Fully activated structure of the sterol-bound Smoothened GPCR-Gi protein complex.

Smoothened (SMO) is an oncoprotein and signal transducer in the Hedgehog signaling pathway that regulates cellular differentiation and embryogenesis. As a member of the Frizzled (Class F) family of G protein-coupled receptors (GPCRs), SMO biochemically and functionally interacts with Gi family proteins. However, key molecular features of fully activated, G protein-coupled SMO remain elusive. We present the atomistic structure of activated human SMO complexed with the heterotrimeric Gi protein and two sterol ligands, equilibrated at 310 K in a full lipid bilayer at physiological salt concentration and pH. In contrast to previous experimental structures, our equilibrated SMO complex exhibits complete breaking of the pi-cation interaction between R4516.32 and W5357.55, a hallmark of Class F receptor activation. The Gi protein couples to SMO at seven strong anchor points similar to those in Class A GPCRs: intracellular loop 1, intracellular loop 2, transmembrane helix 6, and helix 8. On the path to full activation, we find that the extracellular cysteine-rich domain (CRD) undergoes a dramatic tilt, following a trajectory suggested by positions of the CRD in active and inactive experimental SMO structures. Strikingly, a sterol ligand bound to a shallow transmembrane domain (TMD) site in the initial structure migrates to a deep TMD pocket found exclusively in activator-bound SMO complexes. Thus, our results indicate that SMO interacts with Gi prior to full activation to break the molecular lock, form anchors with Gi subunits, tilt the CRD, and facilitate migration of a sterol ligand in the TMD to an activated position.

The mechanism for ligand activation of the GPCR-G protein complex.

G protein­coupled receptors (GPCRs) activate cellular responses ranging from odorants to neurotransmitters. Binding an agonist leads to activation of a heterotrimeric G protein (GP) that stimulates external signaling. Unfortunately, the mechanism remains unknown. We show for 15 class A GPCRs, including opioids, adrenergics, adenosines, chemokines, muscarinics, cannabinoids, serotonins, and dopamines, that interaction of an inactive GP, including Gs, Gi, Go, G11, and Gq, to the inactive GPCR, containing the intracellular ionic lock between transmembrane (TM) helices 3 and 6, evolves exothermically to form a precoupled GPCR-GP complex with an opened TM3-TM6 and the GP-α5 helix partially inserted into the GPCR but not activated. We show that binding of agonist to this precoupled GPCR-GP complex causes the Gα protein to open into its active form, with the guanosine diphosphate exposed for signaling. This GP-first paradigm provides a strategy for developing selective agonists for GPCRs since it is the pharmacophore for the precoupled GPCR-GP complex that should be used to design drugs.

Association between exercise habits and incident type 2 diabetes mellitus in patients with thyroid cancer: nationwide population-based study.

BACKGROUND: We investigated the association between exercise habits before or after thyroidectomy and incident type 2 diabetes mellitus (T2DM) in patients with thyroid cancer. METHODS: An observational cohort study of 69,526 thyroid cancer patients who underwent thyroidectomy for the treatment of thyroid cancer between 2010 and 2016 was performed using the Korean National Health Information Database. Regular exercise was defined as mid-term or vigorous exercise at least 1 day in a week based on a self-reported questionnaire. Patients were divided into four groups according to exercise habits before and after thyroidectomy: persistent non-exercisers, new exercisers, exercise dropouts, and exercise maintainers. RESULTS: During a median follow-up of 4.5 years, 2,720 (3.91%) patients developed T2DM. The incidence of T2DM per 1,000 person years was lower in patients who performed regular exercise before or after thyroidectomy than in persistent non-exercisers (10.77 in persistent non-exerciser group, 8.28 in new exerciser group, 8.59 in exercise dropout group, and 7.61 in exercise maintainer group). Compared with the persistent non-exerciser group, the new exerciser group (hazard ratio [HR] 0.87, 95% confidence interval [CI] 0.78-0.97), the exercise dropout group (HR 0.81, 95% CI 0.72-0.91), and the exercise maintainer group (HR 0.84, 95% CI 0.76-0.93) had lower risks of incident T2DM. Exercising < 1,500 MET-minutes/week in the exercise maintainer group was associated with a lower risk of incident T2DM compared with persistent non-exercisers (< 500: HR 0.80, 95% CI 0.67-0.96, P = 0.002; 500 to < 1,000: HR 0.81, 95% CI 0.71-0.93, P < 0.001; 1,000 to < 1,500: HR 0.81, 95% CI 0.69-0.94, P < 0.001). CONCLUSIONS: Regular exercise before or after thyroidectomy was associated with a lower risk of incident T2DM in patients with thyroid cancer.

