Antiplatelet therapy with aspirin (ASA) is widely used in cardiovascular disease (CVD) prevention because it inhibits platelet cyclooxygenase-1 (COX-1) and thromboxane A2 (TXA2)-mediated platelet aggregation, but not everyone fully benefit from ASA due to incomplete TXA2 suppression. Among the mechanisms to explain ASA failure (poor ASA response or ASA resistance), oxidative stress and inflammation have emerged as relevant factors. The relationship of urinary 11-dehydro-thromboxane B2 (11dhTxB2), an inactive stable metabolite of TxA2, and oxidative stress (urinary 8-isoPGF2α) in a cohort of stable coronary artery disease (CAD) patients on ASA treatment with the risk of adverse outcomes (mortality) was investigated. In this session, the effect of aspirin on platelet cyclooxygenase COX-1 and thromboxane (TxA2) inhibition, and its use in cardiovascular disease prevention will be reviewed. This review will include current laboratory measurement of urinary 11dhTxB2, a stable inactive metabolite of TxA2, to assess ASA response as a risk factor of CVD including mortality. Study results confirmed that laboratory measurement of urinary 11dhTxB2 represents a strong independent risk factor for all-cause mortality among stable CAD patients on ASA therapy. These results also indicate that 8-isoPGF2α and COX-2 thromboxane production not affected by ASA may maintain platelet hyperactivity irrespective of COX-1 inhibition. Poor ASA inhibition of 11dhTxB2 (ASA resistance) may prompt additional anti-oxidative and/or anti-inflammatory therapy in these patients aimed at modifying 5-year mortality risk.

The GeneStrat® Genomic Test is a blood-based series of ddPCR tests for DNA (EGFR, KRAS, BRAF) and multiplexed assays for RNA variants (EML4-ALK, ROS1, RET).  In this report we will update on test and specimen collection kit design considerations, feasibility and product development studies. We will report on post-launch test performance using the included nucleic acid preserving blood collection tubes.

Free 25OH Vitamin D has received increasing attention over the last four years. More and more studies support the importance of free 25OH Vitamin D, suggesting that this fraction of 25OH Vitamin D is a better measurement of Vitamin D status rather than the total 25OH Vitamin D in pregnancy, renal disease, liver disease and intensive care units. This presentation will review recent evidence-based studies in the areas of supplementation strategy with Vitamin D2 / D3, evaluation of Vitamin D status during pregnancy, respiratory diseases, and monitoring of chronic kidney disease (CKD) patients. 

QuantiFERON TB (QFT) was the first interferon gamma release assay (IGRA) utilizing whole blood stimulation as a test for TB infection, gaining FDA approval in 2001. Subsequently there have been several improvements to the first test in terms of accuracy, ease of use and clinical utility.  There are currently over 1200 independent publications on QFT and it is the most studied IGRA in the world on contact evaluation, epidemiology, vaccine and Pharma development and biomarker research. The 4th generation (QFT-Plus) is currently FDA approved in the USA and is CE marked in Europe. QFT-Plus provides higher specificity than the tuberculin skin test (TST) by incorporating highly specific antigens that do not cross react with BCG and most environmental non tuberculosis mycobacteria. In addition, it has new antigens that stimulate CD8 T cells.

Published studies now supports a role for CD8+ T cells participating in the host defense to MTB by producing IFN-γ and other soluble factors, which activate macrophages to suppress growth of MTB, kill infected cells, or directly lyse intracellular MTB (1-3). MTB-specific CD8+ cells have been detected in subjects with LTBI and with active TB disease where IFN-γ producing CD8+ cells may be frequently found (4-6). Moreover, ESAT-6 and CFP-10 specific CD8+ T lymphocytes are described as being more frequently detected in subjects with active TB disease versus LTBI, and may be associated with a recent MTB exposure (7-9). In addition, MTB-specific CD8+ T cells producing IFN-γ have also been detected in active TB subjects with HIV co-infection (10, 11) and in young children with TB disease (12).  How the incorporation of novel CD8 antigen stimulation technology has proved beneficial in independent performance studies with QFT-Plus in active TB and contact investigations will be discussed.  Maintaining the performance and ease of use of whole blood stimulation builds on the value of QFT and will provide further benefit in the management of latent tuberculosis and TB control programs.

