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Unique Legal Blog
Changing Uses for Bypass Trusts
A Discussion of Wills, Part 3: Beware of "Simple" Estate Plans
What Happens If Your Heir Doesn't Want What You Are Giving?
Monday, March 02, 2015
Minneapolis Estate Planning Lawyer Explains the Reasons Why You May Want a Bypass Trust
Every year, each individual who dies in the U.S. can leave a certain amount of money to his or her heirs before facing any federal estate taxes. For example, in 2013, a person who died could leave $5.25 million to his or her heirs (or a charity) estate tax free, and everything over that amount would be taxable by the federal government. Transfers at death to a spouse are not taxable.
Therefore, if a husband died owning $8 million in assets in 2013 and passed everything to his wife, that transfer was not taxable because transfers to spouses at death are not taxable. However, if the wife died later that year owning that $8 million in assets, everything over $5.25 million (her exemption amount) would be taxable by the federal government. Couples would effectively have the use of only one exemption amount unless they did some special planning, or left a chunk of their property to someone other than their spouse.
Estate tax law provided a tool called "bypass trusts" that would allow a spouse to leave an inheritance to the surviving spouse in a special trust. That trust would be taxable and would use up the exemption amount of the first spouse to die. However, the remaining spouse would be able to use the property in that bypass trust to live on, and would also have the use of his or her exemption amount when he or she passed. This planning technique effectively allowed couples to combine their exemption amounts.
For the year 2013, each person who dies can pass $5.25 million free from federal estate taxes. This exemption amount is adjusted for inflation every year. In addition, spouses can combine their exemption amounts without requiring a bypass trust (making the exemptions "portable" between spouses). This change in the law appears to make bypass trusts useless, at least until Congress decides to remove the portability provision from the estate tax law.
However, bypass trusts can still be valuable in many situations, such as:
(1) Remarriage or blended families. You may be concerned that your spouse will remarry and cut the children out of the will after you are gone. Or, you may have a blended family and you may fear that your spouse will disinherit your children in favor of his or her children after you pass. A bypass trust would allow the surviving spouse to have access to the money to live on during life, while providing that everything goes to the children at the surviving spouse's death.
(2) State estate taxes. Currently, 13 states and the District of Columbia have state estate taxes. If you live in one of those states, a bypass trust may be necessary to combine a couple's exemptions from state estate tax.
(3) Changes in the estate tax law. Estate tax laws have been in flux over the past several years. What if you did an estate plan assuming that bypass trusts were unnecessary, Congress removed the portability provision, and you neglected to update your estate plan? You could be paying thousands or even millions of dollars in taxes that you could have saved by using a bypass trust.
(4) Protecting assets from creditors. If you leave a large inheritance outright to your spouse and children, and a creditor appears on the scene, the creditor may be able to seize all the money. Although many people think that will not happen to their family, divorces, bankruptcies, personal injury lawsuits, and hard economic times can unexpectedly result in a large monetary judgment against a family member.
Although it may appear that bypass trusts have lost their usefulness, there are still many situations in which they can be invaluable tools to help families avoid estate taxes.
Don't pay unnecessary taxes, call now to discuss your options with an attorney.
Monday, February 23, 2015
"I just need a simple will." It's a phrase I hear at least once a week. What could be wrong with that? This post explains the many common situations in which a "simple will" may not be a good fit for your family tells the cautionary tale of one family who relied on a will purchased at a stationary store.
Monday, February 16, 2015
Beneficiaries may not want the asset left to them? Why? And what happens if they reject it? I explain the reasons why someone may reject an inheritance and what happens to it if they do.
Monday, February 09, 2015
Should you bring your kids to meet with your estate planning attorney? I discuss the issues related to having them present and what an estate planning attorney should do if you want the children involved.
Monday, January 26, 2015
You have a timeshare in a warm sunny place and want to leave it to a loved one to inherit. How can you do this? What are the issues unique to owning a timeshare?
Monday, January 12, 2015
A Minneapolis Probate Lawyer Discusses the Issue of Using a Will Copy in a Probate
Many people keep their important documents at home where they are easily accessible. It's not at all uncommon to find people with a filing cabinet or even a shoe box containing passports, account statements, deeds, tax returns, birth certificates and social security cards. Wills are often added to these files once the estate planning process is completed. In choosing to store your important estate planning documents at home, however, you risk having the originals lost or destroyed in the case of fire, flooding or theft. So what happens if the original version of your will is lost or ruined?
When a person dies, Minnesota law determines what must happen in the state probate proceeding. In most cases, the "original" of the will must be submitted to the probate court in the county where the person resided. If the original of the will cannot be located and provided to the court, Minnesota's probate code does permit the submission of a photocopy of that signed will though it may cause a delay.
Should you lose the original copy of your will, the best practice would be for you to execute a new will which would make things easier for your family and loved ones upon your death. In that case there would be better assurances that your wishes were followed and carried out. Preparing a new will should not take much time for your attorney. If you work with Unique Estate Law, we can easily finalize a new original for you. In addition, if you have our Foundational Estate Plan, then you received a free account with Legal Vault and copies of your documents should all be online for your, or your loved ones, to access in case of emergency. If for some reason this is not done, you may wish to execute a document stating the original was destroyed in a flood or fire but that you did not intend to revoke it.
Another option to consider to keep the originals of your estate planning documents safe, even in the face of disaster, is purchasing a fireproof/waterproof safe for your home or rent a safe deposit box with a local bank where you can still easily access your documents but keep them secure off-site. Many of my clients have gun safes and have decided to put their plan in the safe. Also, each county in Minnesota will, for a small fee, store your original will.
If you have any questions on storage of your documents, please contact an estate planning attorney at Unique Estate Law.
Wednesday, December 17, 2014
Many people are confused by advance directives. They are unsure what type of directives are out there, and whether they even need directives at all, especially if they are young. There are several types of advance directives. One is a living will, which communicates what type of life support and medical treatments, such as ventilators or a feeding tube, you wish to receive. Another type is called a health care power of attorney. In a health care power of attorney, you give someone the power to make health care decisions for you in the event are unable to do so for yourself. A third type of advance directive for health care is a do not resuscitate order. A DNR order is a request that you not receive CPR if your heart stops beating or you stop breathing. Depending on the laws in your state, the health care form you execute could include all three types of health care directives, or you may do each individually.
