Comparison of quantitative and qualitative anti-dsDNA assays.

OBJECTIVE: In evaluation of systemic lupus erythematosus (SLE), anti-double-stranded DNA antibodies (anti-dsDNA) play a significant role in diagnosis, monitoring SLE activity, and assessing prognosis. However, evaluations of the performance and limitations for recently developed methods for anti-dsDNA assessment are sparse. METHODS: Specimens used for antinuclear antibody testing (n = 129) were evaluated for anti-dsDNA assay comparability across 4 medical centers in the United States. The methods compared were Werfen Quanta Lite dsDNA, Zeus Scientific dsDNA Enzyme Immunoassay, Bio-Rad multiplex immunoassay (MIA) dsDNA, ImmunoConcepts Crithidia, and Bio-Rad Laboratories Crithidia. RESULTS: For quantitative anti-dsDNA measurements, Spearman's correlation coefficient was highest between Zeus and Werfen (ρ = 0.86; CI, 0.81-0.90; P < .0001). Comparison of MIA to Werfen or Zeus yielded similar results to each other (ρ = 0.58; CI, 0.44-0.68; P < .0001; and ρ = 0.59; CI, 0.46-0.69; P < .0001, respectively), but lower than the correlation between Zeus and Werfen. Positive concordance between assays ranged from 31.4% to 97.1%, and negative concordance between assays ranged from 58.5% to 100%. The detection of anti-dsDNA in those with SLE diagnosis ranged from 50.9% to 77.4% for quantitative assays and 15.1% to 24.5% for Crithidia assays. CONCLUSION: Current quantitative anti-dsDNA assays are not interchangeable for patient follow-up. Crithidia-based assays demonstrate high negative concordance and lack positive concordance among the methods.

Trends in co-prescribed opioids and benzodiazepines, non-prescribed opioids and benzodiazepines, and schedule-I drugs in the United States, 2013 to 2019.

Concurrent opioid and benzodiazepine users are at increased risk of overdose death, compared to opioid-only users. The objective of this study was to understand recent time trends in opioid and benzodiazepine concurrent use, misuse, and schedule-I drug use, and how these differ by age, sex and geographic region. Commercial, United States medical insurance claims data and urine drug test results from 2013 to 2019 were used to study the outcomes of concurrent use (n = 756,258), schedule-I drug use (n = 746,672) and prescription misuse (n = 452,523). Drug use outcomes were studied at quarterly time points for each year. Data analysis included joinpoint regression models to estimate quarterly drug use rates, determined by positive urine tests for corresponding drug categories, and was conducted from November 2021 through January 2022. Concurrent use decreased from 19.3% to 9.8%, misuse generally decreased from 75.6% to 55.1%, and schedule-I use increased from 8.9% to 13.8%, from 2013 to 2019. Concurrent use decreased at greater rates after 2016, after the Centers for Disease Control and Food and Drug Administration guidelines against concurrent use were released, while schedule-I use increased, notably after the 2014 hydrocodone reschedule. This indicates a potential shift from prescription use to non-prescribed drug use, where most affected groups included males, younger individuals, and those residing in Northeastern regions. Study results support public health initiatives focused on policy that increases access to multimodal pain management and substance use disorder management programs-critical steps in preventing patients from seeking non-prescribed drugs for self- medicating due to pain or addiction.

Retrospective comparison of false-positive result frequencies of 3 syphilis serology screening tests in pregnant and nonpregnant patients at an academic medical center in Appalachia.

OBJECTIVE: This study retrospectively compared false-positive result frequencies of 3 syphilis serology screening tests and assessed whether false positivity was associated with pregnancy and age. METHODS: Results for 3 screening tests were retrieved from the laboratory database, including rapid plasma reagin (RPR) assay between October 2016 and September 2019, BioPlex 2200 Syphilis Total immunoassay between May 2020 and January 2022, and Alinity i Syphilis TP assay between February 2022 and April 2023. The false-positive result frequencies were calculated based on testing algorithm criteria. RESULTS: False-positive result frequency for BioPlex was 0.61% (90/14,707), significantly higher than 0.29% (50/17,447) for RPR and 0.38% (55/14,631) for Alinity (both P < .01). Patients with false-positive results were significantly older than patients with nonreactive results for RPR (median age: 36 vs 28, P < .001), but not for BioPlex or Alinity. For all 3 tests, the positive predictive values in pregnant women were lower than those in nonpregnant women or men. However, pregnant women did not exhibit a higher false-positive result frequency. CONCLUSION: Although false-positive result frequencies were low overall for all 3 syphilis serology tests, there is a significant difference between different tests. Pregnancy was not associated with more false-positive results for all 3 tests.

