For the complete article, please refer to The Swazi Observer.

South Africa will treat all HIV-positive babies and expand testing, the president announced Tuesday, a dramatic and eagerly awaited shift in a country that has more people living with HIV than any other.

President Jacob Zuma’s speech on World AIDS Day was viewed as a definitive turning point for a nation where the previous administration distrusted drugs developed to keep AIDS patients alive and instead promoted garlic treatments. One Harvard study said that resulted in more than 300,000 premature deaths.

Zuma compared the fight against AIDS to the decades-long struggle against the apartheid government, which ended in 1994 with the election of Nelson Mandela in the country’s first multiracial elections.

“At another moment in our history, in another context, the liberation movement observed that the time comes in the life of any nation when there remain only two choices: submit or fight,” Zuma said. “That time has now come in our struggle to overcome AIDS. Let us declare now, as we declared then, that we shall not submit.”

Zuma was greeted with a standing ovation when he entered a Pretoria exhibition hall filled with several thousand people.

In some ways, Zuma is an unlikely AIDS hero. In 2006, while being tried on charges of raping an HIV-positive family friend, he was ridiculed for testifying that he took a shower after sex to lower the risk of AIDS. He was acquitted of rape.

Zuma, a one-time chairman of the country’s national AIDS council, may never live down the shower comment. But he has won praise for appointing Dr. Aaron Motsoaledi as his health minister. AIDS activists say Motsoaledi trusts science and is willing to learn from past mistakes.

UNAIDS executive director Michel Sidibe, who took the podium shortly before Zuma, told the president: “What you do from this day forward will write, or rewrite, the story of AIDS across Africa.”

On Tuesday, in response to a plea from Zuma, the United States announced it was giving South Africa $120 million over the next two years for AIDS treatment drugs.

Zuma said in a speech broadcast across South Africa on state radio and television that the new policy changes would take effect in April.

“It means that people will live longer and more fulfilling lives,” he said.

South Africa, a nation of about 50 million, has an estimated 5.7 million people infected with HIV.

The new steps include treatment for all HIV-positive children under 1 year old, and earlier treatment for patients infected with both the virus that causes AIDS and tuberculosis, and for women who are pregnant and HIV-positive.

Zuma said all health institutions, not just specialist centers, would provide counseling, testing and treatment.

He also called on South Africans to get tested for HIV. But, contrary to speculation in recent days, he did not take an HIV test Tuesday.

“I have taken HIV tests before and I know my status,” he said. “I will do another test soon as part of this new campaign. I urge you to start planning for your own tests.”

The health minister under Zuma’s predecessor distrusted drugs developed to keep AIDS patients alive, instead promoting garlic treatments. Zuma’s government has set a target of getting 80 percent of those who need AIDS drugs on them by 2011.

A Harvard study of the years under President Thabo Mbeki, who questioned the link between HIV and AIDS, concluded that more than 300,000 premature deaths in South Africa could have been prevented had officials here acted sooner to provide drug treatments to AIDS patients and to prevent pregnant women with HIV from passing the virus to their children.

After Zuma won a power struggle within the governing African National Congress, the party forced Mbeki to step down late last year after almost a decade as president. Zuma took over after elections in April.

Setjhaba Ranthako brought his 4-year-old daughter Tshegofatso to hear Zuma’s speech, saying education should start early.

“I’ve see in President Zuma a person who’s willing to listen, and say, `Here I am, come with your views, and let’s turn your views into an effective campaign to combat the spread” of AIDS, said Ranthako, who works with a group that raises awareness about AIDS among men.

After listening to his president, advertising consultant Tedson Tibani said the steps Zuma outlined could significantly reduce infections within a few years. Tibani said putting more people on drugs would cost money, but said he was hopeful others would follow the U.S. in donating money.

“There’s a kind of hope the president has instilled,” Tibani said. “I’m very happy with that. We’ve never had that before.”

The crowd that had greeted Zuma like a rock star before his speech rose to their feet when Zuma finished Tuesday. Then he danced along with a choir that sang: “Zuma, you are blessed.”