If you are 18 or over, it's time to establish your health care directives. Although no one thinks they will be in a medical situation requiring a directive at such a young age, it happens every day in the United States. People of all ages are involved in tragic accidents that couldn't be foreseen and could result in life support being used. If you plan in advance, you can make sure you receive the type of medical care you wish, and you can avoid a lot of heartache to your family, who may be forced to guess what you would want done.
Many people do not want to do health care directives because they may believe some of the common misperceptions that exist about them. People are often frightened to name someone to make health care decisions for them, because they fear they will give up the right to make decisions for themselves. However, an individual always has the right, if he or she is competent, to revoke the directive or make his or her own decisions. Some also fear they will not be treated if they have a health care directive. This is also a common myth – the directive simply informs caregivers of the person you designate to make health care decisions and the type of treatment you'd like to receive in various situations. Planning ahead can ensure that your treatment preferences are carried out while providing some peace of mind to your loved ones who are in a position to direct them.
Monday, November 24, 2014
A Minneapolis Estate Planning Lawyer Defines a Successor Trustee and Explains Why You Should Have One
You did everything right. You sat down with a lawyer, paid her to draft your estate plan, created a living trust and named each other as trustees. But, the unthinkable happened and your spouse died before you did. You were so sure it would be you first. Your lawyer now explains that you are the successor trustee and that you must now administer your spouse's trust. What does she mean by a successor trustee?
Monday, November 10, 2014
A Minneapolis Probate Lawyer Discusses Selecting An Executor Post Mortem
The death of a loved one is a difficult experience no matter the circumstances. It can be especially difficult when a person dies without a will. If a person dies without a will and there are assets that need to be distributed, the estate will be subject to the process of administration instead of probate proceedings.
Monday, October 13, 2014
Families with significant net worth who have a tradition of philanthropy often consider establishing a charitable foundation as part of their estate plans. While there are a number of advantages to using family foundations as a philanthropic vehicle, families need to seek guidance from estate planning and tax professionals to ensure it is the best option for achieving their objectives.
Read more . . .
Monday, September 29, 2014
Stepparent adoption is the most common form of adoption in the United States. Once the adoption is finalized, the stepparent assumes full financial and legal responsibility for his or her spouse's child and the non-custodial parent's rights and responsibilities are terminated.
Previous Posts
When to Involve Adult Children in the Estate Planning Process
Leaving a Timeshare to a Loved One
A Discussion About Wills, Part 2: Is a copy of a will sufficient?
A Discussion About Wills, Part 1: You Really Do Need a Will
Do I Really Need Advance Directives for Health Care?
What is a Successor Trustee
What To Do After Death, Part I
Blog Categories
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Cabin Planning
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Medical Assistance
Non-Traditional Estate Planning
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Blog Links
Archived Posts
From within Hennepin County Unique Estate Law represents estate planning and elder law clients throughout Minnesota, including Minneapolis, Edina, Bloomington, St. Louis Park, Minnetonka, Plymouth, Wayzata, Maple Grove, St. Paul, and Brooklyn Park. The Minnesota law firm of Unique Estate Law focuses on all aspects of estate planning, including specialized wills, trusts, powers of attorney and medical directives for married couples, young families, blended families, single parents, gay families and those going through a divorce. Unique Estate Law also handles probate administration, asset protection, Medical Assistance planning, elder law, business succession planning, adoptions and cabin planning.
© 2015 Unique Estate Law | Disclaimer 3800 American Blvd., Suite 1500, Bloomington, MN 55431 | Phone: 952-955-7623 333 Washington Avenue North, Minneapolis, MN 55401 | Phone: 952-955-7623 5775 Wayzata Blvd., St. Louis Park, MN 55416 | Phone: 952-955-7623 Asset Protection | Business Succession Planning | Cabin Planning | Estate Planning | Estate Planning for High Net Worth Individuals | Estate Planning for Unmarried Couples | Guardianships | Incapacity Planning | Planning for Children | Special Needs Planning | Small Business Assistance | Advanced Estate Planning | Estate Planning | Fees | Probate
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This season's form skier Tina Maze put herself on track for a grand slam of crystal globes with victory in the slalom at Ofterschwang Germany.
Maze was second after the morning run but fought back in style in the second to snatch her 10th victory of the World Cup season, her second in a
slalom. The Slovenian's win lifted her to the top of the world cup slalom standings and also increased her all-time points record in the overall World Cup standings to 2.254.
Maze remains in contention to win all four discipline World Cups on top of the overall title.
In the slalom standings she now leads second-placed Mikaela Shiffrin by seven points with only one race left at the World Cup finals in Lenzerheide next week.
Shiffrin – the American teenage sensation who won slalom gold at the world championships in Schladming early last month and three slalom races on the World Cup circuit this year – had to settle for third place on the day.
The 17-year-old from Vail, Colorado led after the first leg but was third fastest in the second to slip down the podium behind Maze and Swiss skier Wendy Holdener.
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Nick Jonas Tickets
Nick Jonas Biography & News
See all pop ticketsNick Jonas Details
Genre | Official site |
Pop |
About Nick Jonas
About Nick Jonas
Nick Jonas has always been the most successful of the Jonas Brothers, a pop-rock band that was embraced the world over by a throng of shrieking pre-teen and teenage girls. Yet while Kevin and Joe were always looked as just part of the band, it was Nick who broke off to become a singer, songwriter, actor and all-around superstar.
The story goes that Nick was 'discovered' in a barber shop while his mother was getting her hair cut. He had a business manager by the age of six, began performing on Broadway by the age of seven, and went on to star in the Disney Channel original series "Jonas L.A." alongside his brothers. Next came the Disney Channel original movies "Camp Rock" and "Camp Rock 2: The Final Jam," and along with other small screen projects he currently stars in the DirectTV drama series, "Kingdom," which premiered on the Audience Network in the fall of 2014.
Though the past year has had its ups and downs for Nick, he pushed along after a "deep rift" within the Jonas Brothers resulted in a tour cancellation. Next their Twitter page went offline, then the brothers' individual accounts disappeared. Despite the fact that Joe and Nick originally took to social media to ask fans to "bear with [them]," the Jonas Brothers officially confirmed their split on October 29, 2013.
Nick spent parts of 2014 as the musical and creative director for Demi Lovato's Neon Lights tour, joining her on select dates to perform the songs "Stop the World," "Here We Go Again," and "Catch Me." He also released two singles – "Chains" and "Jealous" off his upcoming self-titled album.