The North American opioid epidemic: opportunities and challenges for clinical laboratories.

Since 1999, the opioid epidemic in North America has resulted in over 1 million deaths, and it continues to escalate despite numerous efforts in various arenas to combat the upward trend. Clinical laboratories provide drug testing to support practices such as emergency medicine, substance use disorder treatment, and pain management; increasingly, these laboratories are collaborating in novel partnerships including drug-checking services (DCS) and multidisciplinary treatment teams. This review examines drug testing related to management of licit and illicit opioid use, new technologies and test strategies employed by clinical laboratories, barriers hindering laboratory response to the opioid epidemic, and areas for improvement and standardization within drug testing. Literature search terms included combinations of "opioid," "opiate," "fentanyl," "laboratory," "epidemic," "crisis," "mass spectrometry," "immunoassay," "drug screen," "drug test," "guidelines," plus review of PubMed "similar articles" and references within publications. While immunoassay (IA) and point-of-care (POC) test options for synthetic opioids are increasingly available, mass spectrometry (MS) platforms offer the greatest flexibility and sensitivity for detecting novel, potent opioids. Previously reserved as a second-tier application in most drug test algorithms, MS assays are gaining a larger role in initial screening for specific patients and DCS. However, there are substantial differences among laboratories in terms of updating test menus, algorithms, and technologies to meet changing clinical needs. While some clinical laboratories lack the resources and expertise to implement MS, many are also slow to adopt available IA and POC tests for newer opioids such as fentanyl. MS-based testing also presents challenges, including gaps in available guidance for assay validation and ongoing performance assessment that contribute to a dramatic lack of standardization among laboratories. We identify opportunities for improvement in laboratory operations, reporting, and interpretation of drug test results, including laboratorian and provider education and laboratory-focused guidelines. We also highlight the need for collaboration with providers, assay and instrument manufacturers, and national organizations to increase the effectiveness of clinical laboratory and provider efforts in preventing morbidity and mortality associated with opioid use and misuse.

Interplay of Antibody and Cytokine Production Reveals CXCL13 as a Potential Novel Biomarker of Lethal SARS-CoV-2 Infection.

The SARS-CoV-2 pandemic is impacting the global population. This study was designed to assess the interplay of antibodies with the cytokine response in SARS-CoV-2 patients. We demonstrate that significant levels of anti-SARS-CoV-2 antibody to receptor binding domain (RBD), nucleocapsid, and spike S1 subunit of SARS-CoV-2 develop over the first 10 to 20 days of infection. The majority of patients produced antibodies against all three antigens (219/255 SARS-CoV-2+ patient specimens, 86%), suggesting a broad response to viral proteins. Antibody levels to SARS-CoV-2 antigens were different based on patient mortality, sex, blood type, and age. Analyses of these findings may help explain variation in immunity between these populations. To better understand the systemic immune response, we analyzed the levels of 20 cytokines by SARS-CoV-2 patients throughout infection. Cytokine analysis of SARS-CoV-2+ patients exhibited increases in proinflammatory markers (interleukin 6 [IL-6], IL-8, IL-18, and gamma interferon [IFN-γ]) and chemotactic markers (IP-10 and eotaxin) relative to healthy individuals. Patients who succumbed to infection produced decreased IL-2, IL-4, IL-12, RANTES, tumor necrosis factor alpha (TNF-α), GRO-α, and MIP-1α relative to patients who survived infection. We also observed that the chemokine CXCL13 was particularly elevated in patients who succumbed to infection. CXCL13 is involved in B cell activation, germinal center development, and antibody maturation, and we observed that CXCL13 levels in blood trended with anti-SARS-CoV-2 antibody levels. Furthermore, patients who succumbed to infection produced high CXCL13 and had a higher ratio of nucleocapsid to RBD antibodies. This study provides insights into SARS-CoV-2 immunity implicating the magnitude and specificity of response in relation to patient outcomes.IMPORTANCE The SARS-CoV-2 pandemic is continuing to impact the global population, and knowledge of the immune response to COVID-19 is still developing. This study assesses the interplay of different parts of the immune system during COVID-19 disease. We demonstrate that COVID-19 patients produce antibodies to three proteins of the COVID-19 virus (SARS-CoV-2) and identify many other immunological proteins that are involved during infection. The data suggest that one of these proteins (CXCL13) may be a novel biomarker for severe COVID-19 that can be readily measured in blood. This information combined with our broad-scale analysis of immune activity during COVID-19 provides new information on the immunological response throughout the course of disease and identifies a novel potential marker for assessing disease severity.

