.g., manufactured with semiconductor processing tools)4,5. While solid-state nanopores were suggested as potentially more physically robust and can be used over a wide range of solution conditions, protein nanopores thus far offer greater sensitivity, more resistance to fouling, greater bandwidth, better chemical selectivity, and a greater signal to noise ratio.
A variety of protein ion channels, such as the one formed by Staphylococcus aureus α-hemolysin (αHL), can be used to detect single molecules, including ions (e.g., H+ and D+)2,3, polynucleotides (DNA and RNA)6,7,8, damaged DNA9, polypeptides10, proteins (folded and unfolded)11, polymers (polyethylene glycol and others)12,13,14, gold nanoparticles15,16,17,18,19, and other synthetic molecules20.
We recently demonstrated that the αHL nanopore can also easily detect and characterize metallic clusters, polyoxometalates (POMs), at the single molecule level. POMs are discrete nanoscale anionic metal oxygen clusters that were discovered in 182621, and since then, many more types have been synthesized. The different sizes, structures, and elemental compositions of polyoxometalates that are now available led to a wide range of properties and applications including chemistry22,23, catalysis24, material science25,26, and biomedical research27,28,29.
POM synthesis is a self-assembly process typically carried out in water by mixing the stoichiometrically required amounts of monomeric metal salts. Once formed, POMs exhibit a great diversity of sizes and shapes. For example, the Keggin polyanion structure, XM12O40q- is composed of one heteroatom (X) surrounded by four oxygens to form a tetrahedron (q is the charge). The heteroatom is centrally located within a cage formed by 12 octahedral MO6 units (where M = transition metals in their high oxidation state), which are linked to one another by neighboring shared oxygen atoms. While tungsten polyoxometalates structure is stable in acidic conditions, hydroxide ions lead to the hydrolytic cleavage of metal-oxygen (M-O) bonds30. This complex process results in the loss of one or more MO6 octahedral subunits, leading to the formation of monovacant and trivacant species and eventually to the complete decomposition of the POMs. Our discussion here will be limited to the partial decomposition products of 12-phosphotungstic acid at pH 5.5 and 7.5.
The goal of this protocol is to detect discrete metal oxygen clusters at the single molecule limit using a biological nanopore-based electronic platform. This method allows the detection of metallic clusters in solution. Multiple species in solution can be discriminated with greater sensitivity than conventional analytical methods33. With it, subtle differences in POM structure can be elucidated, and at concentrations markedly lower than those required for NMR spectroscopy. Importantly, this approach even allows the discrimination of isomeric forms of Na8HPW9O341.
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Note: The protocol below is specific to the Electronic BioSciences (EBS) Nanopatch DC System. However, it can be readily adapted to other electrophysiology apparatus used to measure the current through planar lipid bilayer membranes (standard lipid bilayer membrane chamber, U-tube geometry, pulled microcapillaries, etc.). The identification of commercial materials and their sources is given to describe the experimental results. In no case does this identification imply recommendation by the National Institute of Standards and Technology, nor does it imply that the materials are the best available.
1. Solution and Analyte Preparation
- Prepare all electrolyte solutions with 18 MΩ-cm water from a Type-1 water purification system to remove trace organic species and then filter all electrolyte solutions through a 0.22 μm vacuum filter immediately before ion channel recordings.
Note: The water quality is a critical factor for the stability and longevity of the membrane nanopore system.
- Wild Type αHL.
- Follow MSDS precautions when handling the αHL toxin protein.
- Mix lyophilized wild-type monomeric S. aureus α-Hemolysin (αHL) powder with 18 MΩ-cm water at 1 mg/mL. Distribute 10 to 30 μL aliquots of the sample into cryo-safe centrifuge tubes, quickly flash freeze in liquid nitrogen and then store at -80 °C. Alternatively, use purified preformed heptameric αHL31.
- Dissolve the lipid 1,2-Diphytanoyl-sn-Glycero-3-Phosphocholine (DPhyPC) to 0.2 mg/mL in n-decane in a 4 mL glass scintillation vial with a polytetrafluoroethyleneeflon-coated cap. Store the solution at 4 °C for repeated use for up to one month.