The role of the microbiology laboratory in the processing of positive patient blood cultures has become more complicated with the increasing demand for rapid information to assist in the management of sepsis.  Many laboratories have turned to using molecular systems that report identification and resistant marker results that supply actionable information but no susceptibility data.  These systems have proven over time to be useful with the management of Gram positive bloodstream infections but vary in the ability to properly guide therapy choices for Gram negative bacteremia.   In our institution, phenotypic antimicrobial susceptibility results are often necessary for optimization of therapy. This is particularly true due to emerging resistance mechanisms in Gram negative bacilli, such as AmpC and porin alterations.  The Accelerate Pheno™ system is a truly unique and divergent technology that has the potential to produce final susceptibility results directly from a positive blood culture within 7 hours.  This is a reduction in time to results from conventional susceptibility methods of 2-3 days.   The Accelerate Diagnostics system was FDA cleared in February of 2017.  Clinical trial and scientific presentations have shown the identification and MIC susceptibility results to be reliable and can be used to improve patient care.  Rapid final MIC results provide invaluable information to a stewardship program to rapidly optimize the therapy of sepsis.

This webinar builds and expands on the recent labroots webinar by Dr. Gary Horowitz to emphasize how relatively in expensive assay standardization procedures can be incorporated into clinical and research laboratories.  Serum 25-hydroxyvitamin D [25(OH)D] is a very difficult metabolite to measure.   Assay variation continues to confound the clinical assessment of vitamin D deficiency. The Vitamin D Standardization Program (VDSP) was founded in 2010 by the NIH Office of Dietary Supplements in collaboration with organization and researchers around the world to promote the standardized laboratory measurement of 25(OH)D.  In this webinar, we will discuss the (1) meaning of assay standardization, (2) VDSP Reference Measurement System; (3) Cost-effective Steps to Standardize 25(OH)D Measurement; (4) VDSP Performance Guidelines – A Weakness; and (5) Emerging vitamin D standardization issues.

Acute and chronic pain are often managed with drug therapy, which may include extended use of drug classes such as opioids and benzodiazepines.  Pain does not go away, and may intensify during pregnancy.  A woman who uses drugs (prescribed and/or non-prescribed) during pregnancy may expose her unborn child to compounds that could adversely affect neonatal development, may lead to premature delivery, and may precipitate a characteristic withdrawal syndrome called neonatal abstinence syndrome (NAS).  In the United States, the Centers for Disease Control and Prevention has reported that among 28 states surveyed, the overall incidence of NAS increased by 300% from 1999 to 2013 (https://www.cdc.gov/mmwr/volumes/65/wr/mm6531a2.htm).  A pregnant woman that has a known addiction to opioids may be managed with opioid substitution agents such as buprenorphine or methadone, which may reduce the incidence and severity of NAS.  Nonetheless, treatment of NAS may require several weeks of intensive care for the newborn, as well as extensive follow-up care and social management.  As such, accurate and timely detection of in utero substance exposure is a critical component of pre- and post-natal care and can assure provision of appropriate medical and social services for the newborn, the mother, and the associated caregivers.  Testing biological specimens for drugs is an important tool for detection and confirmation of drug exposure.  This presentation will describe pros and cons of several specimen types used to monitor exposure to medications during pregnancy and after delivery.  Common analytical methods and challenges with interpretation of results will also be discussed.

Clostridium difficile infection (CDI) has become the leading cause of healthcare associated infections in the United States and currently accounts for 15% to 20% of infectious diarrhea in the healthcare setting.  Accurate diagnosis of CDI is criticalin promoting effective management of patients and implementing appropriate infection control measures to disrupt and prevent transmission.  Reliable diagnostic tests are needed to confirm or negative a presumptive clinical diagnosis of CDI.  Nucleic acid amplification tests (NAAT) have become available within the last ten years and have been used in one of two ways: 1) Direct testing, 2) algorithmic testing as a supplement to enzyme immunoassay (EIA) for the detection of glutamate dehydrogenase and Toxin A/B.  Much controversy exists as to which approach, direct NAAT or algorimthic is optimal for the laboratory diagnosis of CDI.  The latest NAAT-based test to enter the market is the Simplexa C. diffcile Direct Assay,   An evaluation of this technology was conducted for the purpose of determining its overall performance and potential integration into two-step algorithm testing format.  The assay was evaluated and compared to two other NAAT platforms.  The results of this evaluation are the subject of this presentation.

This session will review and describe the basic steps of a biosafety risk assessment for clinical laboratories.  The risk of exposure to infectious agents exists in every clinical laboratory. Laboratories must minimize that risk and conduct activities as safely as possible. The starting point is performance of a risk assessment specific for the tests performed and instrumentation used in the laboratory. The risk assessment process relies on accumulating factual information about the pathogens and the testing procedures to define the health effects to workers from exposure.  Integral to the risk assessment process is knowledge of the biological agent, facility, safety equipment, and work processes. To perform a risk assessment requires gathering all available information on pathogens that the lab can reasonably expect to encounter and evaluating procedures used in the laboratory to determine the possible risks associated with the activities.  While the risk assessment is the initial step, it is only one component of the biosafety plan, which includes the selection of mitigation tools, employment of biosafety competencies, biosafety training, safety audits and occupational health services. 

Dr. Tony Hu’s research focuses on developing and validating integrated nanotechnique-based strategies for marker discovery and molecular diagnostics in peripheral blood samples to provide a translatable solution for personalized medicine. His innovations aim to fill current gaps in early detection, real-time therapy monitoring and effective prognostics. Instead of competing on lowering the detection limit of those known markers with poor specificity on disease diagnosis, Hu’s focus is to utilize nanotech-induced capabilities to discover those unknowns and to address major unsolved problems in oncology and infectious diseases.

The integration of next-generation sequencing (NGS) tests into clinical laboratory practice across medical disciplines provides medically actionable data that in some cases are not otherwise attainable.  This presentation will describe evolving guidance and private/public funded initiatives developed to promote the effective and safe use of NGS for patient care.  Integration of NGS into clinical laboratory practice created novel challenges and approaches for test validation, quality assurance, result interpretation, reporting, and compliance with regulatory requirements.   Data analysis, storage, and medical coding presents other challenges that require significant informatics expertise not traditionally a component of laboratory practice.  The capability to sequence the human genome, pathogens, and the human microbiome provides the opportunity to collect data having immediate and potentially future clinical relevance, blurring the line between medical and research applications.  In some instances, the laboratory assumes an enhanced consultative role to advise clinicians about test selection, variant interpretation, and the limitations of NGS; knowledge required to inform clinical decision-making.  This professional consultative role is essential toward assuring appropriate test utilization and as a resource for clinicians who are less familiar with NGS and do not have the time to establish and maintain expertise in this rapidly evolving area of laboratory medicine.

The advent of next generation sequencing has transformed the way we identify mutations with clinical implications. As the compendium of variants expands, gathering the most current and accurate information is critical to computing variant classifications. Manual curation is laborious, time consuming, and error prone, as information critical to variant interpretation may be missed. The QIAGEN knowledgebase includes the most comprehensive database of variant specific publications, as well as data from third party sources such as ClinVar, HGMD, CentoMD, and OMIM. In this webinar, you will learn how the QIAGEN knowledgebase functions as the cornerstone of our clinical decision support tool (QIAGEN Clinical Insight – Interpret; QCI-I), facilitating rapid variant filtering, interpretation, and reporting across multiple public and propriety databases.

