Characterizing ability of serum potassium (K) and the serum K reference interval to flag hypokalemia or hyperkalemia as observed in plasma: A simulation study.

OBJECTIVES: Serum potassium (K) exhibits a positive shift relative to plasma K due to a variable amount of K release associated with clotting. Because of this variation, plasma K results outside of the reference interval (RI) for plasma (hypokalemia or hyperkalemia) in individual samples may not produce classification-concordant results in serum according to the serum RI. We examined this premise from a theoretical standpoint by simulation. DESIGN & METHODS: We used textbook K reference intervals for plasma (PRI = 3.4-4.5 mmol/L) and serum (SRI = 3.5-5.1 mmol/L). The difference between PRI and SRI is characterized by a normal distribution: serum K = plasma K + 0.35 ± 0.308 mmol/L. This transformation was applied by simulation to an observed patient data distribution for plasma K to generate a corresponding theoretical serum K distribution. Individual samples were tracked for comparison with respect to classification (below, within, above RI) for plasma and serum. RESULTS: Primary data were an all-comers plasma K patient distribution (n = 41,768; median = 4.1 mmol/L; 7.1% below PRI (hypokalemia); 15.5% above PRI (hyperkalemia)). Simulation to obtain the associated serum K yielded a right-shifted distribution (median = 4.4 mmol/L; 4.8% below SRI; 10.8% above SRI). Sensitivity for detection in serum (flagged below SRI) for samples originating as hypokalemic in plasma was 45.7% (specificity = 98.3%). Sensitivity for detection in serum (flagged above SRI) for samples originating as hyperkalemic in plasma was 56.6% (specificity = 97.6%). CONCLUSIONS: Simulation results indicate that serum K should best be thought of as an inferior substitute marker for plasma K. These results follow simply from the variable component of serum K compared to plasma K. Plasma should be the preferred specimen type for K assessment.

Does Diabetes Alter CSF Total Protein Levels? A Retrospective Cohort Study.

Background and Purpose: Elevation of total protein level in cerebrospinal fluid (CSF-TP) in diabetic patients is often disregarded by clinicians. However, existing studies on the topic have significant limitations, and therefore we aimed to explore the relationship between diabetes and CSF-TP in a large database of CSF samples. Methods: Retrospective review of all diagnostic lumbar punctures at the Ottawa Hospital between 1996-2016. Patients were excluded if they had elevated CSF cell counts, or a condition known to elevate CSF-TP. Multivariate linear regression modeling considered the effects of age, sex, and diabetes. Results: Among 6124 patients (746 with diabetes, 5378 without), mean CSF-TP did not differ significantly between groups (0.39 and 0.35 mmol/L, p = 0.2). When controlled for age and sex, there was no significant effect of diabetes on CSF-TP and no significant correlation between mean serum glucose and CSF-TP (R2 = 0.12). Conclusions: CSF-TP did not differ significantly between diabetic and non-diabetic groups, once the influence of age and sex was controlled. Elevated CSF-TP should be regarded as pathologic, even in the setting of diabetes.

Limited Evidence for Use of a Black Race Modifier in eGFR Calculations: A Systematic Review.

BACKGROUND: Commonly used estimated glomerular filtration rate (eGFR) equations include a Black race modifier (BRM) that was incorporated during equation derivation. Race is a social construct, and a poorly characterized variable that is applied inconsistently in clinical settings. The BRM results in higher eGFR for any creatinine concentration, implying fundamental differences in creatinine production or excretion in Black individuals compared to other populations. Equations without inclusion of the BRM have the potential to detect kidney disease earlier in patients at the greatest risk of chronic kidney disease (CKD), but also has the potential to over-diagnose CKD or impact downstream clinical interventions. The purpose of this study was to use an evidence-based approach to systematically evaluate the literature relevant to the performance of the eGFR equations with and without the BRM and to examine the clinical impact of the use or removal. CONTENT: PubMed and Embase databases were searched for studies comparing measured GFR to eGFR in racially diverse adult populations using the Modification of Diet in Renal Disease or the 2009-Chronic Kidney Disease Epidemiology Collaboration-creatinine equations based on standardized creatinine measurements. Additionally, we searched for studies comparing clinical use of eGFR calculated with and without the BRM. Here, 8632 unique publications were identified; an additional 3 studies were added post hoc. In total, 96 studies were subjected to further analysis and 44 studies were used to make a final assessment. SUMMARY: There is limited published evidence to support the use of a BRM in eGFR equations.