Nick Jonas Discography
Album name | Release date |
Who I Am | 2010 |
Top 10 Nick Jonas Songs
Rank | Track Name | Play Count | Listener Count |
1 | Who I Am | 18,045 | 7,737 |
2 | Rose Garden | 6,941 | 2,840 |
3 | Last Time Around | 6,147 | 2,686 |
4 | Vesper's Goodbye | 6,755 | 2,563 |
5 | Tonight | 5,008 | 2,422 |
Next 4 Concerts
Nick Jonas Drops New Single, Announces Solo Tour for This Fall
Nick Jonas is having a busy early September. First, he sang the national anthem on Sunday to start off the US Open Women's Finals, (to mixed reactions), then on Monday he released a new single of his upcoming solo album (which you can listen to below) and now he's released a bunch dates for his…
Read moreNick Jonas Ticket Prices
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Nick Jonas Tour Dates 2014
date city state venue 12/31/2014 Las Vegas NV Drais Rooftop Beach Club 12/22/2014 Tampa FL Amalie Arena 12/21/2014 Sunrise FL BB&T Center 12/19/2014 Buffalo NY First Niagara Center 12/18/2014 Rosemont IL Allstate Arena 12/15/2014 Washington DC Verizon Center 12/14/2014 Boston MA TD Garden 12/12/2014 New York NY Madison Square Garden 12/11/2014 Rochester NY Blue Cross Arena 12/10/2014 Philadelphia PA Wells Fargo Center 12/09/2014 Charlotte NC The Fillmore Charlotte 12/05/2014 Los Angeles CA Staples Center 12/03/2014 San Jose CA SAP Center 11/30/2014 Grand Prairie TX Verizon Theatre at Grand Prairie 11/06/2014 Brooklyn NY Rough Trade NYC 11/05/2014 Brooklyn NY Music Hall of Williamsburg 11/04/2014 New York NY Gramercy Theatre 10/30/2014 West Hollywood CA Troubadour 10/29/2014 Los Angeles CA The Satellite 10/08/2014 Boston MA Wilbur Theatre 10/06/2014 Toronto ON Lee's Palace 10/05/2014 Pontiac MI Crofoot Ballroom 10/03/2014 Minneapolis MN Fine Line Music Cafe 10/02/2014 Chicago IL Bottom Lounge 09/30/2014 Dallas TX Trees 09/29/2014 Houston TX Fitzgerald's 09/27/2014 Mesa AZ Mesa Arts Center 09/24/2014 San Francisco CA Great American Music Hall 09/22/2014 Seattle WA The Showbox Nick Jonas Tour Dates 2013
date city state venue 07/20/2013 Wantagh NY Nikon at Jones Beach Theater Nick Jonas Tour Dates 2012
date city state venue 07/26/2012 Cambridge Cam Cherry Hinton Hall Nick Jonas Tour Dates 2011
date city state venue 08/13/2011 Bethlehem PA Sands Riverplace


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Spinelessjellyfish, a multiple stakes winner who was grade I stakes-placed, will enter stud at Steve Sahadi's Cardiff Stud Farms near Atascadero, Calif.Bred in California by Cardiff Farm Management, Spinelessjellyfish earned $779,367 of his career total of $889,591 on grass and made a habit of distinguishing himself in several California turf stakes for trainer Jenine Sahadi. He won two consecutive runnings of Khaled Stakes and two runnings of the Sensational Star Handicap. He also captured the California Turf Championship Handicap and once ran third in it, and was second in the Crystal Water Handicap three times. Against graded competition, he placed in the Shoemaker Breeders' Cup Mile Stakes (gr. IT) and the San Simeon Handicap (gr. IIIT). Overall, he won or placed in 15 stakes.Retired sound, Spinelessjellyfish ended his career with 10 wins from 48 starts and earnings of $889,591. He raced the first part of his career for Steve Sahadi and Barton and Ronelle Heller and the remainder for Sahadi. A fee will be announced later, and there is a possibility he will be syndicated.
Spinelessjellyfish to Cardiff in California
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Dying Too Young: Trends show poverty plays a role, many deaths are preventable
Here are some trends found in a review of child deaths in Oklahoma:
* Poverty often plays a role.
"If they died from a car wreck ... a drowning, intentional abuse or sudden illness ... a commonality that you see more times than not is that this family is poor," said Annette Wisk Jacobi, a state Health Department official who has served for years on the state Child Death Review Board. "A lot of the time we're looking at children left alone or left with a child not old enough to care for everyone else or ... left with somebody who is not a very good care provider. ... As the economy kind of goes south, we're going to see more of this."
In many child murder cases, the killer is unemployed or financially stressed from reasons such as divorce, court records show. One out-of-work defendant awaiting trial admitted to police he became frustrated caring for his two stepchildren and infant son in a motel room day after day while his wife worked. He said he slapped, threw, kicked and stomped his 2-year-old stepdaughter Liliana because she wouldn't eat her lunch.
* The number of child deaths in Oklahoma each year dropped from more than 1,000 in the early 1980s to fewer than 800 in recent years.
* Almost 100 children have been homicide victims since the start of 2006, according to the state medical examiner. But those statistics can be misleading. A drowning death caused by neglect was listed by the medical examiner as manner unknown. And a smoke inhalation death — where the batteries for the smoke alarms had been used for toys — was considered an accident. Many deaths are considered to be from natural causes.
* Some deaths from possible abuse or neglect go unpunished. No arrests were made in 17 of 47 child homicides studied in 2007 by the Child Death Review Board, according to its latest report. Prosecutors have told the board they sometimes can't identify a perpetrator because too many people were around a child around the time of an injury.
* Dozens of deaths could have been prevented, especially those caused by parents and infants sleeping together.
In case after case, an autopsy found a baby died after a sleeping parent rolled over on him or her. Alcohol or drugs are sometimes involved.
Continue reading this story on the...
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submitted 2 years ago by SteezyKiiid
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[–]Senor_Wilson 4 points5 points6 points 2 years ago (2 children)
She's so god damn trashy.
[–]YaDamnSkippy 3 points4 points5 points 2 years ago (0 children)
Trashy works for me
[–]RulerOf -1 points0 points1 point 2 years ago (0 children)
Money has a way of letting you get away with that whenever you want.
It also helps that she's probably more obsessed with fitness than half of the subeddits dedicated to the same.
π Rendered by PID 32574 on app-64 at 2015-03-06 08:34:25.321069+00:00 running 9d9577f country code: US.


text:
Credits & Info
- Uploaded
- Feb 25, 2013 | 3:14 PM EST
Author Comments
You were traveling through the desert when your car broke down. You walked for miles and miles. A town is in sight, maybe you can get some help!
Rated 1.5 / 5 stars
If you get the gold coin before you get the paint thinner, you can't use any items in your inventory.
Rated 2.5 / 5 stars
An okay game, not too hard but one of the puzzles was challenging. However the bug effecting the music was a definite downer.
Rated 2.5 / 5 stars
Make the hints pop somehow.. build the walkthrough into the game if you have to, dont leave us with a "figure it out" game and not enough to figure out.
Rated 2 / 5 stars
Music cuts off and has to be restarted every few minutes. Also, this game relies heavily on the walkthrough. the only thing I figured out was the cash register puzzle, and I still had to use a calculator to figure it out. Modify this a bit and it'll have a better rating.
Rated 2 / 5 stars
some of the puzzles were clever, the bottles and the cash register. But it seemed a little too easy, I did get stumped for a few minutes trying to find the next gold coin, I found the carpet just by clicking around randomly in each room. it was decent but it was way too quick and little too obvious for the most part.


text:
updated 3 weeks ago
Total Listed: 54291 | Last Hour: 8686 | Trade Offers: 45440 |
hah this isn't even priced
http://www.tf2outpost.com/trade/24871137 ~ 1.2
http://www.tf2outpost.com/trade/25091542 ~ 1
surprisingly all I can find…
http://backpack.tf/vote/id/54d31e6ab98d88a2178b476f Countering.
Classifieds at the time of posting this suggestion. http://gyazo.com/3437936668d7d8916975c01c85969025…
2nd suggestion for me, sold mine for this:
http://oi59.tinypic.com/1zld93c.jpg (it's 6.7 keys)
On outpost all Pro. killstreaks, on the market starts at…
Ref suggestion with real Proof...
Don't sue me, Post rage below and be mad >:L
Sellers Below or low end Backpack.tf prices
0.11 are oos... Should be considered as a qs
http://www.tf2outpost.com/trade/24644989 - 0.12
Seller's BP before
Seller's BP after
Needs a slight drop in price, had one myself and it was very hard to sell. Tried to sell it for pure for a total of 4 days but I didn`t get a single offer,…
i m demomn giv me al ur mony.
1 @ 1.44 Ref
4 @ 1.55…


text:
The Modern World:a History 4th Edition Pearson - FindersCheapers.com
The Modern World:a History 4th Edition Pearson Price comparison. Compare and save at FindersCheapers.com. |
Filetype: Submitter: fatenahmed |
Marlinez Jr.
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Ralph Langley Elementary Home of the Lobos
Spring Break March 15-19 March 15-19 will be a student/ staff holiday. The Langley Staff wishes you a safe and happy Spring Break! Chick-Fil-A Night We have made good ... |
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LEON VALLEY ELEMENTARY school may 2010 A TEA Exemplary Campus
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text:
Long before people, ancient creatures roamed the earth. They were known as dinosaurs. Follow Opie and the boy as they travel to England to work with the eccentric Benjamin Waterhouse Hawkins and Richard Owen to build the world's first dinosaur models. Join Opie as he learns all about the dinosaurs and discover tons of prehistoric fun by the end of A Possum's Prehistoric Adventure.
©2011 Jamey M. Long (P)2011 Tate Audio
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Newark Mayor Cory Booker Responds to 'Meet the Press' Controversy on 'Tonight Show' (Video)
The New Jersey Democrat says President Obama was "the first person to step forward and say 'hey, you're my friend, we all say things that don't come out as we mean them, and I'm still with you.'"
Cory Booker says he and President Obama remain on good terms after he blasted campaign tactics against Mitt Romney on NBC's Meet the Press.
In an appearance on Monday's edition of The Tonight Show, Booker told Jay Leno that Obama is a "good person" who was also the "first person to step forward and say 'hey, you're my friend, we all say things that don't come out as we mean them, and I'm still with you.'"
PHOTOS: 20 Biggest Political Players in Hollywood
The Newark, N.J., mayor drew criticism from the Obama campaign and its supporters following his May 21 Meet the Press appearance, where he complained about negative campaigning on both sides, condemning the president's attack ads on Romney's career at Bain Capital as "nauseating."
Republicans immediately pounced on Booker's comments, and the mayor responded with a video on YouTube in which he said it was fair for Obama to single out Romney's private-equity past.
On the Tonight Show, Booker -- a surrogate for Obama during the 2012 presidential election -- said "the reaction after the show, to be really 100 percent sincere, my biggest concern was my President."
He also spoke about heroically rescuing a woman from a fire, and co-starring with New Jersey Governor Chris Christie in an amusing web video sending up their competitive rivalry.
Watch the Leno clip below.
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Given the atmosphere of suspicion and distrust that so often characterizes U.S.-China relations, it is vitally important that Chinese foreign policy and military capabilities be calmly and carefully assessed.
Issues / War & Peace
The Bush administration's defense review affords an opportunity to overhaul the nation's military strategy, forces, and equipment plans in light of the challenges and opportunities of the new century.
Violence and warfare in Colombia are often blamed on the drug trade, but their roots run much deeper and go back well over five decades.
The militarization of Washington's Latin America policy is being led by the drug war, training programs, arms transfers, and a wide array of "military-to-military contact" efforts.
The militarization of Latin America, which begins at the U.S.-Mexico border, is undermining recent trends toward greater democratization and respect for human rights while doing little to stanch the flow of drugs into the United States.
U.S. officials have sought to relax restrictions on intelligence sharing with Andean governments at a time when these provisions need to be strengthened.
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Under Qaddafis rule, Libya has made impressive gains in health care, education, housing, womens rights, and basic social services.
The Bush administrations Andean Regional Initiative (ARI)largely an expansion of U.S. support for Plan Colombiapassed the House of Representatives in late July, largely intact.
Poverty, social disruption and destruction stemming from these wars, and the proliferation of small arms and light weapons are major factors in expanding the use of child soldiers.


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An integrated microfluidic device for influenza and other genetic analyses.