Currently, prostate cancer is the second leading cause of death from cancer in North America, the most frequent malignancy in men from all ethnicities, surpassing lung cancer. Prostate tumors are highly heterogeneous and there are many studies evaluating the molecular profile or ‘fingerprint’ of prostate cancer.  Conventional analysis reviews the protein expression of key markers via serum analysis and immunohistochemistry (IHC) of the biopsy.  In this presentation we will review how RNA in situ hybridization (ISH) analysis of prostate cancer can unravel the molecular heterogeneity of the tumor, identify molecular subsets and detect key fusions and mutations of the RNA.  We will also present a comparison of IHC and ISH technology and the combination of dual color IHC and dual color ISH can be used to enhance simultaneous evaluation of DNA, RNA and protein, revealing the molecular profile of prostate cancer.

Despite the fact that screening for many cancers is associated with net harm, healthcare consumers continue to be screened. PSA is the most commonly performed laboratory screening test for cancer in men, but is associated with harms including complications such as erectile dysfunction, urinary incontinence and bowel dysfunction that may result from prostatectomy or radiation therapy after being diagnosed. In addition, various psychosocial morbidities may result from positive or false-positive results, and a cascade of follow-up tests and treatments. Appropriate use of laboratory screening tests has been documented to improve health outcomes.

This presentation will describe/list:

•    Major epidemiological biases that positively impact perception of screening effectiveness including lead-time and length biases
•    Current recommendations in the United States for PSA screening for prostate cancer
•    Criteria adopted by the World Health Organization (WHO) for effectiveness of any health screening program and how well the PSA test meets each.

These relate to:

o    Societal burden of the disease
o    Clinical performance of the screening test
o    Acceptability and feasibility
o    Cost-effectiveness
o    Ability for disease intervention
o    Informed consent
o    Better population health outcome with screening

The presentation will also note the consequences from screening and the dilemmas with laboratory-based screening tests such as PSA, while providing some recommendations for future use of these tests. The discussions are aimed at raising awareness about various issues surrounding prostate cancer screening using the PSA test to promote better use of laboratory screening services. 

Novel psychoactive substances (NPS) emerged in the United States around 2000.  Initially, synthetic cannabinoids and stimulants prevailed.  Now, designer opioids, benzodiazepines and others have appeared.  As a result of pain management, in addition to common drug abusers, a whole new set of individuals in desperate search for pain medications have become abusers of the NPS version of designer opioids helping spur the current opioid crisis in the United States.  The adverse effects of NPS have become significant public health and public safety concerns.  Adverse effects from these substances range from psychosis to death with abusers often committing crimes or injuring themselves.  From a forensic toxicological concern, deaths from NPS are now commonplace.  The landscape of these compounds changes frequently, making it difficult for toxicology laboratories to stay relevant.  The analytical tools required are costly and laboratories need to constantly update the scope of compounds within the assay.  The history of NPS, their associated effects and the breadth of the problem from social and scientific perspectives will be addressed along with future outlooks for this class of substances that have become a costly part of the drug use and abuse scenes in respect to lives lost and eradication.

Many direct immunoassays in use today for testosterone analysis are little better than a guess at the low levels that are sometimes required.  This has led many clinical societies to advocate for research into mass spectrometry-based testing.  Given this recommendation, and the often high volume of testing, many clinical laboratories are considering  the development of their own in-house mass spectrometry-based testosterone analytical methods.  This consideration requires selecting an instrument and workflow that fits laboratory operations and the requisite analytic specifications of the assay.  The trade-off in these considerations is typically between more advanced, and more expensive, instruments or more involved and time consuming sample preparation techniques.  This webinar will present research on different liquid chromatography tandem mass spectrometry testosterone  analytical methods that can be performed on low-, mid- and high-sensitivity mass spectrometers.  With each of these analytical methods, the increasing cost of the instrument comes with the advantage of decreasing sample preparation.  The improvements in mass spectrometers will be discussed to provide a better understanding of what makes for more sensitive instruments; the operational challenges and costs of various sample preparation strategies will also be discussed.  The goal is for clinical researchers to be aware of these tradeoffs when evaluating instruments in their research so they can select the instrument and workflow that best suits their laboratory’s needs.

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The products mentioned are for Research Use Only. Not for use in diagnostic procedures.

Pregnancy represents a time of rapid bodily change, which include physical proportions, physiology and responsibility. Arguably, nothing during these times changes more than the requirement and metabolism of vitamin D, although from current Institute of Medicine (IOM) recommendations one would never surmise this fact.  Do the current recommendations for the requirements of vitamin D during these critical time periods reflect emerging data?  Sadly, no.  The minimal recommendations by the IOM and the zero recommendation by The World Health Organization (WHO), which was echoed by a recent Cochrane Review, stating that there are simply no requirement for vitamin D during pregnancy, are contrary to expanding published data (later reviewed in this chapter) that suggest otherwise. Why these recommendations continue to be made remains unclear, but clearly not recognizing a problem exists suggests that one does not need to address the “problem.”

During these dramatic times of physiologic change, the roles of vitamin D in the pregnant versus, for example, the lactating woman are quite different.  In the pregnant women, we believe the primary role of vitamin D to be an immunomodulatory – rather than a calcium-regulating factor, although, it would also retain that function.  Further, vitamin D inadequacy in early life is clearly an instance of the “Barker Hypothesis”.  This theory states that certain adult-onset diseases might have their roots in nutritional insults sustained in the perinatal period (either in utero or in the early months of infancy or both).  Clearly, conditions associated with vitamin D deficiency such as asthma, multiple sclerosis and other neurological disorders would qualify.

If one is reading this review to relive the past 50 years of vitamin D recommendations during pregnancy, we suggest you read other recent publications for history in this area.  However, if the reader is interested in gaining new insight into the vitamin D requirements and function during pregnancy supported by recent data, we suggest you read on. Also, with regard to randomized controlled trials (RCT’s) for vitamin D, they are largely doomed to fail.  The reasons for this are many and specific cases of this failure will be presented.
 

Many Americans have succumbed to the scare tactics of the dermatological community, and avoid sunlight for fear of skin cancer and premature aging. Therefore, most of us suffer from vitamin D deficiency, which causes daily aches, pains, and fatigue, worsens serious illnesses, and diminishes the quality of everyday life. Drawing on decades of research, Dr. Michael f. Holick reveals a well-kept secret: our bodies need a sensible amount of unprotected sun exposure. 

In 2012, Consumers Reports and the American Board of Internal Medicine Foundation launched the Choosing Wisely campaign. This initiative, which attracts media attention, assists physicians, nurses, pharmacists, and medical laboratory scientists identify overused diagnostic tests and treatments. Over 70 medical specialty societies now collaborate with CR and ABIM to generate diagnostic and treatment guidelines. These include the American Association for Clinical Laboratory Science, the American Association of Blood Banks, the American Society for Hematology, and the American Society for Clinical Pathology, who focus on laboratory test utilization and the blood product administration, CR and ABIM, with support from the Robert Wood Johnson Foundation, support local "Champtions" who develop means for applying Choosing Wisely recommendations. We examine the effects of Choosing Wisely on medical laboratory test ordering and interpretation. 

Monitoring unfractionated heparin (UFH) using the activated partial thromboplastin time (APTT) or the anti-factor Xa (anti-Xa) chromogenic assay still seems to be a controversy.  Is the anti-Xa less affected by pre-analytical variables in monitoring subjects on UFH and the low molecular weight heparins (LMWH)?  Which assay is more accurate in monitoring heparin to achieve therapeutic levels, cost effective in time spent monitoring patients, use of blood products, time in the hospital, and overall cost which may include laboratory testing time.  The APTT still is the most commonly used assay to monitor UFH therapy but cannot be used to monitor the LMWHs.  Previous studies have cited interference in monitoring UFH with the  APTT from oral anticoagulants, oral contraceptives, lupus anticoagulants, factor deficiencies, elevated acute phase reactants (fibrinogen, FVIII), low anti-thrombin, the APTT reagent sensitivity and instrumentation.  The anti-Xa assay may be affected by hyperbilirubinemia, hyperlipidemia (can be cleared by ultracentrifugation) and decreased anti-thrombin levels (assay dependent). The anti-Xa assay can be used to monitor all heparin analogues and direct oral anticoagulants (DOACs).  This presentation will discuss the pros and cons of both methods and which may be most effective in patient care.