- Prepare phosphotungstic acid solutions.
- Follow MSDS precautions when handling the phosphotungstic acid powder and prepare a 2 mM phosphotungstic acid stock solution by dissolving 57.6 mg of H3PW12O40 into 10 mL of a 1 M NaCl and 10 mM NaH2PO4 solution, which constitutes the stock solution.
- Take 5 mL of this solution and adjust the pH to 5.5 with 3 M NaOH. Adjust the pH of the other 5 mL of the stock solution to 7.5 with 3 M NaOH.
Note: At pH 5.5, 12-phosphotungstic acid (PTA, H3PW12O40) decomposes primarily into the monovacant anion [PW11O39]7-.
2. Test Cell Assembly
- Assemble the test cell per the manufacturer's instructions.
- Soak one Ag/AgCl wire in bleach (sodium hypochlorite) for 10 minutes after abrading it with 600 grit sandpaper. Position the electrode inside the quartz nanopore membrane (QNM).
- Place a cylindrical AgCl pellet electrode embedded in a silver wire outside of the QNM.
- Once the test cell is set up, turn on the power supply and data acquisition program. Ensure that the DC current reading is 0 pA in the absence of solution in the test cell.
- Use a syringe connected to the test cell via a fluid line to add buffered electrolyte solution above the face of the QNM and to ensure that the ionic current saturates the amplifier. If it does not, the QNM may be clogged. Apply a pop voltage (± 1 V) and/or a pressure greater than 300 mm Hg to clear it. If that works, reduce the voltage and pressure.
3. Lipid Bilayer Formation
- Fill the solution in the test cell so that the solution level is well above the face of the QNM. Then lower the solution level via syringe to below the face, such that the current decreases to zero.
- Dip a 10 μL pipette tip into the lipid vial. Push on the back end of the pipette tip and tap it on the side of the vial to remove all visible lipid.
- Touch the pipette tip onto the air-water interface of the solution in the test cell when the solution level is above the QNM's face and wait two to five minutes for the lipid to spread uniformly.
- Slowly lower the solution level below the face of the QNM until the current saturates, and then slowly raise the solution level past the face of the QNM to form a lipid bilayer membrane.
- Once a bilayer appears to form (i.e., when the current goes to zero), try popping it several times by increasing the pressure and ensure that the QNM is not clogged. To reform the lipid bilayer membrane, lower the solution level below the face of the QNM and slowly raise it.
- If the lipid bilayer membrane did not form the first time, lower the solution below the face and raise it again. If it does not form after 3 trials, add more lipids as described in 3.2 and 3.3.
- After forming a membrane, set the current offset to zero when the applied potential is zero.
4. αHL Pore Formation
- Add 2.5 ng of purified preformed αHL heptameric protein sample (or 250 ng of monomeric αHL) to the test cell (volume ≈ 200 μL) to enable channel formation.
- Increase the pressure on the bilayer with a gas-tight syringe (Figure 1) after a bilayer is formed, to expand the membrane from the QNM, and facilitate nanopore insertion. Raise the applied back pressure typically between 40 to 200 mmHg, depending on each QNM.
Note: The EBS software has an automated insertion feature that applies a higher bias (typically 200 to 400 mV) to induce pore formation and then automatically reduces the desired voltage to the measurement bias once a single channel forms.
- After a nanopore forms, reduce the back-pressure to about 1⁄2 of the insertion pressure. If multiple channels are observed, remove them by significantly reducing the pressure.
5. Metallic Cluster Partitioning in the Nanopore
- To account for electrode imbalances, set the DC offset voltage such that when the applied potential is set to zero there is no measured current.
- Prior to adding the POM sample, perform a control experiment to ensure there are no contaminants (e.g., trace POMs from a previous experiment) in the reservoir. Specifically, acquire an ionic current trace under an applied potential of +120 mV to -120 mV in the absence of any POMs to verify that no spontaneous current blockades are present.
Note: Due to the asymmetric structure of the αHL channel (Figure 1), the magnitute of the ionic current will differ for positive and negative applied potentials. The ratio of the measured current above this applied voltage is indicative of the orientation of the αHL nanopore in the membrane.