ABSTRACT An integrated microfluidic device capable of performing a variety of genetic assays has been developed as a step towards building systems for widespread dissemination. The device integrates fluidic and thermal components such as heaters, temperature sensors, and addressable valves to control two nanoliter reactors in series followed by an electrophoretic separation. This combination of components is suitable for a variety of genetic analyses. As an example, we have successfully identified sequence-specific hemagglutinin A subtype for the A/LA/1/87 strain of influenza virus. The device uses a compact design and mass production technologies, making it an attractive platform for a variety of widely disseminated applications.
- Microchimica Acta 10/2013; 181(13-14):1611-1631. · 3.72 Impact Factor
Article: A droplet microfluidic approach to single-stream nucleic acid isolation and mutation detection.
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ABSTRACT: In this work, a droplet microfluidic platform for genetic mutation detection from crude biosample is described. Single-stream integration of nucleic acid isolation and amplification is realized on a simple fluidic cartridge. Subsequent DNA melting curve is employed with signal normalizing algorithm to differentiate heterozygous K-ras codon 12 c.25G>A mutant from the wildtype. This technique showcases an alternative to modular bench-top approaches for genetic mutation screening, which is of interest to decentralized diagnostic platforms.Microfluidics and Nanofluidics 08/2014; 17(2):425-430. · 2.67 Impact Factor - SourceAvailable from: Byung Hyun Park
Article: Integration of sample pretreatment, μPCR, and detection for a total genetic analysis microsystem
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ABSTRACT: Since the emergence of lab-on-a-chip technology, a variety of chemical and biochemical assays were successfully implemented on microdevice platforms. Among the chip-based applications, genetic analysis based on the polymerase chain reaction (PCR) has been extensively developed in order to accomplish the goal of cheap, rapid, high-throughput, and point-of-care DNA testing. We are summarizing here several formats of the miniaturized PCR systems including the integration of units for sample pretreatment and downstream analytical detection. The various sections cover (a) miniaturized PCR systems, (b) integrated sample pretreatment-PCR microsystems, (c) integrated PCR-detection microsystems, and (d) integrated sample pretreatment-PCR-detection microsystems. Respective microdevices were successfully introduced recently in the form of a fully integrated microsystem for genetic analysis with sample-in-answer-out capability. Contains 120 references. Figure Microchimica Acta 10/2013; · 3.72 Impact Factor
Page 1
Lab on a Chip
An integrated microfluidic device for influenza and other
genetic analyses
Rohit Pal, Ming Yang, Rongsheng Lin, Brian N. Johnson,
Nimisha Srivastava, S. Zafar Razzacki,
Kenneth J. Chomistek, Dylan Heldsinger, Razi-
Ul Muhammad Haque, Victor Ugaz, Prasanna Thwar,
Zheng Chen, Kenneth Alfano, Moon-Bin Yim,
Madhavi Krishnan, A. Oveta Fuller, Ronald G. Larson,
David T. Burke and Mark A. Burns
We have designed and fabricated an integrated device to
perform genetic analyses including the detection and subtyping
of the influenza virus. The device is inexpensive and self-
contained, suggesting that this platform could be used for
wide-scale dissemination of genetic tests. Author - please edit contents entry text to fit
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Page 2
An integrated microfluidic device for influenza and other genetic analyses
Rohit Pal,{aMing Yang,{aRongsheng Lin,aBrian N. Johnson,aNimisha Srivastava,aS. Zafar Razzacki,a
Kenneth J. Chomistek,aDylan Heldsinger,aRazi-Ul Muhammad Haque,aVictor Ugaz,aPrasanna Thwar,a
Zheng Chen,aKenneth Alfano,aMoon-Bin Yim,aMadhavi Krishnan,aA. Oveta Fuller,bRonald G. Larson,a
David T. Burkecand Mark A. Burnsad
Received 4th May 2005, Accepted 22nd July 2005
First published as an Advance Article on the web
DOI: 10.1039/b505994a
An integrated microfluidic device capable of performing a variety of genetic assays has been
developed as a step towards building systems for widespread dissemination. The device integrates
fluidic and thermal components such as heaters, temperature sensors, and addressable valves to
control two nanoliter reactors in series followed by an electrophoresis separation. This
combination of components is suitable for a variety of genetic analyses. As an example, we have
successfully identified sequence-specific hemagglutinin A subtype for the A/LA/1/87 strain of
influenza virus. The device uses a compact design and mass production technologies, making it an
attractive platform for a variety of widely disseminated applications.
The Human Genome Project has catalyzed research in the area
of miniaturized analytical systems, or lab-on-a-chip techno-
logies, for genetic analyses. The development of these systems
involves the integration of microfabricated components,1–5
such as microscale separations,6–10reactions,11–15microvalves
and pumps,16–23and various detection schemes,24–29into fully
functional devices. Several devices that have integrated DNA
amplification reactions and electrophoretic separations have
been developed.30–34
Devices that employ hybridization
fluorescence detection have been fabricated to perform sample
preparation, various fluidic manipulations, and polymerase
chain reaction (PCR).35Integration of multiple steps of
biological assays on a single device provides significant
advantages in terms of sample/reagent consumption, process
automation, analysis speed and efficiency, and contamination
Integrated genetic analysis devices can find applications in
two broad categories. The first class is centralized laboratory-
based high throughput systems for applications such as
genomics, proteomics, metabolomics, and drug discovery.36
These devices are traditionally in the microarray format
though there are some exceptions. High throughput systems
are designed to yield a large amount of information from a
single experiment. They normally are customized, involving
complex chemistry/biochemistry processes (e.g., DNA probes
for microarrays) in manufacturing. High throughput systems
need not be portable and typically require auxiliary equipment
for large information processing.
The other class of lab-on-a-chip devices is the group
designed for widespread distribution for applications such as
infectious disease diagnosis, chemical/biological warfare detec-
tion, agriculture pest isolation, and forensic identification.37–41
Unlike the high throughput systems, these devices are typically
used to provide extremely specific and simple information
about a small number of samples. Widespread dissemination
of these devices for point-of-care use by untrained personnel
can be one goal and requires a high degree of integration and
process automation.
A particular example of a widely disseminated application is
the identification of the influenza viral strain. Influenza is a
significant long-term public health threat because of its genetic
mutability, rapid transmission, and ability to move from
species to species.42,43Influenza pandemics occurred several
times in the 20th century (1918, 1957 and 1968), and all had
significant death tolls. In each case, a newly assorted influenza
strain to which humans have little immunity was introduced
into the human population, presumably from an animal
source. Recent outbreaks with pandemic potential in East
Asia also introduced new H5 avian influenza strains. Given the
rapidly changing influenza genome, an adaptable, worldwide
surveillance of specific types and subtypes is a high priority,44
and genetic analysis based on PCR and sequence-specific
detection provides the most accurate method for typing
influenza and other viruses.