Reproducible manuscript preparation with RMarkdown application to JMSACL and other Elsevier Journals.

INTRODUCTION: With the rising complexity of modern multimarker analytical techniques and notable scientific publication retractions required for erroneous statistical analysis, there is increasing awareness of the importance of research transparency and reproducibility. The development of mature open-source tools for literate programming in multiple langauge paradigms has made fully-reproducible authorship possible. OBJECTIVES: We describe the procedure for manuscript preparation using RMarkdown and the R statistical programming language with application to JMSACL or any other Elsevier journal. METHODS: An instructional manuscript has been prepared in the RMarkdown markup language with stepwise directions on preparing sections, subsections, lists, tables, figures and reference management in an entirely reproducible format. RESULTS: From RMarkdown code, a submission-ready PDF is generated and JMSACL-compatible LaTeX code is generated. These can be uploaded to the Editorial Manager. CONCLUSION: A completely reproducible manuscript preparation pipeline using the R and RMarkdown is described.

Continuous reference intervals for pediatric testosterone, sex hormone binding globulin and free testosterone using quantile regression.

Testosterone (T), sex hormone binding globulin (SHBG), free testosterone (FT) and bioavailable testosterone (BAT) are commonly employed tests in pediatric endocrinology and all require age-dependent reference intervals for interpretation. The common methods used to derive these reference intervals require decisions about data shape and/or age partition thresholds, which can result in sharp differences between age groups, particularly for pubescent children. Partitioning also results in a form of data loss, where data from one age-bin is completed disconnected from the adjacent age-bins. Non-parametric continuous reference intervals methods have previously been developed to avoid some of these drawbacks. These strategies use all the available data and smooth transitions between ages avoiding partitioning. However, the fitting process involves selection and adjustment of many parameters and it can be difficult to maintain a reproducible approach. Here we provide a workflow for non-parametric continuous reference intervals applied to T, FT, BAT, and SHBG using the R language quantregGrowth package. T measurements were determined by LC-MS/MS, FT and BAT were calculated, and SHBG was measured on the Roche Cobas e601. The continuous interval methodology is described in detail with code examples and illustrations for reproducibility.

Effective interventions to improve the quality of critically high point-of-care glucose meter results.

OBJECTIVES: Point-of-care testing (POCT) is testing performed outside the traditional laboratory, often at the patient bedside. In hospital settings, blood glucose is the most common POCT. Staff performing POCT are not usually laboratory trained; they are clinical staff with a primary focus on treating patients. Clinical staff find POCT quality assurance (QA) practices burdensome and are often non-compliant. In hospitals within EORLA (Eastern Ontario Regional Laboratories Association), all critically high POCT glucose results must be repeated prior to acting, according to policy. Compliance with this policy is audited regularly. DESIGN: and methods: All POCT glucose tests performed in participating sites between January and June 2018 and June and December 2019 were audited for compliance with the critical repeat policy. The discordant repeat rate was also determined for each audit period. Between January and May 2019, there were interventions aimed at improving compliance with the repeat policy. RESULTS: Compliance with the critical repeat policy increased from 30 to 57% in 2019 compared to 2018, following nursing education and implementation of notifications on the glucose meters themselves. The rate of discordant repeat results (>20% different from initial) also improved at most sites in 2019 compared to 2018. Nurses cited insufficient cleaning of patient hands prior to initial testing as the primary reason for discordant repeats. CONCLUSIONS: Operator compliance with POCT QA policies is an ongoing challenge requiring continual audit, feedback and education. A strong POCT multi-disciplinary committee with supports from senior and clinical leadership in an organization are key to improving compliance.

An international multi-center serum protein electrophoresis accuracy and M-protein isotyping study. Part II: limit of detection and follow-up of patients with small M-proteins.

Background Electrophoretic methods to detect, characterize and quantify M-proteins play an important role in the management of patients with monoclonal gammopathies (MGs). Significant uncertainty in the quantification and limit of detection (LOD) is documented when M-proteins are <10 g/L. Using spiked sera, we aimed to assess the variability in intact M-protein quantification and LOD across 16 laboratories. Methods Sera with normal, hypo- or hyper-gammaglobulinemia were spiked with daratumumab or elotuzumab, with concentrations from 0.125 to 10 g/L (n = 62) along with a beta-migrating sample (n = 9). Laboratories blindly analyzed samples according to their serum protein electrophoresis (SPEP)/isotyping standard operating procedures. LOD and intra-laboratory percent coefficient of variation (%CV) were calculated and further specified with regard to the method (gel/capillary electrophoresis [CZE]), gating strategy (perpendicular drop [PD]/tangent skimming [TS]), isotyping (immunofixation/immunosubtraction [ISUB]) and manufacturer (Helena/Sebia). Results All M-proteins ≥1 g/L were detected by SPEP. With isotyping the LOD was moderately more sensitive than with SPEP. The intensity of polyclonal background had the biggest negative impact on LOD. Independent of the method used, the intra-laboratory imprecision of M-protein quantification was small (mean CV = 5.0%). Low M-protein concentration and high polyclonal background had the strongest negative impact on intra-laboratory precision. All laboratories were able to follow trend of M-protein concentrations down to 1 g/L. Conclusions In this study, we describe a large variation in the reported LOD for both SPEP and isotyping; overall LOD is most affected by the polyclonal immunoglobulin background. Satisfactory intra-laboratory precision was demonstrated. This indicates that the quantification of small M-proteins to monitor patients over time is appropriate, when subsequent testing is performed within the same laboratory.

An international multi-center serum protein electrophoresis accuracy and M-protein isotyping study. Part I: factors impacting limit of quantitation of serum protein electrophoresis.

Background Serum protein electrophoresis (SPEP) is used to quantify the serum monoclonal component or M-protein, for diagnosis and monitoring of monoclonal gammopathies. Significant imprecision and inaccuracy pose challenges in reporting small M-proteins. Using therapeutic monoclonal antibody-spiked sera and a pooled beta-migrating M-protein, we aimed to assess SPEP limitations and variability across 16 laboratories in three continents. Methods Sera with normal, hypo- or hypergammaglobulinemia were spiked with daratumumab, Dara (cathodal migrating), or elotuzumab, Elo (central-gamma migrating), with concentrations from 0.125 to 10 g/L (n = 62) along with a beta-migrating sample (n = 9). Provided with total protein (reverse biuret, Siemens), laboratories blindly analyzed samples according to their SPEP and immunofixation (IFE) or immunosubtraction (ISUB) standard operating procedures. Sixteen laboratories reported the perpendicular drop (PD) method of gating the M-protein, while 10 used tangent skimming (TS). A mean percent recovery range of 80%-120% was set as acceptable. The inter-laboratory %CV was calculated. Results Gamma globulin background, migration pattern and concentration all affect the precision and accuracy of quantifying M-proteins by SPEP. As the background increases, imprecision increases and accuracy decreases leading to overestimation of M-protein quantitation especially evident in hypergamma samples, and more prominent with PD. Cathodal migrating M-proteins were associated with less imprecision and higher accuracy compared to central-gamma migrating M-proteins, which is attributed to the increased gamma background contribution in M-proteins migrating in the middle of the gamma fraction. There is greater imprecision and loss of accuracy at lower M-protein concentrations. Conclusions This study suggests that quantifying exceedingly low concentrations of M-proteins, although possible, may not yield adequate accuracy and precision between laboratories.

Age matters: Impact of data-driven CSF protein upper reference limits in Guillain-Barré syndrome.

Objective: We conducted a retrospective review of patients with a diagnosis of Guillain-Barré syndrome (GBS) to assess the diagnostic impact of applying age-adjusted upper limits for CSF total protein (CSF-TP) supported by a systematic literature review. Methods: Cases coded as GBS or inflammatory neuropathy for the period 2001-2016 at The Ottawa Hospital were reviewed. Cases were included if they met the Brighton criteria for GBS with a diagnostic certainty level 1 or 2 and had contemporaneous CSF-TP data. We excluded cases with CSF pleocytosis >50 and cases with Miller-Fisher syndrome. Age-adjusted reference limits were compared with conventional 0.45 and 0.6 g/L upper limits. Results: One hundred thirty-eight cases met the study criteria, with a mean age of 47 years. The mean interval from symptom onset to lumbar puncture was 7.9 days, and mean CSF-TP was 1.23 g/L. There was a strong correlation between rising CSF-TP and time to lumbar puncture. Age-adjusted CSF-TP had a significantly lower sensitivity of only 45% in the first week (32% in the first 3 days) compared with 70% in the first week for the 0.45 g/L limit. All upper limits gained high sensitivity after the first week. Conclusions: The low sensitivity of CSF-TP for the diagnosis of GBS is exacerbated by age-adjusted upper limits. The main role of lumbar puncture in GBS in the first week may be to help exclude other inflammatory or neoplastic etiologies of acute neuropathy. After the first week, the magnitude of the CSF-TP rise reduces the effect of different upper reference limits.

A Survey of Cerebrospinal Fluid Total Protein Upper Limits in Canada: Time for an Update?

BACKGROUND: The antiquated standard reference range of 0.15-0.45 g/L for cerebrospinal fluid total protein (CSF-TP) is well entrenched in medical literature and laboratory operating procedures across the world. METHODS: We conducted a web-based survey with a response rate of 34.9% through the listserv of the Canadian Neurological Sciences Federation. Additional laboratory reference data were collated by telephone interview of hospital laboratory technologists across Canada. RESULTS: A total of 142 site responses were obtained: 64.1% from academic/tertiary hospitals and 35.9% from community hospitals. A strong majority (80.4%) of both types of institutions reported using a CSF-TP upper reference limit of 0.45 g/L or less. As a rule, no age adjustments were implemented in CSF-TP-level interpretation. CONCLUSIONS: Recent well-powered laboratory reference studies have documented CSF-TP upper reference limits that are above 0.6 g/L starting at age 50, with incremental limits partitioned by subsequent decades of age. The conventional 0.45 g/L limit could lead to false positive results. Our survey suggests there is a need to consider a wide adoption of data-driven, rather than historical, reference values.

Adult CSF total protein upper reference limits should be age-partitioned and significantly higher than 0.45 g/L: a systematic review.

