Nerenz, Robert D

Abstract Background Historically, 25-hydroxyvitamin D (25OHD) assays have under- or over-recovered 25-hydroxyvitamin D2 (25OHD2), but assay manufacturers have modified their reagents to address this problem. In this study, we compared the second- and third-generation Roche assays as well as two contemporary offerings from Diasorin and Beckman against liquid chromatography–tandem mass spectrometry (LC-MS/MS). Methods We identified 50 remnant serum samples with 25OHD concentrations from across the analytical range of the second-generation Roche assay. To increase 25OHD2 representation, we identified 25 additional samples from individuals prescribed high-dose vitamin D2 supplements. We tested samples on Roche assays and circulated to laboratories performing Beckman and Diasorin 25OHD assays. We tested samples by LC-MS/MS to obtain concentrations for 25OHD2 and 25-hydroxyvitamin D3. Results Mean overall bias for each assay was 5.1 ng/mL or less against the LC-MS/MS measurement; mean proportional bias was 8.7% to 12.1%. Some individual specimens had much larger bias. 25OHD2 was under-recovered on average, but the bias for the third-generation Roche assay represents a significant improvement over the previous assay, and mean bias for current generation assays was no worse than −3.2 ng/mL. In most cases, clinical classification by automated assay values agreed with clinical classification by LC-MS/MS; where present, disagreements occurred near classification thresholds. Conclusions Automated 25OHD assays continue to improve, and 25OHD2 recovery no longer appears to be a significant concern for the assays evaluated here. All assays evaluated were adequate for clinical classification of vitamin D nutritional status and are suitable for routine use, including in patients prescribed high-dose vitamin D2.

Abstract Background We took advantage of our verification of a new biotin-resistant 25-hydroxyvitamin D (vitamin D) immunoassay formulation to compare analytical performance of the outgoing formulation and assays from Roche, Diasorin and Beckman. Liquid chromatography with tandem mass spectrometry (LCMS) was an independent arbiter of vitamin D concentration and provided quantification of vitamin D2 and vitamin D3. Methods Twenty-five specimens from patients on high-dose vitamin D2 supplementation (50 000 IU/week) and fifty additional specimens spanning our measurable range were identified by the Dartmouth Hitchcock Analytics Institute. Aliquots were tested by the Roche cobas Vitamin D Total II and Vitamin D Total III assays, deidentified, split, and shipped to other study sites for measurement by LCMS, the Diasorin Liaison 25 OH Vitamin D Total assay, and the Beckman Coulter Access 25(OH) Vitamin D Total assay. Estimated vitamin D2 was calculated by subtracting the LCMS-measured vitamin D3 result from the immunoassay total vitamin D result. Bias calculations assume LCMS measurements as the true concentration. Statistical analyses were performed in R. This study was approved by the Dartmouth Hitchcock Medical Center Institutional Review Board. Results No significant differences were observed between platforms in absolute or relative bias in total vitamin D or estimated vitamin D2. Clinical classification was unanimous in 62/75 specimens. For thirteen specimens in which at least one disagreement was observed, immunoassay results for discrepant classification were no more than 10.2 ng/mL from the classification cutoff (median 2 ng/mL). Two specimens with 68 and 70 ng/mL vitamin D were classified as toxic (>100 ng/mL) by one immunoassay each. Conclusion Current vitamin D immunoassays continue to improve over their predecessors and offer good agreement across platforms. Vitamin D2 recovery no longer represents a clinically-concerning obstacle. These immunoassays display positive bias for total vitamin D relative to LCMS but generally result in correct clinical classification.

Abstract Background The AACC Academy revised the reproductive testing section of the Laboratory Medicine Practice Guidelines: Evidence-Based Practice for Point-of-Care Testing (POCT) published in 2007. Methods A panel of Academy members with expertise in POCT and laboratory medicine was formed to develop guidance for the use of POCT in reproductive health, specifically ovulation, pregnancy, premature rupture of membranes (PROM), and high-risk deliveries. The committee was supplemented with clinicians having Emergency Medicine and Obstetrics/Gynecology training. Results Key recommendations include the following. First, urine luteinizing hormone (LH) tests are accurate and reliable predictors of ovulation. Studies have shown that the use of ovulation predicting kits may improve the likelihood of conception among healthy fertile women seeking pregnancy. Urinary LH point-of-care testing demonstrates a comparable performance among other ovulation monitoring methods for timing intrauterine insemination and confirming sufficient ovulation induction before oocyte retrieval during in vitro fertilization. Second, pregnancy POCT should be considered in clinical situations where rapid diagnosis of pregnancy is needed for treatment decisions, and laboratory analysis cannot meet the required turnaround time. Third, PROM testing using commercial kits alone is not recommended without clinical signs of rupture of membranes, such as leakage of amniotic fluid from the cervical opening. Finally, fetal scalp lactate is used more than fetal scalp pH for fetal acidosis due to higher success rate and low volume of sample required. Conclusions This revision of the AACC Academy POCT guidelines provides recommendations for best practice use of POCT in fertility and reproduction.