Many people blame the policies of the former South Africa President, Thabo Mbeki, for the widespread AIDS epidemic in the country. Mbeki often disputed the link between HIV and AIDS and was against the widespread distribution of antiretroviral medications. Instead he encouraged HIV-positive people to eat garlic and beet roots, and to drink lemon juice to deter HIV. People believe that Mbeki’s attitudes and actions could have contributed to as many as 330,000 early deaths.

President Zuma, who was the Vice President to Mbeki, is trying to lengthen the gap between his and his predecessor’s policies. South Africa offers anti-retroviral (ARV) treatments to 700,000 HIV patients, over double the 216,000 patients from last year. At roughly $1,500 a year for each treatment it can get a bit pricey. Fortunately, foreign donors have often provided treatments free of charge, and the US ambassador to South Africa has announced that the United States will contribute another $120 million for next year’s AIDS treatment and research.

* For the complete article, please visit http://stdtestingblog.com/original-articles/

Human Immunodeficiency Virus, or HIV, is the virus that causes AIDS, and is at the forefront of research in many fields.  One of the most interesting topics of research is anti-retroviral therapy (ART) implemented in HIV positive patients in order to reduce the effect of the virus.

The newest breakthrough entails using snake and insect venom as a form of ART. A major component in bee venom inhibits replication of both CXCR4 and CCR5 HIV-1 in human CD4 cells. Phospholipase A₂ (PLA₂), which is found in the venom of many snakes, has been shown to block viral entry into cells.

The exact mechanism, whether enzymatic or simply competing for a binding site, is still in the process of being worked out.  Aside from this, the most important details have shown promising signs for the field of research dedicated to HIV treatment.

This discussion further reiterates the needs and necessity for regular, comprehensive STD testing.  If everybody got tested for STDs on a regular basis, the incidence of HIV (among other STDs) would be considerably lower.  Testing is simple, and should be done (as according to the CDC) every six months to one year, or in between sexual partners.

For the complete article, please refer to the original articles page.

Bedenbaugh was sentenced to six years in prison for not telling his ex-wife, to whom he was married for five years, that he had HIV.  This exposed her to possible infection.  Instead, he told her that the medicine that he took was for a blood disease.  She remains uninfected.

It is illegal to knowingly engage in sexual intercourse with another person without first informing them of an HIV infection.

In 2006, Bedenbaugh was convicted for  inappropriate contact with a 13-year-old girl in November 2006.  More recently, in 2008, he was investigated for  an alleged sexual assault on a juvenile in 2008.  This brought about attention to his medical history, which indicated that he had HIV.

For the original article, please refer to original articles.

According to the CDC, everyone ages 13 to 64 should do an HIV test at least once. People who have had more than one partner since their last STD test, and people who’s partner has had more than one partner should get tested regularly. Also at higher risk are men who have sex with men (MSM) and people who already have an STD. These people should also get tested at least once a year.

Women should get an annual Pap test beginning when they are 21-years of age, or earlier if they are sexually active. The Pap test is a screening tool used by doctors to detect cancerous changes of the cervix, which is a common result of the common human papillomavirus (HPV). HPV usually does not show any symptoms; however, it can lead to genital warts in both men and women. Unfortunately there is no screening test for HPV in men. Men who think they have symptoms of HPV should see someone about a physical exam.

In addition to HPV, the CDC suggests that sexually active women under 25 should get tested for chlamydia and gonorrhea at least once a year. Regular screening is also recommended for men who have sex with men, older women who have had multiple partners, and pregnant women.

Syphilis rates reached a record low in year 2000; however, they have been increasing since causing the CDC to suggest syphilis screening for individuals who feel they are at risk.

*For the complete article, please visit http://hivtestingblog.com/original-articles/

*For the complete article, please visit http://hivtestingblog.com/original-articles/

Stanford did not have standing to file suit because, the court determined, Roche owned the patents at issue.