We have designed and tested an integrated genetic analysis
device. The device is designed to perform two independent
serial biochemical reactions, followed by an electrophoretic
separation (Fig. 1). The key components (phase change
valves,45thermally isolated reaction chambers,46gel electro-
phoresis,47and pulsed drop motion) that were developed for
this device are electronically addressable and simple to
operate, properties that can lead to eventual autonomous
operation. We have successfully performed influenza viral
aDepartment of Chemical Engineering, University of Michigan, Ann
Arbor, MI 48109, USA
bDepartment of Microbiology and Immunology, University of Michigan,
Ann Arbor, MI 48109, USA
cDepartment of Human Genetics, University of Michigan, Ann Arbor,
dDepartment of Biomedical Engineering, University of Michigan, Ann
Arbor, MI 48109, USA
{ These authors contribute equally to the paper.
PAPER www.rsc.org/loc | Lab on a Chip
This journal is ? The Royal Society of Chemistry 2005Lab Chip, 2005, 5, 1–9 | 1
Page 3
strain (A/LA/1/87 and A/Sydney/5/97) subtyping on this device
using a PCR-Restriction Fragment Length Polymorphism
(PCR-RFLP) that distinguished the polymorphism in the
hemagglutinin coding region. Other PCR-based analyses
including amplification from human genomic DNA (D1S80
locus) and mouse plasmid DNA (SNRPN locus) were also
Materials and methods
Glass microfabrication
The microfabricated devices are assembled by bonding silicon
and glass substrate components (Fig. 2). Borofloat glass wafers
(500 mm thick, 10 cm diameter) are annealed at 560 uC for 1 h.
Chromium and gold (600/4000 A˚) are evaporated on the glass
wafer. Photoresist (MicropositSC 1827; Shipley Co.,
Marlborough, MA) is spun on at 3000 rpm and soft baked
at 100 uC for 5 min. The fluidic channel pattern is then exposed
on the wafer using a mask aligner (Cannon PLA 501FA,
405 nm wavelength, 500 light integral). The lithography is
finished by developing the wafer in MF319 (Shipley Co.) for
1.5 min. The photoresist is hard baked at 135 uC for 30 min.
The gold is etched for 4 min and the chromium is etched for
2 min, each in its respective etchant (gold etchant TFA;
Transene Co and chromium etchant CR-14; Cynatek Corp.,
Fremont, CA). The glass is etched in hydrofluoric acid (49%
HF, CMOS grade; J.T. Baker, Philipsburg, NJ) to obtain the
desired channel depth using an estimated etch rate of
7 mm min21. The access holes in the wafer are laser drilled
(Accu-Tech Laser Processing Inc., San Marcos, CA). In some
cases the wafer is coated with a 2 mm thick parylene layer by
chemical vapor deposition (CVD). The glass wafer is diced to
obtain the individual devices.
Silicon microfabrication
Silicon fabrication begins by growing a 2000 A˚thick thermal
oxide on a silicon wafer (<100>, 500 mm thick). Lithography
for the heater elements involves spinning, soft baking,
exposing, developing the photoresist, and is similar to the
glass fabrication, except that titanium and platinum (Ti/Pt;
Fig. 1
device. There are three liquid entry channels (''L'', sample, PCR
reagents and RD reagents), several metering channels, drop mixing
intersections, a sealed PCR chamber, an open RD chamber, and an
electrophoresis channel. Each valve (''V'') is individually and electro-
nically addressable. (b) Photograph of an assembled device. The
discrete liquid drops controlled in this device are 100 to 240 nl, with
fluidic channel dimensions of 200–600 mm wide and 50 mm deep. The
electrodes and diodes shown in the figure were not used in this work.
(a) Schematic representation of an integrated microfluidic
Fig. 2
device. (a) Silicon side fabrication; (b) glass side fabrication.
Process steps for the fabrication of the hybrid glass–silicon
2 | Lab Chip, 2005, 5, 1–9This journal is ? The Royal Society of Chemistry 2005
Page 4
300/1000 A˚) are evaporated on the wafer after the resist is
patterned. The wafer is left in acetone (CMOS grade; J.T.
Baker) for 15 min to liftoff unwanted metal. A 2 mm thick
parylene layer is deposited on the wafer by CVD. Lithography
for the electrical contact holes is performed and the parylene is
etched using oxygen plasma reactive ion etching (RIE 2000;
South Bay Technologies, 100 W and 100 mT oxygen plasma).
The silicon side is coated with a protective layer of photoresist
and diced to obtain the individual devices.
Glass–silicon device assembly
A custom designed and mass produced printed circuit board
(PCB, Advanced Circuits, Aurora, CO) serves as the platform
for electrical connections. After receiving a supply of PCBs,
each one has an area removed from the center (using a small
router bit or scroll saw) to allow access to the back side of
device for active cooling. The diced silicon devices are cleaned
(with acetone, isopropyl alcohol, and de-ionized water) and
fixed onto the PCB using standard quick cure epoxy. The
devices are wire bonded (Kulicke & Soffa 4124 Ball Bonder)
using 1.0 ml gold wire. The cleaned glass device component is
visually aligned to the silicon side and UV curable optical glue
(NOA 72; Norland Products, Cranbury, NJ) is wicked between
the glass and silicon sides through the edges. The device is
cured in UV light (365 nm) for 6 h and then in an oven at 50 uC
for 12 h. The wirebonds are encapsulated with a non-
fluorescent epoxy (EP939; Thermoset, Indianapolis, IN).
O-rings are attached to each end of the separation channel
to act as buffer reservoirs.