Abstract Background Loperamide (Imodium®), a commonly used anti-diarrheal, is a mu opioid receptor agonist that, like all opioids, reduces gastrointestinal tract peristalsis. Loperamide is considered to have low abuse potential as it does not produce an analgesic or euphoric effect due to low bioavailability and first-pass metabolism. However, reports of individuals misusing loperamide through the use of super-therapeutic doses, alone or in combination with P-glycoprotein and/or CYP450 enzyme inhibitors, is increasing. We hypothesized that loperamide could potentially cross-react with laboratory immunoassay drug screens. Methods Drug-free urine was spiked with loperamide or its principal metabolite, N-desmethyl loperamide (dLop), and assayed on multiple fentanyl and buprenorphine assays. Fentanyl immunoassay screen-positive results at one institution were examined by high-resolution mass spectrometry (MS) for the presence of loperamide and quantified by liquid chromatography– tandem MS when positive. Results Loperamide produced positive results on the Thermo DRI Fentanyl and Immunalysis Fentanyl assays at concentrations greater than 5.72 mg/L and 23.7 mg/L. dLop generated positive results for the Thermo DRI and Immunalysis fentanyl assays at concentrations exceeding 6.9 mg/L and 35.7 mg/L. dLop also produced positive buprenorphine results on the Thermo CEDIA buprenorphine assay at concentrations exceeding 12.2 mg/L. High-resolution MS analysis of 225 fentanyl immunoassay positives (Thermo DRI) yielded 5 specimens containing loperamide and/or dLop, 4 of which contained measurable quantities of fentanyl in addition to loperamide/dLop. Conclusions Laboratories using these assays should be aware of the potential for false-positive screening results due to the presence of high concentrations of loperamide and its metabolite dLop.

Abstract Background Despite recommendations to limit vitamin D testing to specific clinical scenarios, test volume remains high in many clinical laboratories. Automated total vitamin D immunoassays frequently under- or over-recover 25-hydroxyvitamin D2 [25(OH)D2], making accurate assessment of vitamin D status difficult in patients taking high-dose 25(OH)D2 supplements. Mass spectrometry-based methods offer excellent recovery of 25(OH)D2 but are not practical for use in all laboratories. In this study, we evaluated 2 automated immunoassays against an LC–MS/MS method performed at a national reference laboratory. Methods A method comparison against LC–MS/MS was performed for the Roche Elecsys Vitamin D total II assay and the IDS-iSYS 25 VitDS immunoassays using 49 patient specimens submitted for clinical 25(OH)D measurement. Mean bias was calculated, and vitamin D status was determined for each specimen according to the 2011 Endocrine Society clinical practice guidelines. Results Theil–Sen regression lines relative to LC–MS/MS were y = 0.88x + 2.94 for Roche and y = 1.03x + 2.48 for IDS. Mean bias (±SD) in samples with 25(OH)D2 concentrations less than 5 ng/mL was −0.25 ng/mL (±6.30) for Roche and −1.45 ng/mL (±6.82) for the IDS. Mean bias (±SD) in samples with 25(OH)D2 concentrations greater than 5 ng/mL was −3.19 ng/mL (±6.61) for Roche and 5.52 ng/mL (±6.36) for IDS. Median percentage recovery of 25(OH)D2 was 87.1% (interquartile range 76.0–111.3) for Roche and 120.6% (interquartile range: 105.3–133.4) for IDS. Vitamin D status was misclassified in 7 samples by the Roche assay and 3 by the IDS assay. For all but one of the discordant pairs, the immunoassay result was within 1.7 ng/mL of the diagnostic cutoff. Conclusions The automated immunoassays evaluated here demonstrate improved recovery of 25(OH)D2 relative to previous generations. Both are acceptable for use in the determination of vitamin D status.