A lower court, the US District Court for the Northern District of California, was instructed to dismiss the suit. The district court had ruled in 2008 that the patents were invalid, but did not agree with Roche’s claims of ownership. According to the appeals court, the district should not have addressed the patents’ validity because Stanford didn’t own the IP to begin with.

This was considered, by many, to be a complete victory for Roche.

Stanford first sued Roche in 2005, seeking more than $200 million for the alleged infringement of three patents assigned to Stanford — US Nos. 5,968,730; 6,503,705; and 7,129,041. The three patents descend from a common parent application and share the same title: “Polymerase Chain Reaction Assays for Monitoring Antiviral Therapy and Making Therapeutic Decisions in the Treatment of Acquired Immunodeficiency Syndrome.”

The test was developed, and is currently used, to measure the viral load of HIV in a person’s blood, by measuring the amount of HIV RNA present in the bloodstream. The original description of the use of the PCR to measure HIV RNA was published in the Journal of Infectious Disease in 1991.

For the complete article, please refer to www.hivtestingblog.com/original-articles/

“I went to the doctor and had an AIDS test and he told me it was positive,” he said, in an interview in this month’s Glamour magazine. “That was one of the worst days of my life.”

The doctor did a confirmatory test, which came back negative.  The doctor attributed the results first test, the false positive, to the heavy drinking and drug-taking lifestyle he used to enjoy tampering with his immune system.

“It turned out that because I was drinking and using drugs so much, my immune system had dropped so that it was a borderline result. When I went back to be tested again it was negative.”

It is also believed that Ozzy’s daughter Kelly referred to him when she broke down at an AIDS charity benefit in London two years ago.

“This charity is really important to me because one of my family is HIV positive,” she said at the time. “And I’m so proud of him.”

Ozzy is currently promoting his autobiography, I Am Ozzy, and many revelations have come out about his life, including the news that he still enjoys conjugal trysts with wife Sharon but struggles to bring the liaisons to a satisfactory conclusion.

*For the original article, please refer to http://www.hivtestingblog.com/original-articles/

A Clarksville woman was arrested Thursday.  She was charged with knowingly exposing a man to HIV.

The woman, Donyel Da’Shawn Brown, was charged with criminal exposure to HIV, and her bond was set at $1 million.

The arrest warrant states a man reported Brown knowingly had unprotected sex with him for four years without telling him she was infected.

Also, Brown and the man had a child during the time she was diagnosed with HIV, the warrant said, but has not been determined if the child has HIV. Whether the child and man were infected has not been determined, Knoll said.

The state law, which took effect in 1994, does not require “the actual transmission of HIV” for someone to be convicted. Brown can be acquitted if she can prove that her partner had prior knowledge of her HIV status.

If convicted, Brown could face jail time of three to six years.

For the complete article, please refer to www.hivtestingblog.com/original-articles/

The test, which was approved by the U.S. Food and Drug Administration in September 2009 is licensed for screening donated blood and blood specimens, and for screening specimens from organ donors. Prior to 1985, there were no tests available to screen for HIV in blood and organ donations, but nowadays donors and organs go through extensive testing. “The risk of acquiring HIV from a blood transfusion… is estimated to be 1 in 4 for every 600,000 transfusions. The risk… from organ transplantation is probably similar.”

The two most common blood tests used to previously detect HIV were the enzyme-linked immunosorbent assay (ELISA), and the Western blot assay, which are both antibody tests. These tests can only tell if a person has been infected with HIV, not how long they’ve been infected or if they have AIDS (acquired immune deficiency syndrome, the final stage of HIV disease).

This new HIV test will help decrease the number of HIV transmissions through blood transfusions and organ donations, and  help make the world a safer place.

*For the complete article, please refer to http://hivtestingblog.com/original-articles/

For more than 20 years, the Burlington nonprofit has helped provide Vermonters with HIV/AIDS everything from a ride to the doctor’s office to a new home. Vermont CARES also will pay the rent for an infected individual, if the need arises. The organization held a rally Saturday to show support for more than 450 Vermonters diagnosed with the disease. People at the rally wore red shirts and stood in a ribbon formation at the University of Vermont’s Redstone Campus.