Pulsed drop motion
Lab air supply, controlled by a pressure regulator (Matheson
Gas Products Inc, Irving, TX, Model Number 3701), provides
the pneumatic pressure used to move drops in the micro-
channel. The regulated air that outlets from the pressure
regulator is connected via a solenoid valve (Numatics Inc.,
Model Number LS02L6H00B) to a pipette tip. The final
connection to the chip is made by either epoxying or manually
holding a pipette tip at the appropriate access hole in the glass
device. The solenoid valve is in a normally closed position and
may be opened by applying a +12 V DC voltage. Opening and
closing of the solenoid valve are controlled through a
combination of a DC power supply (Electro Industries,
Model Digi 35A) and a relay board (National Instruments,
TX, Model ER-16). Functioning of the relay board is con-
trolled through a program written in LabVIEW (National
Instruments, TX) and a Digital I/O card (PCI-DIO-96). The
pulsed pressure system is also used to operate the phase change
valves. The phase change material used in the valves is a
custom mixture of M1595 microcrystalline wax and Lycojet
Temperature control
The setup for performing PCR and restriction digest thermal
reactions consists of a DC power supply (B+K Precision
Model 1760, Yorba Linda, CA), two data acquisition (DAQ)
boards (National instruments PCI 6031E and PCI-6704,
Austin, TX), two connector blocks (National instruments
SCB-100 and SCB-68, Austin, TX), a custom signal condition-
ing circuit, a computer and two LabVIEW programs (National
instruments, Austin, TX). To calibrate the temperature sensors
the entire device is incubated in a convection oven whose
temperature is ramped from room temperature to 100 uC. The
resistance is recorded in a LabVIEW program by sending a
current of 3 mA through two leads of the four point sensors
with the voltage being measured across the other two leads.
The slope and intercept from a linear fit of the resistance and
temperature data is read into the control algorithms for
temperature control. Temperature is controlled by regulating
the voltage across the microfabricated heaters using a
proportional-integral (PI) algorithm. The heaters are con-
nected to the power supply through the signal conditioning
circuit that boosts the supply voltage from the computer with
an op-amp gain of 3. Thermal isolation of the PCR chamber
from other components on the device is achieved by removing
the PCB under the areas requiring low temperatures and
refilling the cavity with a high-conductivity silicone-based
thermal grease (Product #54013, AOS Thermal Compounds,
Eatontown, NJ). During the operation, the entire device is
placed on a probe station or a Peltier device with temperature
maintained at 10 uC.
Influenza DNA amplification
The PCR-RFLP assay used for the identification of influenza
viral strain A/LA/1/87 consists of two biochemical reactions: a
PCR followed by a restriction digestion. The hemaggultinin
gene (HA1) region of influenza viral RNA was reverse
transcribed, PCR amplified, ligated into pGEM-T vector,
and cloned into E. coli. The cloned plasmid was used to
synthesize RNA in vitro with T7 RNA polymerase. The RNA
samples are then subject to reverse transcription to produce the
DNA samples that were used in the device. For the on-chip
influenza reaction, the PCR primers amplify a 690 bp fragment
of the coding region from the cloned, transcribed HA1 RNA,
following reverse transcription. The reaction mixture (y240 nl)
consists of 2 ng ml21DNA template, 0.2 mM each dNTP,
60 mM Tris-HCl, 15 mM NH4SO4, 1.5 mM MgCl2, 0.1%
TX-100, 0.3 mM each primer, and Taq DNA polymerase
75 units ml21(Invitrogen, Carlsbad, CA). The thermocycling
protocol applied to the device consists of 92 uC for 30 s, and
then 35 cycles of the following: 92 uC for 5 s, 55 uC for 10 s,
and 72 uC for 20 s, and finally 72 uC for 60 s, for a total cycling
time of 22 min. A portion of the PCR product (y60 nl) is
subsequently subjected to a restriction endonuclease digestion
within the same device. The digestion reaction mixture consists
of 500 units ml21HpaI restriction endonuclease (New England
Biolabs, Beverly, MA), 1X NEBuffer 4, 10 mg ml21bovine
serum albumin (BSA), and 0.01 mM YOYO-1 fluorescent dye
(Molecular Probes Inc., Eugene, OR). Successful HpaI
endonuclease digest cuts the PCR product into two smaller
fragments of 145 bp and 545 bp. The restriction digest reaction
is performed at 37 uC for 10 min. A 690 bp fragment of the
hemagglutinin coding regions of influenza A/Sydney/5/97
DNA can be amplified by the same primer set as the A/LA/
1/87 strain, but is not digested with HpaI endonuclease.
This journal is ? The Royal Society of Chemistry 2005Lab Chip, 2005, 5, 1–9 | 3
Page 5
Human and mice DNA amplification
Amplification of the human genomic DNA uses a commercial
amplification kit(EDVO-Kit#
Bethesda, MD). The template for this reaction is an anony-
mous human genomic DNA sample. The amplicon is approxi-
mately 650 bp. The reaction mixture consists of 2.5 ng ml21
human genomic DNA, 0.2 mM each
dNTP, 1X Taq DNA polymerase buffer, 1.5 mM MgCl2,
0.5 mM each primer, and Taq DNA polymerase 100 units ml21
(Invitrogen, Carlsbad, CA). The reaction mixture is subject to
thermocycling at 94 uC for 5 s, 65 uC for 10 s, and 72 uC for 20 s
for 35 cycles. The template for the mouse plasmid DNA
amplification is mouse Snrpn gene DNA cloned from genomic
Mus spretus into a pGEM vector. The amplicon is 996 bp. The
same PCR reaction mixture composition and thermocycling
parameters have been used for mouse Snrpn PCR as for
Influenza PCR except the genomic templates and primers. The
PCR primers used to amplify each genetic marker are
presented in Table 1, along with the expected product sizes.
334, EDVOTEK,West
Microgel electrophoresis
Electrophoretic separations were performed using photo-
Amersham Pharmacia Biotech). Two gel formulations were
used: ReproGel High Resolution (8%T) and ReproGel Long
Read (7%T). The gel solution was prepared by mixing one part
of the acrylamide stock solution with two parts of the
photoinitator solution, and is loaded into the channel network
through the inlet hole at one end of the separation channel.
Gel casting was performed by placing the device under a UV
lamp for a period of 10 min using a previously described
masking procedure which allows the gel interface to be
precisely positioned inside the electrophoresis channel.47An
electric field of around 26 V cm21was used for the separation.