Interlaboratory Agreement of Anti-Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Serologic Assays in the Expedited College of American Pathologists Proficiency Testing Program.

CONTEXT.­: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a recently emerged, currently pandemic virus, and the etiologic agent of coronavirus disease 2019 (COVID-19). Clinical testing for antibodies to SARS-CoV-2 has rapidly become widespread, but data regarding the interlaboratory performance of these serologic assays are limited. OBJECTIVE.­: To describe the development and initial results of the College of American Pathologists (CAP) SARS-CoV-2 Serology Survey. DESIGN.­: Members from the CAP Microbiology and Diagnostic Immunology and Flow Cytometry Committees formed a working group to support development of a new proficiency testing survey for anti-SARS-CoV-2 antibody assays. Supplemental questions in the survey assessed the state of SARS-CoV-2 serologic testing among participating laboratories as of July 2020. Results were analyzed for agreement by immunoglobulin (Ig) isotype tested, assay manufacturer, and methodology. RESULTS.­: A total of 4125 qualitative results were received from 1110 laboratories participating in the first survey. Qualitative agreement for assays measuring anti-SARS-CoV-2 total antibodies or IgG was greater than 90% for all 3 samples in the survey. Qualitative agreement for IgM and IgA for the negative sample was greater than 95%, but lacked consensus for the other 2 samples. CONCLUSIONS.­: These initial data suggest overall excellent agreement and comparable performance for most qualitative anti-SARS-CoV-2 IgG and total antibody assays across all participating clinical laboratories, regardless of specific target antigen or assay methodology.

Drug Positivity Findings from a Universal Umbilical Cord Tissue Drug Analysis Program in Appalachia.

BACKGROUND: West Virginia has high rates of opioid-related health crises and deaths that extend to pregnant women and newborns. Our institutional screening approach has included universal umbilical cord tissue drug analysis (UCTDA) since 2013. The objective of this study was to retrospectively report incidence of in utero drug exposure using UCTDA data. METHODS: Two sequential UCTDA data sets (October 2013 to September 2015, and October 2016 to September 2018) represent interrupted epochs given changes in interfaced data availability. UCTDA positivity (by drug class and parent drug) and numbers of drugs detected in each specimen were retrospectively analyzed. THC was removed from the analysis because of discontinuous testing, and 4 opioids were separated from the data set given the potential for both therapeutic and illicit use. RESULTS: UCTDA specimens that were positive for drugs (22% overall) decreased between Epochs 1 and 2, from 25% to 20%. Increased positivity was noted for hydrocodone (+407%), oxycodone (+240%), amphetamines (+506%), and cocaine (+417%). Fentanyl and morphine positivity decreased by 75% and 18%, respectively, whereas buprenorphine detection increased 195%. Most positive specimens (80% overall) had 1 drug present, but specimens positive for 2 to 6 discrete drugs were found. CONCLUSION: Universal UCTDA allows for unbiased assessment of drug exposure in infants. With the additional knowledge of therapeutic indications for drug use, UCTDA may allow for analysis of trends in illicit drug use and the impact of interventions to curb neonatal abstinence syndrome.

Retrospective evaluation of 99th percentile hCG results to adjust clinical decision points.

