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  Clinical Abstracts





December 2012

Deborah Sesok-Pizzini, MD, MBA
Tina Ipe, MD, MPH

Digital PCR analysis of maternal plasma to detect sickle cell anemia Digital PCR analysis of maternal plasma to detect sickle cell anemia

Sickle cell anemia is caused by homozygosity for the hemoglobin S mutation, which results in polymerization of the hemoglobin S protein when it is deoxygenated. Prevalence of the mutation is highest in African countries, but due to the changing mobility of populations, sickle cell anemia is becoming more common in other countries, such as the United Kingdom. In fact, sickle cell anemia is now the most common indication for invasive prenatal diagnostic testing in the United Kingdom. Finding a noninvasive test for prenatal diagnosis will help with the management of these affected fetuses and provide a safer method for determining fetuses at risk. The authors designed probes to differentiate between wild-type hemoglobin A and mutant hemoglobin S alleles encoded by the hemoglobin beta gene in cell-free fetal DNA (cffDNA) isolated from maternal plasma. Maternal plasma contains about 10 percent of cffDNA. Using cffDNA is an alternative to sampling chorionic villi or amniocytes, both of which carry risks of miscarriage. The authors noted that the definitive diagnosis of autosomal recessive conditions in cases in which both parents carry the same mutation is more complex because the majority of mutant alleles in plasma will be maternal in origin. Therefore, prenatal diagnosis with cffDNA in these circumstances necessitates the determination of allelic ratios in maternal plasma rather than the presence or absence of a mutation not carried by the mother. For their study to determine the feasibility of using digital PCR for noninvasive prenatal diagnosis in pregnancies at risk for sickle cell anemia, the authors used real-time PCR and informative markers for male and female fetuses. While digital PCR was used to quantify the fractional fetal DNA concentrations for male fetuses, female fetuses required additional informative indel markers. The study showed that using noninvasive prenatal diagnosis correctly classified fetuses at risk for sickle cell anemia with an accuracy rate of 80 percent. When a fractional fetal DNA concentration of more than seven percent was used, the correct classification was 100 percent. The authors noted that promptly processing samples and optimizing sample preparation may improve the reliability of results. They concluded that additional studies and informative markers are needed to expand the use of noninvasive prenatal diagnosis to autosomal recessive diseases, especially for female fetuses.

Barrett AN, McDonnell TCR, Allen Chan KC, et al. A digital PCR analysis of maternal plasma for noninvasive detection of sickle cell anemia. Clin Chem. 2012;58(6):1026–1032.

Correspondence: Lyn S. Chitty

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MicroRNAs in CSF identify brain cancers and reflect disease activity Quality standards for sample collection in coagulation testing

Preanalytical variables are often the cause of testing errors in clinical pathology laboratories. Monitoring sample collection and the quality of samples submitted to the laboratory is important for ensuring the accuracy of test results. For many types of test results, the same quality measures apply, such as proper patient identification, proper use of sample collection tubes, and correct technique. For laboratories performing coagulation testing, additional preanalytical variables must be considered and controlled to minimize erroneous and variable results. The authors reviewed preanalytical errors that may impact coagulation testing and offered evidence-based suggestions and recommendations for obtaining high-quality test results. The authors’ recommendations addressed patient preparation factors, as well as prevention of misidentification errors, use of correct techniques, appropriate venous stasis, appropriate devices and needles, prevention of hemolyzed specimens, order of blood draw, and tube mixing. The authors noted that the most common preanalytical problems are samples not arriving in the lab, hemolyzed specimens, clotting, and inappropriate tube filling. Due to activation of the clotting cascade, hemolyzed specimens are of particular concern. The authors support the Clinical and Laboratory Standards Institute’s recommendations to reject these samples. The authors performed their own study to evaluate bias in coagulation test results when specimen collection tubes were incompletely filled. Their results showed that there was clinically significant bias for activated partial thromboplastin time (APTT) at less than 89 percent of fill volume, fibrinogen at less than 78 percent, and coagulation factor VIII at less than 67 percent. Prothrombin time (PT) and activated protein C resistance were still accurate even at fill volumes of 67 percent. The authors recommended rejecting specimens due to compromised patient identification, hemolyzation of specimens, wrong specimen tube, or inappropriate blood-to-additive ratio.

Lippi G, Salvagno GL, Montagnana M, et al. Quality standards for sample collection in coagulation testing. Semin Thromb Hemost. 2012;38(6):565–575.

Correspondence: Dr. Giuseppe Lippi at or

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Very early diagnosis of chest pain using point-of-care testing Screening for prostate cancer: USPSTF recommendation statement

Prostate cancer is the most commonly diagnosed non-skin cancer in American men, with a lifetime risk of 15.9 percent. However, the lifetime risk of dying from this cancer is only 2.8 percent. The majority of deaths due to prostate cancer occur after age 75. The author reviewed the current recommendations of the U.S. Preventive Services Task Force (USPSTF) with regard to prostate cancer screening and provided an update of the 2008 recommendation statement. The task force reviewed recent findings on the benefits and harms of prostate-specific antigen (PSA)-based screening and the benefits and harms of treatment for localized disease. The author, reporting on behalf of the USPSTF, stated that PSA-based screening for prostate screening is no longer recommended, regardless of a man’s age. However, the task force noted that PSA screening for surveillance after diagnosis or treatment may still be indicated, but this decision is beyond the scope of this current recommendation. Prior to this publication, recommendations for prostate cancer screening incorporated serum PSA levels, digital rectal exams, and ultrasonography. However, the USPSTF concluded that there was convincing evidence that PSA-based screening programs detected many cases of asymptomatic cancer that were expected to remain asymptomatic for a man’s lifetime. To that end, it is believed that PSA screening caused an overdiagnosis in many men. Two of the largest trials estimated an overdiagnosis rate of 17 to 50 percent for prostate cancer screening. The author and her colleagues noted that the PSA test often produces false-positive results, leading to unnecessary anxiety in men. PSA levels that trigger a biopsy may result in complications associated with the procedure. In some cases, this may even result in unnecessary surgery, with its inherent risks.

Moyer VA, on behalf of the U.S. Preventive Services Task Force. Screening for prostate cancer: U.S. Preventive Services Task Force recommendation statement. Ann Intern Med. 2012;157:120–134.


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Clinical pathology abstracts editors: Deborah Sesok-Pizzini, MD, MBA, associate professor, Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, and medical director, Blood Bank and Transfusion Medicine, Children’s Hospital of Philadelphia; Tina Ipe, MD, MPH, resident, Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania.