In sickness and in health: pivotal role of vitamin D

Within the last several years, frequency of vitamin D testing has multiplied substantially all over the world, since it has been shown to have an important role in many diseases and conditions. Even though liquid chromatography - tandem mass spectrometry (LC-MS/MS) has been identified as “gold standard” method for vitamin D measurement, most laboratories still use immunochemistry methods. Besides analytical problems (hydrophobicity, low circulating concentrations, ability to bind to lipids, albumins and vitamin D binding protein, presence of multiple vitamin D metabolites and variable ratios of 25(OH)D2 and 25(OH)D3 in the blood), vitamin D shows great preanalytical variability, since its concentration is drastically influenced by seasonal changes, exposure to sun, type of clothes or sun block creams. Vitamin D is mostly measured in serum or plasma, but new studies are showing importance of measuring vitamin D in pleural effusions, breast milk, urine, synovial fluid and saliva. Besides the main role in calcium homeostasis and bone metabolism, many studies linked vitamin D deficiency with cancer, cardiovascular diseases, diabetes, fertility and many other conditions. However, even though initial observational studies indicated that supplementation with vitamin D might be beneficial in disease development and progression; first results of well-designed randomized controlled prospective studies did not find differences in frequency of cardiovascular events or invasive cancer between patients taking vitamin D supplementation compared to placebo. In the light of these recent findings, validity of excessive vitamin D testing remains an open question.

Impact of an Andean breakfast on biochemistry and immunochemistry laboratory tests: an evaluation on behalf COLABIOCLI WG-PRE-LATAM

Introduction: In Andean countries, specifically in Ecuador, a food transition in the population has been observed because of economic growth. The Working Group for Preanalytical Phase in Latin America (WG-PRE-LATAM) of the Latin America Confederation of Clinical Biochemistry (COLABIOCLI) was established in 2017, and its main purpose is to study preanalytical variability and establish guidelines for preanalytical procedures in order to be implemented by clinical laboratories and healthcare professionals in Latin America. The aim of this study on behalf of COLABIOCLI WG-PRE-LATAM was to evaluate whether an Andean breakfast can interfere with routine biochemistry and immunochemistry laboratory tests. Materials and methods: We studied 20 healthy volunteers who consumed an Andean breakfast containing a standardized amount of carbohydrates, proteins and lipids. We collected blood specimens for laboratory tests before breakfast and 1, 2, and 4 hours thereafter. Significant differences between samples were assessed by the Wilcoxon ranked-pairs test. Results: The Andean breakfast statistically (P ≤ 0.05), modified the results of the following tests: triglycerides, insulin, cortisol, thyroid stimulating hormone, free thyroxine, total protein, albumin, urea, creatinine, lactate dehydrogenase, alkaline phosphatase, amylase, lipase, total bilirubin, direct bilirubin, iron, calcium, phosphorus, magnesium, and uric acid. Conclusions: Andean breakfast can influence the routine biochemistry and immunochemistry laboratory tests and might expose patient safety to some risks. Therefore, the COLABIOCLI WG-PRE-LATAM calls attention and highlights that the fasting time needs to be carefully considered when performing blood testing in order to prevent spurious results and thus, reduce laboratory errors.

The Biospecimen Preanalytical Variables Program: A Multiassay Comparison of Effects of Delay to Fixation and Fixation Duration on Nucleic Acid Quality

Context.—Despite widespread use of formalin-fixed, paraffin-embedded (FFPE) tissue in clinical and research settings, potential effects of variable tissue processing remain largely unknown.Objective.—To elucidate molecular effects associated with clinically relevant preanalytical variability, the National Cancer Institute initiated the Biospecimen Preanalytical Variables (BPV) program.Design.—The BPV program, a well-controlled series of systematic, blind and randomized studies, investigated whether a delay to fixation (DTF) or time in fixative (TIF) affects the quantity and quality of DNA and RNA isolated from FFPE colon, kidney, and ovarian tumors in comparison to case-matched snap-frozen controls.Results.—DNA and RNA yields were comparable among FFPE biospecimens subjected to different DTF and TIF time points. DNA and RNA quality metrics revealed assay- and time point–specific effects of DTF and TIF. A quantitative reverse transcription–polymerase chain reaction (qRT-PCR) assay was superior when assessing RNA quality, consistently detecting differences between FFPE and snap-frozen biospecimens and among DTF and TIF time points. RNA Integrity Number and DV200 (representing the percentage of RNA fragments longer than 200 nucleotides) displayed more limited sensitivity. Differences in DNA quality (Q-ratio) between FFPE and snap-frozen biospecimens and among DTF and TIF time points were detected with a qPCR-based assay.Conclusions.—DNA and RNA quality may be adversely affected in some tumor types by a 12-hour DTF or a TIF of 72 hours. Results presented here as well as those of additional BPV molecular analyses underway will aid in the identification of acceptable delays and optimal fixation times, and quality assays that are suitable predictors of an FFPE biospecimen's fit-for-purpose.

Effects of Fecal Sampling on Preanalytical and Analytical Phases in Quantitative Fecal Immunochemical Tests for Hemoglobin

Background Information on preanalytical variability is mandatory to bring laboratories up to ISO 15189 requirements. Fecal sampling is greatly affected by lack of harmonization in laboratory medicine. The aims of this study were to obtain information on the devices used for fecal sampling and to explore the effect of different amounts of feces on the results from the fecal immunochemical test for hemoglobin (FIT-Hb). Methods Four commercial sample collection devices for quantitative FIT-Hb measurements were investigated. The volume of interest (VOI) of the probes was measured from diameter and geometry. Quantitative measurements of the mass of feces were carried out by gravimetry. The effects of an increased amount of feces on the analytical environment were investigated measuring the Hb values with a single analytical method. Results VOI was 8.22, 7.1 and 9.44 mm3 for probes that collected a target of 10 mg of feces, and 3.08 mm3 for one probe that targeted 2 mg of feces. The ratio between recovered and target amounts of devices ranged from 56% to 121%. Different changes in the measured Hb values were observed, in adding increasing amounts of feces in commercial buffers. Conclusions The amounts of collected materials are related to the design of probes. Three out 4 manufacturers declare the same target amount using different sampling volumes and obtaining different amounts of collected materials. The introduction of a standard probes to reduce preanalytical variability could be an useful step for fecal test harmonization and to fulfill the ISO 15189 requirements.

Mobile phone exposure influences some erythrocytes parameters in vitro. A novel source of preanalytical variability?

The effect of radiofrequency exposure on human health and health care equipment is a matter of ongoing debate. This study was planned to investigate the influence of radiofrequency (RF) waves emitted by a commercial mobile phone on red blood cells (RBC) in vitro.The study population consisted of 16 ostensibly healthy volunteers. Two whole blood specimens were collected from each volunteer. One sample was placed in a plastic rack, 1 cm distant from the chassis of a commercial mobile phone which was activated by a remote phone call lasting 30 min. The other blood sample was placed in another plastic rack, but was kept distant from any type of RF source. The main RBC parameters including RBC count, hematocrit (Ht), hemoglobin, mean corpuscular platelet volume (MPV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC) and RBC distribution width (RDW-CV) were assessed with an Advia 2120.The exposure of whole blood to the mobile phone call significantly increased Ht, hemoglobin, MCV and MCH, whereas the RBC count, MCHC and RDW-CV remained unchanged. A significant correlation was observed between variation of Ht and those of hemoglobin (p=0.008), MCV (p=0.009) or MCH (p=0.037), as well as between hemoglobin and MCV (p=0.048). Increased values were found in 13/16 (81%) samples for both Ht and hemoglobin, 14/16 (88%) samples for MCH and 16/16 (100%) samples for MCV.These results suggest that close mobile phone exposure may be an unappreciated and possibly underestimated cause of preanalytical bias in RBC testing.

Preanalytical Variables Affecting the Integrity of Human Biospecimens in Biobanking

BACKGROUND Most errors in a clinical chemistry laboratory are due to preanalytical errors. Preanalytical variability of biospecimens can have significant effects on downstream analyses, and controlling such variables is therefore fundamental for the future use of biospecimens in personalized medicine for diagnostic or prognostic purposes. CONTENT The focus of this review is to examine the preanalytical variables that affect human biospecimen integrity in biobanking, with a special focus on blood, saliva, and urine. Cost efficiency is discussed in relation to these issues. SUMMARY The quality of a study will depend on the integrity of the biospecimens. Preanalytical preparations should be planned with consideration of the effect on downstream analyses. Currently such preanalytical variables are not routinely documented in the biospecimen research literature. Future studies using biobanked biospecimens should describe in detail the preanalytical handling of biospecimens and analyze and interpret the results with regard to the effects of these variables.

Harmonization in hemolysis detection and prevention. A working group of the Catalonian Health Institute (ICS) experience

Hemolysis is the main cause of non-quality samples in clinical laboratories, producing the highest percentage of rejections in the external assurance programs of preanalytical quality. The objective was to: 1) study the agreement between the detection methods and quantification of hemolysis; 2) establish comparable hemolysis interference limits for a series of tests and analytical methods; and 3) study the preanalytical variables which most influence hemolysis production.Different hemoglobin concentration standards were prepared using the reference method. Agreement was studied between automated methods [hemolytic indexes (HI)] and reference method, as well as the interference according to hemolysis degree in various biochemical tests was measured. Preanalytical variables which could influence hemolysis production were studied: type of extraction, type of tubes, transport time, temperature and centrifugation conditions.Good agreement was obtained between hemoglobin concentrations measured using the reference method and HI, for the most of studied analyzers, particularly those giving quantitative HI. The hemolysis interference cut-off points obtained for the majority of tests studied (except LDH, K) are dependent on the method/analyzer utilized. Furthermore, discrepancies have been observed between interference limits recommended by the manufacturer. The preanalytical variables which produce a lower percentage of hemolysis rejections were: centrifugation at the extraction site, the use of lower volume tubes and a transport time under 15 min at room temperature.The setting of interference limits (cut-off) for each used test/method, and the study of preanalytical variability will assist to the results harmonization for this quality indicator.