While accurate measurement of chorionic gonadotropin (hCG) is necessary, so are appropriate clinical decision points (CDPs) for patients of all ages. The CDP for hCG is intended to identify early pregnancy in patients of child bearing age; non-pregnant patients who are older frequently yield hCG results > 5 IU/L, making the use of a low hCG CDP problematic for these patients. Using a retrospective review of all hCG results generated over a 32-month period, 8507 hCG results from non-pregnant females of all ages were analyzed. Patients < 40 years of age comprised 74% of hCG measurements, and produced hCG results ≥ 5 IU/L 1% of the time, but this frequency increased in patients 40-49 (17% of hCG results; 4% ≥5 IU/L) and ≥50 (9% of hCG results; 20% ≥ 5 IU/L). While only 3% of hCG results were ≥5 IU/L in the overall data set, all (24/24) of hCG results 10-14 IU/L came from patients ≥ 40 years of age and all (3/3) hCG results ≥ 15 IU/L came from patients ≥ 50 years of age. The 99th percentile hCG results in the population were 3 IU/L in patients < 40, 7 IU/L in patients 40-49, and 13 IU/L in patients ≥ 50 years of age. These findings demonstrate a progressive increase in measurable hCG correlating to patient age and demonstrate a proof-of-concept that institutions could assess 99th percentile hCG results to assign more appropriate method-dependent CDPs to different age groups.

The Effect of Proprotein Convertase Subtilisin/Kexin Type 9 Inhibitors on Nonfasting Remnant Cholesterol in a Real World Population.

BACKGROUND: Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors have demonstrated significant effects on low-density lipoprotein (LDL) cholesterol and nonhigh density lipoprotein (HDL) cholesterol. To date, there have been limited reports on the effect of PCSK9 inhibitors on remnant cholesterol. OBJECTIVES: Assess the effect of PCSK9 inhibitors on nonfasting remnant cholesterol in a real world population. Identify whether pretreatment triglyceride levels are associated with PCSK9 inhibition success as indicated by changes in remnant cholesterol levels. METHODS: Patients in our adult lipid clinic (n = 109) receiving PCSK9 inhibition for atherosclerotic cardiovascular disease or familial hypercholesterolemia who had available pre- and post-PCSK9 inhibition standard nonfasting lipid data were, retrospectively, selected for data analysis. Remnant cholesterol was the difference between non-HDL and LDL cholesterol. LDL cholesterol was measured directly and calculated from Friedewald and Martin/Hopkins methods. Data were analyzed using repeated measures ANOVA and multivariable linear regression for differential effects on remnant and LDL cholesterol based upon pretreatment nonfasting triglyceride levels. RESULTS: Remnant cholesterol as well as total, LDL, non-HDL cholesterol, and triglycerides decreased significantly (P<0.001) after PCSK9 inhibition. Patients with higher pretreatment triglyceride levels showed greater decrease in remnant cholesterol after PCSK9 inhibition (P<0.001) than those with lower pretreatment triglycerides. CONCLUSIONS: In patients receiving PCSK9 inhibitors, remnant cholesterol as determined from nonfasting blood was reduced in proportion to pretreatment triglycerides.

Quality Improvement Intervention for Reduction of Redundant Testing.

Laboratory data are critical to analyzing and improving clinical quality. In the setting of residual use of creatine kinase M and B isoenzyme testing for myocardial infarction, we assessed disease outcomes of discordant creatine kinase M and B isoenzyme +/troponin I (-) test pairs in order to address anticipated clinician concerns about potential loss of case-finding sensitivity following proposed discontinuation of routine creatine kinase and creatine kinase M and B isoenzyme testing. Time-sequenced interventions were introduced. The main outcome was the percentage of cardiac marker studies performed within guidelines. Nonguideline orders dominated at baseline. Creatine kinase M and B isoenzyme testing in 7496 order sets failed to detect additional myocardial infarctions but was associated with 42 potentially preventable admissions/quarter. Interruptive computerized soft stops improved guideline compliance from 32.3% to 58% (P < .001) in services not receiving peer leader intervention and to >80% (P < .001) with peer leadership that featured dashboard feedback about test order performance. This successful experience was recapitulated in interrupted time series within 2 additional services within facility 1 and then in 2 external hospitals (including a critical access facility). Improvements have been sustained postintervention. Laboratory cost savings at the academic facility were estimated to be ≥US$635 000 per year. National collaborative data indicated that facility 1 improved its order patterns from fourth to first quartile compared to peer norms and imply that nonguideline orders persist elsewhere. This example illustrates how pathologists can provide leadership in assisting clinicians in changing laboratory ordering practices. We found that clinicians respond to local laboratory data about their own test performance and that evidence suggesting harm is more compelling to clinicians than evidence of cost savings. Our experience indicates that interventions done at an academic facility can be readily instituted by private practitioners at external facilities. The intervention data also supplement existing literature that electronic order interruptions are more successful when combined with modalities that rely on peer education combined with dashboard feedback about laboratory order performance. The findings may have implications for the role of the pathology laboratory in the ongoing pivot from quantity-based to value-based health care.