- Add the POM sample by flushing the reservoir with metallic cluster solution at 1 to 5 mM concentration. Alternatively, load the sample into the capillary prior to cell assembly to study the partitioning of POMs into the other end of the αHL channel.
- Record the ionic current using the manufacturer's software to detect transient current blockades caused by partitioning of individual POMs into the nanopore. Estimate the physical and chemical properties of the molecule from the ionic current blockade depth, event frequency, and the residence time distribution of the blockades.
6. Ion Channel Recordings and Data Analysis
- Acquire the ionic current time series measurements using a high-impedance, low-noise amplifier and data acquisition system. Perform the measurements at an applied voltage of -120 mV (relative to channel cis side) for each pH.
- Apply a low-pass 100 kHz 8-pole Bessel filter to the signal, which is subsequently digitized at 500 kHz (i.e., 2 ms/point). Extract events from the time series and analyze events using the ADEPT algorithm in the MOSAIC software package32,33.
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Representative Results
Over the past two decades, membrane-bound protein nanometer-scale pores have been demonstrated as versatile single-molecule sensors. Nanopore-based measurements are relatively straightforward to execute. Two chambers filled with electrolyte solution are separated by a nanopore embedded in an electrically insulating lipid membrane. Either a patch-clamp amplifier or an external power supply provides an electrostatic potential across the nanopore via Ag/AgCl electrodes immersed in the electrolyte reservoirs. The electric field drives individual charged particles into the pore, which produces transient reductions in the ionic current that depend on the size, shape, and charge of the particles. A computer program controls the applied voltage and monitors, in real time, the ionic current blockades caused by molecules reversibly partitioning into the pore. The current is amplified and converted to voltage with a low-noise, high impedance field-effect transistor and digitized using a data acquisition card.
Here, we provide a general procedure for detecting polyoxometalates with a biological nanopore. As seen in Figure 2, prior to the addition of POMs the unobstructed channel has a mean open channel current of ~ 100 pA at an applied potential of -120 mV. The addition of POMs produces transient blockades and decreases the ionic current by approximately 80%. As expected, because these particles are negatively charged, the blockades are not observed when the polarity of the applied potential is reversed. Note that if the POMs didn't interact with the pore wall, they would diffuse through the pore's in about 100 ns, which is far too brief to be detected with a conventional patch clamp amplifier. Thus, most of the time a given particle spends in the pore is a direct consequence of the interaction between the particle and the pore. The duration of an ionic current blockade event is defined as the residence time, tau (τ).
To illustrate the utility of this method, we discuss the use of an αHL nanopore to monitor the decomposition of 12-phosphotungstic acid (PTA, H3PW12O40) at pH 5.5 and pH 7.5. This decomposition can be observed with 31P NMR measurements, but the concentration needed is 2 mM while nanopore measurements need less than 30 μM, because of the nanopore measurement sensitivity. At pH 5.5, [PW11O39]7- is the predominant species30.
The data analysis is performed by calculating a histogram of the relative blockade depth ratio (i.e.,<i>/<io>, where <i> is the mean current with the POM in the pore and <io> is the mean open channel current). The histogram of the mean current blockade depth ratios at -120 mV and pH 5.5 exhibits a minor peak at <i>/<io> ≈ 0.06 and major peak at <i>/<io> ≈ 0.16 (Figure 3, green). We assume these peaks correspond to [P2W5O23]6- and [PW11O39]7-, respectively, based on 31P NMR. 31P NMR studies suggest that increasing the pH changes the relative concentration of these two species, and this is borne out by the change in the area of the two peaks shown in Figure 3.
When the POM solution is titrated to pH 7.5 ex situ, the total POM concentration decreases due to the partial degradation of the two-principal species to inorganic salts (i.e., free phosphate, HxPO4−3+x and tungstate, WO42− ions). The histogram of the relative blockade depth ratio also shows two principal peaks (Figure 3, orange), but with 20-fold fewer events (which suggests the total POM concentration at pH 7.5 is approximately 20-fold less than that at pH 5.5, if the nanopore's capture efficiency for POMs is the same at the two pH values). It is interesting to note that at pH 7.5 and greater, the POM species observed here were not detected in the 31P NMR spectrum due to their low concentration caused by their dissociation into phosphate and tungstate ions.