An innovative microcoring technology: A novel approach to acne scar treatment.

BACKGROUND: Acne scars present a complex challenge in dermatology and cosmetics, despite advancements in technological interventions such as fractional lasers, microneedling, and surgical procedures. Effective treatment remains elusive for many individuals. OBJECTIVE: This study aims to evaluate the efficacy of rotational fractional resection using 1 mm diameter rotating scalpels as a primary treatment for icepick and boxcar scars on the cheeks and glabella region. METHODS: Three patients with acne scars underwent a single treatment session of rotational fractional resection. Evaluation occurred at the 2-month post-treatment mark to assess improvements in scar appearance and potential skin-related side effects. RESULTS: Following the treatment, significant improvements were observed in the targeted acne scars. Notable enhancements were noted without major skin-related adverse effects, except for minor suture marks. CONCLUSION: The outcomes of this study underscore the potential of rotational fractional resection as an innovative and effective approach in treating acne scars. This single-session cosmetic procedure shows promise in yielding lasting and quantifiable results, offering a hopeful solution for individuals seeking comprehensive acne scar treatment.

Structure and Molecular Mechanism of Signaling for the Glucagon-like Peptide-1 Receptor Bound to Gs Protein and Exendin-P5 Biased Agonist.

The glucagon-like peptide-1 receptor (GLP-1R) is a key regulator of blood glucose and a prime target for the treatment of type II diabetes and obesity with multiple public drugs. Here we present a comprehensive computational analysis of the interactions of the activated GLP-1R-Gs signaling complex with a G protein biased agonist, Exendin P5 (ExP5), which possesses a unique N-terminal sequence responsible for the signal bias. Using a refined all-atom model of the ExP5-GLP-1R-Gs complex in molecular dynamics (MD) simulations, we propose a novel mechanism of conformation transduction in which the unique interaction network of ExP5 N-terminus propagates the binding signal across an array of conserved residues at the transmembrane domain to enhance Gs protein coupling at the cytoplasmic end of the receptor. Our simulations reveal previously unobserved interactions important for activation by ExP5 toward GDP-GTP signaling, providing new insights into the mechanism of class B G protein-coupled receptor (GPCR) signaling. These findings offer a framework for the structure-based design of more effective therapeutics.

Multicenter performance evaluation of the Alinity m CMV assay for quantifying cytomegalovirus DNA in plasma samples.

Monitoring of cytomegalovirus (CMV) viral load is critical for informing treatment decisions in order to prevent the severe health consequences of CMV infection or reactivation of latent CMV in immunocompromised individuals. This first field evaluation examined the analytical and clinical performance of the Alinity m CMV assay. Analytical performance was assessed with a commercially available six-member panel, while the clinical performance evaluation compared the Alinity m CMV assay to the RealTime CMV assay and a laboratory-developed test (LDT) as the test of record at three large hospital-based clinical laboratories. Precision of the Alinity m CMV assay was demonstrated with total standard deviation (SD) between 0.08 and 0.28 Log IU/mL. A total of 457 plasma specimens were tested on the Alinity m CMV assay and compared to the test of record at each site (n = 304 with RealTime CMV and n = 153 with LDT CMV). The Alinity m CMV assay had excellent correlation (correlation coefficient r ≥0.942) in comparison to the RealTime CMV or LDT CMV assays. The mean observed bias ranged from -0.03 to 0.34 Log IU/mL. Median onboard turnaround time of Alinity m CMV was less than 3 h. When the CMV assay is run on the Alinity m system, it has the capacity to shorten time to result and, therefore, to therapy.

The atomistic level structure for the activated human κ-opioid receptor bound to the full Gi protein and the MP1104 agonist.

The kappa opioid receptor (κOR) is an important target for pain therapeutics to reduce depression and other harmful side effects of existing medications. The analgesic activity is mediated by κOR signaling through the adenylyl cyclase-inhibitory family of Gi protein. Here, we report the three-dimensional (3D) structure for the active state of human κOR complexed with both heterotrimeric Gi protein and MP1104 agonist. This structure resulted from long molecular dynamics (MD) and metadynamics (metaMD) simulations starting from the 3.1-Å X-ray structure of κOR-MP1104 after replacing the nanobody with the activated Gi protein and from the 3.5-Å cryo-EM structure of µOR-Gi complex after replacing the 168 missing residues. Using MD and metaMD we discovered interactions to the Gi protein with strong anchors to two intracellular loops and transmembrane helix 6 of the κOR. These anchors strengthen the binding, contributing to a contraction in the binding pocket but an expansion in the cytoplasmic region of κOR to accommodate G protein. These remarkable changes in κOR structure reveal that the anchors are essential for activation.