In all separations, no additional surface treatment of the
microchip glass channels was applied. A solution of 1.0X Tris-
borate-EDTA (Bio-Rad, Hercules, CA, USA) was used as the
running buffer. All reagents were used as received from the
polyacrylamide (ReproGel,
Image processing
The migrating DNA bands were detected by their fluorescence
using an Olympus SZX-12 fluorescence stereoscope with a
mercury arc illumination source, and a CCD camera
(Hamamatsu C2400-08 SIT, Hamamatsu Corporation, USA,
Bridgewater, NJ) for imaging. The camera output was
recorded, digitized, and intensity profiles corresponding to
the migrating bands were obtained by extracting the variation
in fluorescence intensity with time at a fixed location in the gel
using Transform 2D image analysis software (Research
Systems, Inc., Boulder, CO). Snap shots of the gel were
digitized to obtain the electropherogram (intensity profile
along the channel). Control DNA fragments added to the
reaction mixture serve as internal size standards.
Results and discussion
Integrated genetic analysis device
The integrated genetic analysis device (Fig. 1) performs two
temperature controlled serial reactions followed by a gel
electrophoresis. The overall operation of the device, shown in
Fig. 3, begins with loading specified amount of reagents and
sample through the inlet ports (L1, L2, and L3). The test
sample and PCR reagents are mixed and positioned in the
PCR chamber. The reaction chamber is then sealed by flanking
phase-change valves (V3and V5). The solution is thermocycled
using integrated thin film heaters and temperature sensors
under the PCR chamber. After thermal cycling, the valves are
opened and the reaction product is moved forward. A portion
of the PCR reaction product is mixed with an equal volume of
the restriction digest reaction reagent (loaded through L3) and
positioned in the restriction digest chamber where the second
reaction occurs. The restriction digest reaction product is then
moved to the separation section where it is electrophoretically
As a demonstration, we have used the device for influenza
A/LA/1/87 strain identification. Fig. 4 shows an electro-
pherogram of an A/LA/1/87 DNA sample obtained from a
complete on-chip analysis. The band pattern obtained after
135 s (separation distance , 1 mm) shows 145 bp and 545 bp
bands that are Hpa1 restriction digestion products, and 343 bp
and 996 bp bands that are internal markers. This pattern
indicates that the original sample for this run was A/LA/1/87
influenza viral DNA. Runs starting with A/Sydney/5/97 viral
DNA sample produce the two marker bands and a single
amplicon band at 690 bp (data not shown). It should be noted
that we worked with viral DNA samples from reverse
transcription instead of the viral RNA due to the difficulty
of sample handling. In separate experiments we have demon-
strated successful RT-PCR starting with viral RNA.
Besides the influenza A/LA/1/87 strain identification, we
have tested three other amplification reactions on the device.
These include amplification from human genomic DNA
(D1S80 locus), mouse plasmid DNA (SNRPN locus), as well
as plasmid DNA of influenza viral strain A/Sydney/5/97.
D1S80 is a simple sequence repeat polymorphism locus present
on human chromosome 1 that contains a 16-nucleotide
sequence variably repeated between 16 and 40 times in the
human population.46Similar repeat-based polymorphic loci
are used in the most common DNA diagnostic and forensic
tests. The mouse Snrpn gene encodes polypeptide N associated
Table 1 PCR primer sequences and product sizes
AssayForward primer Reverse primerProduct
Human D1S80
Mouse SNRPN
690 bp
y650 bp
996 bp
4 | Lab Chip, 2005, 5, 1–9 This journal is ? The Royal Society of Chemistry 2005


text:
QRV information Between 20 Mar 2015 and 25 Mar 2015 Only SSB. 14,18,21,24,28MHz QSL information Only Direct via QSL manager Please send your QSL card, two GSs and SAE as follows A4maniax c/o Sankei kousinsya Co,.Ltd. 6-20-6, Hirai, Edogawa-ku, Tokyo 132-0035 JAPAN Read more
"NCDXF is pleased to announce a $50,000 grant to the Intrepid-DX Group to activate South Sandwich and South Georgia Islands in 2016." Souce: Facebook The Intrepid-DX Group is proud to announce a major ham radio expedition to two rare entities in January-February 2016. South Sandwich Island and South Georgia Island are two of the most remote places on […] Read more
In the name of the HSDXA under the developed skills, a ham radio of the ham radio of thailand in the dwellings. Shh the signed it if, E20HHK judges the activities. of. – me. have na friends. Ham radio group to broadcast to the cat in name Maeo Island AS- 145 IOTA DXpeditions by […] Read more
Advance registration for those planning to attend the International DX Convention in Visalia, California, April 16-19, ends on April 8. An updated list of those already registered to attend is on the convention website! The Visalia program includes DX University and Contest Academy sessions, ARRL, DX, and Contest forums, and a long list of DX and technical presentations, as well […] Read more
José, LU1FM will be active from San Andres Island. 21 March – 4 April 2015 as 5JØB. He will operate on 160 – 6m SSB, Digital modes. He will parcipicate in CQ WPX SSB Contest. He will use IC 7100 transc. with ALS 500 MRX amplifier. Antennas- Hexbeam on 6-20m, dipoles 160, 80, 40m. […] Read more
Vanuatu as YJ0MT from 21 April till 5 May 2015. Location will be Port Vila, the capital city in Efate Island (OC-035). Operator: JA0RQV, Masato Tamura (Masa) Schedule: 21 April to 5 May 2015 Band/Mode: 40m-6m/CW, SSB mostly CW Rig/ANT: IC-7000(100W)/Vertical Operation style: Business in weekday (Serious in weekend) QSL info.: VIA H/C JA0RQV, Bureau/SASE(w/USD2)/LoTW later [Mailing address] Masato […] Read more
IK0FVC, IK0PRG, S53R and S53T will be active from the Vatican during the CQ WW 160m SSB contest as HV0A. They will also QRV outside the contest on CW, low bands. QSL via IK0FVC. Source: http://www.dx-world.net/ Read more
Audio version Listen to an audio interview with N5UP from the RAIN Report and learn about eQSL.cc Purpose eQSL.cc is the first and only global electronic QSL card exchange for amateur radio operators and SWLs. It is designed to be the fastest, easiest, and cheapest way to exchange QSO confirmations, eliminating the cost and time that regular QSL […] Read more
PR70FEB is the 3rd of 12 Special Event Station to celebrate 70 years end of World War II that we will use in 2015. Each month in 2015 we will activate a different callsign and also send 12 different QSLs. The Special Event Stations are to remember the importance of = PEACE = , and in honour to […] Read more
Event celebrating 450 years of the city of Rio de Janeiro PQ450RIO The city of Rio de Janeiro complete this March / 2015, 450 years of existence. Many events will be held during the month and throughout the year. Hams the city of Rio and around the state can not be left out of […] Read more