*For the complete article, please refer to http://hivtestingblog.com/original-articles/

Alfredo Idiarte of CHAI said his organization is committed to the program and will work towards making it more comprehensive. He also said that CHAI is looking at providing deoxyribonucleic acid Polymerase chain reaction (DNA PCR) tests for infant diagnosis. The DNA PCR test looks for direct (DNA) evidence of the virus rather than the antibodies. Currently the tests do not detect evidence of the virus in children younger than 18 months. Early detection could drastically improve treatment options.

*For the original article please refer to http://hivtestingblog.com/original-articles/.

In laboratory studies, the Amplicor test was able in some cases to detect as few as 400 copies of HIV DNA in a blood sample and could regularly detect m800 or more copies.

Two clinical trials of Amplicor were conducted in patients with advanced HIV infection who had received no antiviral treatment or treatment for less than 16 weeks. The results showed that high viral load before anti-HIV therapy, or large increases in viral load after treatment, correlated with increased risk of disease progression to full-blown AIDS.

Early diagnosis can help get these children onto a treatment plan, which at a young age can prove the difference between “surviving and thriving”. If a child born to an HIV infected mother, the first HIV DNA PCR test is performed at 6 weeks of age. If that PCR test returns a positive result it is repeated for a confirmation. If the PCR test returns a negative result the first time around, a confirmation PCR test is to be performed at 6 months of age. These new developments in testing and treatment options could possibly save many, many young lives.

*For the complete article please visit http://hivtestingblog.com/original-articles/

“Broadly neutralizing antibodies” (bNAbs) are not very common. Only six have been identified so far, but two new ones with an “unusual potency” have been discovered in one sub-Saharan African donor. The antibodies, named PG9 and PG16, have characteristics that open possibilities for AIDS vaccine research. PG9 and PG16 were found to possibly prevent infection in more than 70% of 162 viral strains tested in cell culture. The antibodies were found to work at levels lower than the best characterized bNAbs so far.

The donor himself did not benefit from the antibodies, seeing as how they are no match against HIV once the infection is established, but researchers are hoping that understanding how PG9 and PG16 stop the virus will help vaccine designers further their work on an HIV/AIDS vaccine. While an effective and consistent HIV vaccine may be far from sight, the discovery of these new bNAbs definitely provides a huge leap in progress in research.

*For the complete article, please refer to http://hivtestingblog.com/original-articles/

Healthcare workers are often exposed to the virus at work; however, it is unlikely that they will contract the virus from a patient. Since December 2001, there have been only 57 documented reports of patient-to-worker HIV transmission, mainly due to precautionary guidelines that healthcare workers follow. The main risk of transmission for healthcare workers  is through accidental needle sticks or other injury with a contaminated instrument. However, even here the risk is small. “Researchers estimate that only about 0.0-1% or healthcare workers” contract HIV from an accidental needle stick.

This low statistic can be attributed to post-exposure prophylaxis (PEP), which can be taken immediately after exposure to reduce the risk of transmission. PEP uses antiretroviral therapy (ART) to prevent transmission, but often comes with serious side effects including dizziness, fatigue, nausea, vomiting, diarrhea and more. Current antiretroviral drugs cannot cure HIV infection, nor reduce the risk of transmitting it to someone else, but they can suppress the virus to undetectable levels in some cases. It has been estimated that PEP reduces the infection rate among healthcare workers by 79%.

Post-exposure Prophylaxis should begin immediately after the exposure, seeing as how PEP is most effective when it is initiated within two t0 four hours of exposure. The specific dosage of medication depends on a couple factors including the patient’s overall health, the severity of exposure, the availability of antiretrovirals, and if the patient has any known or possible cross-resistance to any drugs. Treatment normally lasts no less than two weeks and no longer than four. Studies show that almost a quarter of those receiving PEP stop taking the medications early because of side effects. As with all forms of treatment, it is less effective if it ends early.