BACKGROUND: Cerebrospinal fluid total protein (CSF-TP) is measured in the diagnosis of a range of immune or infectious disorders of the nervous system. Most laboratories and the medical literature use an antiquated, age-independent upper limit of 0.45 g/L. Therefore, we performed a systematic review of reference studies in the medical literature, with the primary objective of determining the CSF total protein upper reference limit (URL). Secondary objectives were to assess the effects of age, gender, laboratory methods, and methodological quality. METHODS: A pre-planned and peer-reviewed electronic search strategy was used to search Ovid Medline and EMBASE for 1960-2017. All records underwent title/abstract review, and potentially relevant records underwent independent full-text review by two researchers. The remaining studies underwent quality assessment using a modification of the QUADAS2 revised tool. CSF-TP upper reference limits extracted from these studies were used to compute weighted means. RESULTS: Twenty-two articles were retained for qualitative analysis and 20 for quantitative analysis. The weighted average of CSF-TP URL was 0.55 g/L, in studies with high methodological quality. Studies that examined the effect of age reported consistent correlations with advancing age, and CSF-TP URL values incrementally exceeded 0.60 g/L after age 50. There were no meaningful differences according to gender, laboratory method, or quality assessment score. CONCLUSIONS: There is concordance in available literature to recommend increasing CSF total protein upper reference limits, and to consider implementing age-adjusted values above 0.60 g/L starting at age 50. This information merits worldwide dissemination, to reduce the risk of over-diagnosis.

Adult CSF total protein: Higher upper reference limits should be considered worldwide. A web-based survey.

BACKGROUND: The cerebrospinal fluid total protein level (CSF-TP) is commonly used as a potential marker of infectious or immune disease of the CNS and PNS. Recent laboratory reference studies indicate that the antiquated single upper reference limit of 0.45 g/L commonly used by hospital laboratories and widely quoted in medical literature is a significant underestimation. METHODS: We distributed worldwide a web-based survey comprised of three questions: 1. What is the CSF-TP upper limit used at your institution? 2. What is the source of this upper limit? 3. Do you adjust your upper limit according to age? RESULTS: A total of 473 unique responses were obtained from North America (37.5%), South America (5.5%), Europe (29.4%), Africa (4%), Asia (21.6%) and Oceania (1.7%). A strong preponderance (86.8%) of institutions reported an upper limit of 0.45 g/L or less. Only 4% reported making age-partitioned adjustments. CONCLUSIONS: Worldwide, a strong majority of hospital laboratories presently use an underestimation of CSF-TP upper reference value, particularly for older adults. Recent well powered laboratory reference studies support higher values with age adjustment.

Quality, origins and limitations of common therapeutic drug reference intervals.

Therapeutic drug monitoring (TDM) is used to manage drugs with a narrow window between effective and toxic concentrations. TDM involves measuring blood concentrations of drugs to ensure effective therapy, avoid toxicity and monitor compliance. Common drugs for which TDM is used include aminoglycosides for infections, anticonvulsants to treat seizures, immunosuppressants for transplant patients and cardiac glycosides to regulate cardiac output and heart rate. An essential element of TDM is the provision of accurate and clinically relevant reference intervals. Unlike most laboratory reference intervals, which are derived from a healthy population, TDM reference intervals need to relate to clinical outcomes in the form of efficacy and toxicity. This makes TDM inherently more difficult to develop as healthy individuals are not on therapy, so there is no "normal value". In addition, many of the aforementioned drugs are old and much of the information regarding reference intervals is based on small trials using methods that have changed. Furthermore, individuals have different pharmacokinetics and drug responses, particularly in the context of combined therapies, which exacerbates the challenge of universal TDM targets. This focused review examines the origins and limitations of existing TDM reference intervals for common drugs, providing targets where possible based on available guidelines.

Recognition and management of common, rare, and novel serum protein electrophoresis and immunofixation interferences.

Protein electrophoresis and immunofixation are subject to a variety of analytical interferences that may affect monoclonal protein diagnostics performed in the context of monoclonal gammopathies. Interferences include endogenous substances, such as hemoglobin and fibrinogen, and exogenous compounds, such as radiocontrast dyes, antibiotics, and monoclonal antibody therapies. General approaches to managing interferences begin with recognition of the problem. Provided herein are examples of common, rare, and novel interferences with the goal of providing a comprehensive overview. With each example, specific methods and strategies are provided to manage analytical interferences to ensure that interpretative reports are accurate. Longstanding and newer technologies are also described to contextualize where interferences may be identified and avoided.