Quality Monitoring Approach for Optimizing Antinuclear Antibody Screening Cutoffs and Testing Work Flow.

BACKGROUND: An antinuclear antibody (ANA) testing strategy involving enzyme immunoassay (EIA) screening that reflexed to immunofluorescence assay (IFA) was implemented, monitored, and optimized for clinical utility. METHODS: The clinical utility, test performance, and workload implications of various ANA testing strategies were compared during the following study phases: (a) Preimplementation (n = 469) when IFA was used for all ANA screening, (b) Verification (n = 58) when EIA performance was confirmed, (c) Implementation (n = 433) when a reflexive strategy (EIA screen/IFA confirmation) was implemented, and (d) Postimplementation (n = 528) after the reflexive strategy was optimized. Sequential samples were captured in the Preimplementation, Implementation, and Postimplementation phases for clinical performance evaluation. RESULTS: Clinical performance of the EIA screen, per ROC analysis yielded area under the curve (AUC) of 0.846 in the Implementation phase and increased to 0.934 Postimplementation (P < 0.01); AUC for IFA similarly increased, from 0.678 to 0.808 (P = 0.05). The reflexive testing strategy increased screening sensitivity from 61% Preimplementation (IFA) to 98% (EIA) at Implementation and was maintained after optimization (98%, Postimplementation). Optimization decreased the false-positive rates for both EIA (from 40% to 18%) and IFA (18% to 8%) and was associated with reductions in daily full-time equivalent (by 33%) and IFA slide use (by 50%). CONCLUSIONS: Continuous quality monitoring approaches that incorporate sequential data sets can be used to evaluate, deploy, and optimize sensitive EIA-based ANA screening methods that can reduce manual IFA work without sacrificing clinically utility.

Comparison of POCT and central laboratory blood glucose results using arterial, capillary, and venous samples from MICU patients on a tight glycemic protocol.

BACKGROUND: Point of care (POC) glucose meters are routinely used to monitor glucose levels for patients on tight glycemic control therapy. We determined if glucose values were different for a POC glucose meter as compared to the main clinical laboratory for medical intensive care unit patients on a tight glycemic protocol and whether the site of blood sampling had a significant impact on glucose values. METHODS: Eighty-four patients (114 paired samples) who were on a tight glycemic protocol in the period November 2005 through August 2006 were enrolled. After simultaneous blood draws, we compared the glucose levels for the glucose meter (arterial/venous/capillary), blood gas (arterial/venous), and central clinical laboratory (serum/plasma from arterial/venous samples). RESULTS: The mean glucose levels of all arterial/venous/fingerstick samples using the glucose meter demonstrated a positive bias of 0.7-0.9 mmol/l (12.6-16.2 mg/dl) (p<0.001) relative to central laboratory venous plasma. There was also a smaller positive (0.1-0.3 mmol/l or 1.8-5.4 mg/dl, p<0.05) bias for arterial/venous blood gas samples and laboratory arterial serum/plasma glucose samples. Using Parkes error grid analysis we were able to show that the bias for arterial or venous POC glucose results would have not impacted clinical care. This was not the case, however, for fingerstick sampling where a high bias could have significantly impacted clinical care. Additionally, in 3 fingerstick samples a severe underestimation (<46% of the central laboratory plasma result) was found. CONCLUSION: Glucose meters using arterial/venous whole blood may be utilized in the MICU; however, due to the increased variability of results we do not recommend the routine use of capillary blood sampling for monitoring glucose levels in the MICU setting.