HIV tests should be performed after any risky sexual behavior, even if PEP was used. Immediately after HIV enters the body antibodies are produced to fight off the infection. While these antibodies cannot completely eliminate the virus, we can use their presence to see if HIV is in the blood. Most people develop detectable antibodies within two to eight weeks; however, it may take longer in some people. Most often, the enzyme immunoassay (EIA) test is used to detect HIV antibodies. If a positive result is returned it is confirmed with a follow-up test before making a diagnosis. Typically the Western blot test is used to confirm a positive HIV result. Other testing options include DNA or RNA tests, which instead of looking for antibodies actually look for genetic material of HIV. These tests can be used for early detection of HIV.

With the combination of healthcare precautions and treatment options such as PEP, we have the ability to decrease the number of patient to worker HIV transmissions drastically.

*For the complete article please refer to http://hivtestingblog.com/original-articles/

The report involved a 38-year old gay man who reported having unprotected anal sex with multiple partners in the previous 48 hours. The patient was treated with Truvada as post-exposure prophylaxis. During his treatment the patient reported more episodes of risky sex, causing his treatment to be extended. During his treatment the patient was repeatedly tested for HIV. He received a couple negative HIV results, but after repeated exposures the patient tested HIV-positive.

The patient received three viral load tests shortly after his positive HIV test result. The viral load turned out to be extremely low, and his CD4 count was high. These results were out of the ordinary for someone with an acute HIV infection, and the patient had no HIV seroconversion symptoms. Several more tests were performed on the patient and all of them returned similar findings.

The authors of the article report that the patient’s HIV infection was weaker than usual, and that this result was most likely due to the antiretroviral therapy he was receiving.

*For the complete article please refer to http://hivtestingblog.com/original-articles/

Similar to the guidelines following occupational exposures, the CDC recommends prophylaxis beginning within 72 hours after the initial exposure with any body fluids from an HIV infected person. If 72 hours has passed after the exposure, the CDC recommends not starting prophylaxis. If the HIV status of the contact person is unknown, but the exposure has an elevated risk the CDC suggests the decision of whether to begin prophylaxis be made on a patient-to-patient basis.

Unfortunately many people are unaware that they are infected with HIV, and because of the 72 hour window period for prophylaxis these people are out of luck when they discover they were exposed. People who benefit the most from prophylaxis are those who know they have been exposed, including sexual assault victims and intravenous drug users.

Some clinicians have fears that people will use post exposure prophylaxis as a “safety net” for unprotected sex, but that is not at all it’s intended purpose.

*For the complete article please refer to http://hivtestingblog.com/original-articles/

While the officers believe that Crawford is HIV-positive, they aren’t at any major risk, seeing as how studies have shown that HIV/AIDS is not carried in saliva. Crawford is being charged with two counts of harassment with a bodily substance — a felony.

*To view this article in it’s entirety, please visit http://hivtestingblog.com/original-articles/

The gel was developed to “enable women to protect themselves against HIV without approval of their partner”. To use it the woman would only need to insert the gel a few hours before sex. The gel flows easily at the vaginal pH, and becomes more solid as the pH increases to that of semen. As the gel turns solid it traps AIDS virus particles, preventing them from infecting vaginal cells.

Due to several factors, women often have difficulty convincing their partners to use protections; however, with this new gel the woman wouldn’t even have to negotiate. It is a form of protection she can use on her own, without her partner’s knowledge.

If all goes as planned, the scientists estimate the gel will be tested in humans within three to five years, and it will be available in the years after that.

*For the complete article please visit http://hivtestingblog.com/original-articles/

Polymerase chain reaction (PCR) has rapidly become one of the most widely used techniques in molecular biology and for good reason: it is a rapid, inexpensive and simple means of producing relatively large numbers of copies of DNA molecules from minute quantities of source DNA material–even when the source DNA is of relatively poor quality.

PCR involves preparation of the sample, the master mix and the primers, followed by detection and analysis of the reaction products. These steps are discussed below.