Each event's residence time in the pore is defined by the duration of the individual ionic current blockades. The distribution of residence times provides insight into the different species that are present. It was shown earlier that for blockades caused by a differently-sized polymers of poly(ethylene glycol), the residence time distribution for each size of that polymer is well described by a single exponential. That result suggests the interaction of that polymer is a simple reversible chemical reaction12,13,20.
Figure 4 illustrates that the residence time distributions for the two peaks were well differentiated at pH 5.5 and 7.5. Two features are clear. First, under all conditions, multiple exponentials are required to fit each of the distributions, which suggests there are variations of the POMs within each species. Second, the residence times of the POMs in the pore are much shorter at pH 7.5 compared to those at pH 5.5, which suggests a weakening of the interaction between the pore and POMs. It has been shown previously that a change in pH alters the relative number of fixed charges in or near the αHL channel lumen. These changes will directly alter the interactions with partitioning POMs inside the pore and therefore modify their residence times34,35.
Figure 1: Schematic diagram of the experimental setup. Method for nanopore-based characterization of individual polyoxometalate molecules. A protein nanopore that self-assembles in a 4 nm thick lipid bilayer membrane is bathed by aqueous electrolyte solutions in a glass capillary and larger reservoir. A pressure is applied to the glass capillary with a gas tight syringe to aid nanopore incorporation. A potential V is applied across the membrane with a matched pair of Ag/AgCl electrodes and drives an ionic current (e.g., Na+ and Cl-) through the pore. The current is converted to voltage with a high impedance amplifier, digitized with an analog to digital converter (ADC) and stored on a computer. Computer software controls the applied potential through a digital to analog converter (DAC) and monitors, in real time, the transient current blockades caused by single molecules that partition into the pore. Please click here to view a larger version of this figure.
Figure 2: Nanopore-based detection of individual metallo-nanoparticles. An illustration of ionic current time series traces that occur before and after the addition of a POM solution to the nanopore apparatus. The partitioning of individual anionic POMs into the pore causes transient current reductions in the mean open pore current, <io>. (Right) A typical event, illustrating the mean current of the blockade (<i>) and the residence time (τ) of the particle in the pore. The applied potential was -120 mV, and the solutions contained 1 M NaCl, 10 mM NaH2PO4 at pH 5.5. The cis compartment also contained 30 μM of 12-phosphotungstic acid. The current blockade depth ratio (<i>/<io>) and the residence times (τ) provide information about which POM species are present in solution. Under the conditions we used here, the αHL channel does not gate (spontaneously close) when POMs are not present. Please click here to view a larger version of this figure.
Figure 3: Histograms of the current blockade depth ratio at pH 5.5 and 7.5. Histograms of the POM-induced ionic current blockade depth ratio at pH 5.5 (green) and 7.5 (orange) with an applied potential V = -120 mV. The two peaks present at each pH value correspond to the known predominate POM species in solution under those conditions. The current blockade depth ratios of 0 and 1 correspond to a fully blocked and open pore, respectively. The histograms were created with a bin width of 0.001 and normalized to counts/s by dividing by the data acquisition time. Please click here to view a larger version of this figure.
Figure 4: Residence time distribution and fitting with several exponentials. The distribution of residence times for POM-induced current blockades caused by the two-principal species (peaks 1 and 2 in Figure 4) observed at pH 5.5 and 7.5 in a semi-log plot. For both species, the residence times are markedly shorter at the higher pH value, which suggests the interaction between the pore and POMs changed. The solid lines are fits of an exponential mixture model to the data. Please click here to view a larger version of this figure.