Sample Preparation

PCR is very versatile. Many types of samples can be analyzed for nucleic acids. Most PCR uses DNA as a target, rather than RNA, because of the stability of the DNA molecule and the ease with which DNA can be isolated. By following a few basic rules, problems can be avoided in the preparation of DNA for the PCR. The essential criteria for any DNA sample are that it contain at least one intact DNA strand encompassing the region to be amplified and that any impurities are sufficiently diluted so as not to inhibit the polymerization step of the PCR reaction.

Although any protocol is acceptable for PCR purposes, it is often best to use the fewest steps possible in DNA preparation in order to prevent accidental contamination with unwanted DNA. Usually a 1:5 dilution of the sample with water is sufficient to dilute out any impurities which may result from the purifying protocol.

The simplest method of isolating DNA from cells is as follows:

1. Cells can be obtained by using a toothpick to scrape under the fingernails, swabbing the inside of the mouth or from the roots of plucked hairs. Regardless of source, cells are resuspended in 20 ul of water. Skip to step four.
2. If you are using cells suspended in media, centrifuge at 1200- 1500Xg for 5 minutes. Resuspend the cell pellet in 1 ml of phosphate buffered saline (PBS) and repellet by spinning at 1200- 1500Xg for 5 minutes. Repeat. These PBS washes remove medium, and its inhibitory factors, from the surface of the cells. After the last wash resuspend the cell pellet in 20 ul of distilled water. Be aware that too much cell debris can inhibit the PCR reaction. If this happens, it may be necessary to further dilute the DNA sample. Go to step four.
3. For bacterial samples take a toothpick and scrape the teeth, or swab the throat, ears or between the toes. Resuspend material in 500ul of water. Freeze and thaw sample three times with vigorous shaking or vortexing between repetitions to break the bacterial cell wall. Although not all DNA will be released from the cells, there will be a sufficient quantity for PCR. Go to step four.
4. Place the sample in a 95oC heating block, or in boiling water, for 5 minutes. This step inactivates the DNase molecules that are found in the sample preparation. If left intact, DNase could clip the desired DNA template molecule into fragments which would be unsuitable for PCR. If there is very little DNA in the sample preparation, the DNA can be concentrated by ethanol precipitation. The sample is now ready for PCR.

DNA samples for PCR–regardless of preparation method–are generally run in duplicate in order to provide a control for the relative quality and purity of the original sample. Adding a small amount of DNA to the control just after the master mix step allows the detection of anything in the completed sample prep which would inhibit the PCR reaction.

Preparation of Master Mix

The Master Mix contains all of the components necessary to make new strands of DNA in the PCR process. The Master Mix reagents include:

http://www.accessexcellence.org/LC/SS/PS/PCR/PCR_technology.php

Notes on the Master Mix

The Master mix buffer is often stored as a 10X stock solution (100 mM Tris-HCL, pH 8.3, 500 mM KCL, 1.5 mM MgCl2) which is diluted to 1X for use. Both the Master mix buffer and the purified water can be stored at room temperature. Store deoxynucleotides, primers and Taq DNA polymerase enzyme at -20oC.

Although 100ul of master mix per reaction is generally used, it is possible to use as little as 25 or 50ul to save on cost of reagents. Regardless of the total volume, be certain to keep the final concentrations of reagents constant.

Master mix reagents can be optained from a variety of companies. Often the initial concentration of the reagent will differ depending on which company produced it. It is easy to figure out how much stock reagent to use by following a simple formula:

(initial concentration) X ( volume needed ) =

                                    (final concentration) X (volume of sample)

For example: I have 10X buffer, 10 mM of each nucleotide, 0.5 mM primers and Taq DNA polymerase at 5 Units/ul. I want to make one 50 ul reaction. Calculations are as follows:

10 X buffer: (10X) X (5 ul) = (1X) X (50 ul) Nucleotides: (10,000 uM) X (1 ul) = (200 uM) X (50 ul) (10mM=10,000uM) primers (500uM) X (O.1ul)= (1.0uM) X (50 ul) Since it is impossible to pipet 0.1ul accurately, a dilution needs to be made first. Add 10 ul of stock primer solution to 990 ul of water to get 5uM concentration of primers. This new primer dilution can be stored at 4oC. Calculation for 5uM stock: (5uM) X (10 ul) = (1.0 uM ) X (50 ul) Taq DNA polymerase (5Units/ul) X ( 0.25 ul) = (.025 Units/ul) X (50 ul) 2.5 Units/100ul= Since it is impossible to pipet 0.25ul accurately, a .025 Units/ul dilution needs to be made first. Add 1.25 ul stock to 3.75 ul water to get a 1.25 Units/ul concentration. Discard and make fresh with each use. Calculation for 1.25 Units/ul stock: (1.25 Units/ul) X (1 ul) = (.025 Units/ul) X (50 ul) To make the master mix for one reaction add:

• 5 ul 10X buffer
• 4ul Each nucleotide (1ul each of dATP, dCTP, dGTP, dTTP))
• 20 ul Each primer (10ul of each)
• 1 ul Taq DNA polymerase (Total volume = 30ul)
• add 15 ul of water
• 5 ul of template (Total volume = 50 ul)

If want to make 3 reactions, 3 X 50ul = 150ul. Use this number in the formula for “volume of sample.”

Primers

A primer is a short segment of nucleotides which is complementary to a section of the DNA which is to be amplified in the PCR reaction. Primers are annealed to the denatured DNA template to provide an initiation site for the elongation of the new DNA molecule. Primers can either be specific to a particular DNA nucleotide sequence or they can be “universal.” Universal primers are complementary to nucleotide sequences which are very common in a particular set of DNA molecules. Thus, they are able to bind to a wide variety of DNA templates.

Bacterial ribosomal DNA genes contain nucleotide sequences that are common to all bacteria. Thus, bacterial universal primers can be made by creating primers which are complementary to these sequences.
Examples of bacteria universal primer sequences are:
Forward 5′ GAT CCT GGC TCA GGA TGA AC 3′ (20 mer)
Reverse 5′ GGA CTA CCA GGG TAT CTA ATC 3′ (21 mer)

Animal cell lines contain a particular sequence known as the “alu gene”. There are approximately 900,000 copies of the alu gene distributed throughout the human genome, and multiple copies distributed through the genome of other animal cells, as well. Thus, the alu gene provides the sequence for a universal primer for animal cell lines. The alu primer is especially useful in that it binds in both forward and reverse directions.
The alu universal primer seqeunce is as follows:
5′ GTG GAT CAC CTG AGG TCA GGA GTT TC 3′ (26mer)

When using universal primers the annealing temperature on the thermal cycler is lowered to 40-55 degrees C.

Sometimes primer units are listed in optical density reading (OD). If this is a problem you will need to convert to molarity using the following equations: Change optical density reading of primer to molarity (uM units)-

1. N = # of primer bases
2. SIGMA 260 =~ 10,000 X N/ m X cm
3. Molecular weight =~ 330 X N
4. OD₂₆₀ / SIGMA 260 X 10⁶ = Concentration (uM)

For example- primer is 20 bases long/ OD₂₆₀ = 10.

1. N = 20
2. SIGMA 260 =~ 10,000 X 20/m X cm = 20,000/m X cm
3. molecular weight =~ 330 X 20 = 6,600
4. 10 OD₂₆₀/20,000 m^-1cm^-1 X 10⁶ = 50uM

Detection and analysis of the reaction product

The PCR product should be a fragment or fragments of DNA of defined length. The simplest way to check for the presence of these fragments is to load a sample taken from the reaction product, along with appropriate molecular-weight markers, onto an agarose gel which contains 0.8-4.0% ethidium bromide. DNA bands on the gel can then be visualized under ultraviolet trans-illumination. By comparing product bands with bands from the known molecular-weight markers, you should be able to identify any product fragments which are of the appropriate molecular weight.