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Due to their anionic charge, POMs likely associate with organic counter cations through electrostatic interactions. Therefore, it is important to identify the proper solution conditions and the right electrolyte environments (especially cations in solution) to avoid complex formation with POMs. Particular care is required in the buffer choice. For example, the capture rate of POMs with tris(hydroxymethyl)aminomethane and citric acid-buffered solutions is significantly lower than that in phosphate buffered solution, likely because either of the first two buffers form a complex with the POM that doesn't strongly interact with the nanopore. Moreover, the NaCl electrolyte was purposely used instead of KCl (as well as the other alkali metals) to avoid the precipitation of [PW11O39]7- by K+.
Critical to the accurate measurement of the residence time distributions is the ability to measure the current at a sufficiently high bandwidth. For instance, with exponentially-distributed residence times there are far more blockades with short than long residence times, and an accurate estimation of the residence time distributions is better achieved by collecting a great deal of data (i.e., acquiring it at as high as a bandwidth the system's electrical capacitance allows). To achieve this condition in nanopore spectroscopy, the system capacitance (membrane and stray capacitance) should be minimized. Stray capacitance is reduced by decreasing the length of all connecting cables and using high quality electrical contacts. The membrane capacitance is minimized by decreasing the surface area of the bilayer, increasing the thickness of supporting materials (i.e., quartz, polytetrafluoroethylene, etc.), and decreasing the area of exposed supporting materials to the electrolyte. In practice, a typical instrument's stray capacitance (≈ 2 pF) will limit the noise for membranes ≈ 1 micron to 5 micron in diameter. This constitutes the method's limitation. For example, the detection of small and highly charged single molecules can be challenging due to their relatively short residence times.
The mechanism by which pressure enables control of channel insertion is not completely understood. The quartz microcapillaries have a very small diameter on which the membrane is formed. Applying pressure will cause the membrane to bulge (thereby increasing the membrane surface area) and possibly thin the membrane. Both effects would increase the rate at which channels will form in the membrane. When a single channel spontaneously forms, reduce the pressure to prevent the insertion of additional channels. The removal of non-inserted αHL from the bulk aqueous phase is not required if the αHL concentration is sufficiently low.
The structures and charges of polyoxometalates are currently studied using traditional analytical chemistry techniques, including NMR, Ultraviolet-visible, Infrared and Raman spectroscopy, mass spectrometry, and X-ray diffraction. We expect that nanopore measurements will complement the characterization of these and other physical properties of POMs, as well as the study of their speciation at low concentration, which will help better understand the synthetic pathway of polyoxometalates formation. It was also shown previously that the αHL pore can even distinguish between 2 isomers of the trivacant Keggin form Na8HPW9O3430.
In summary, we have shown that a membrane-bound protein nanopore can be used to detect and characterize tungsten oxide metallic clusters (heteropolytungstates) in solution using a simple high-resolution electrical measurement. The sensitivity afforded by this novel approach permits the tracking of subtle differences in POM structure that arise at different pH values at concentrations that are substantially lower (> 70-fold) than required for traditional methods such as NMR spectroscopy. Due to the single molecule detection capability of nanopores, the actual limit of detection in the method can be made much lower by measuring the current for longer times (the capture rate scales in proportion to the POM concentration).
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We are grateful for financial support from the European Molecular Biology Organization for a postdoctoral fellowship (to J.E.) and a grant from the NIH NHGRI (to J.J.K.). We appreciate the help of Professors Jingyue Ju and Sergey Kalachikov (Columbia University) for providing heptameric αHL, and for inspiring discussions with Professor Joseph Reiner (Virginia Commonwealth University).
|Catalog Number
|Nanopatch DC System
|Electronic Biosciences, Inc., EBS
|Millipore LC-PAK
|Millipore vacuum filter
|1,2-Diphytanoyl-sn- Glycero-3-Phosphocholine (DPhPC)
|Avanti Polar Lipids, Alabaster, AL
|Decane, ReagentPlus, ≥99%,
|List Biological Laboratories, Campbell, CA
|Ag wire
|Alfa Aesar
|2 mm Ag/AgCl disk electrode
|In Vivo Metric
|High-impedance amplifier system
|Electronic Biosciences, San Diego, CA
|quartz capillaries
|custom polycarbonate test cell
|Data Processing and Analysis MOSAIC
|Phosphotungstic acid hydrate
|Sodium Chloride
|sodium phosphate monobasic monohydrate
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