Mechanism of ß-arrestin recruitment by the µ-opioid G protein-coupled receptor.

Agonists to the µ-opioid G protein-coupled receptor (µOR) can alleviate pain through activation of G protein signaling, but they can also induce ß-arrestin activation, leading to such side effects as respiratory depression. Biased ligands to µOR that induce G protein signaling without inducing ß-arrestin signaling can alleviate pain while reducing side effects. However, the mechanism for stimulating ß-arrestin signaling is not known, making it difficult to design optimum biased ligands. We use extensive molecular dynamics simulations to determine three-dimensional (3D) structures of activated ß-arrestin2 stabilized by phosphorylated µOR bound to the morphine and D-Ala2, N-MePhe4, Gly-ol]-enkephalin (DAMGO) nonbiased agonists and to the TRV130 biased agonist. For nonbiased agonists, we find that the ß-arrestin2 couples to the phosphorylated µOR by forming strong polar interactions with intracellular loop 2 (ICL2) and either the ICL3 or cytoplasmic region of transmembrane (TM6). Strikingly, Gi protein makes identical strong bonds with these same ICLs. Thus, the Gi protein and ß-arrestin2 compete for the same binding site even though their recruitment leads to much different outcomes. On the other hand, we find that TRV130 has a greater tendency to bind the extracellular portion of TM2 and TM3, which repositions TM6 in the cytoplasmic region of µOR, hindering ß-arrestin2 from making polar anchors to the ICL3 or to the cytosolic end of TM6. This dramatically reduces the affinity between µOR and ß-arrestin2.

Immunogenicity of SARS-CoV-2 Vaccine in Kidney Transplant Recipients: A Cross-Sectional Study in Korea.

Eighty-five Korean kidney transplant recipients who received three doses of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine were tested with anti-receptor binding domain (RBD) antibody and neutralizing antibody. High anti-RBD antibody (≥ 100 U/mL) and neutralizing antibody responses (≥ 30%) were detected in 51/85 (60.0%) patients. When we divided the patients with the time from transplantation to vaccination (< 1, 1-2.4, 2.5-4.9, and ≥ 5-year), anti-RBD antibody titers were 3.2 U/mL, 27.8 U/mL, 370.2 U/mL, and 5,094.2 U/mL (P < 0.001) and anti-neutralizing antibody levels were 2.2%, 11.6%, 45.6%, and 93.0% (P < 0.001), respectively. Multivariate analysis revealed increased antibody responses when the time from transplantation to vaccination was five years or longer (odds ratio, 12.0; confidence interval, 2.7-52.8). Korean kidney transplant recipients had suboptimal antibody responses after the third dose of SARS-CoV-2 vaccine. A shorter time from transplantation to vaccination was a risk factor for a low antibody response.

Comparison between polynomial regression and weighted least squares regression analysis for verification of analytical measurement range.

OBJECTIVES: Recently, the linearity evaluation protocol by the Clinical & Laboratory Standards Institute (CLSI) has been revised from EP6-A to EP6-ED2, with the statistical method of interpreting linearity evaluation data being changed from polynomial regression to weighted least squares linear regression (WLS). We analyzed and compared the analytical measurement range (AMR) verification results according to the present and prior linearity evaluation guidelines. METHODS: The verification of AMR of clinical chemistry tests was performed using five samples with two replicates in three different laboratories. After analyzing the same evaluation data in each laboratory by the polynomial regression analysis and WLS methods, results were compared to determine whether linearity was verified across the five sample concentrations. In addition, whether the 90% confidence interval of deviation from linearity by WLS was included in the allowable deviation from linearity (ADL) was compared. RESULTS: A linearity of 42.3-56.8% of the chemistry items was verified by polynomial regression analysis in three laboratories. For analysis of the same data by WLS, a linearity of 63.5-78.3% of the test items was verified where the deviation from linearity of all five samples was within the ADL criteria, and the cases where the 90% confidence interval of all deviation from linearity overlapped the ADL was 78.8-91.3%. CONCLUSIONS: Interpreting AMR verification data by the WLS method according to the newly revised CLSI document EP6-ED2 could reduce laboratory workload, enabling efficient laboratory practice.

Evaluation of Serum Beta-2-Microglobulin Assay on Beckman Coulter Chemistry Analyzer.

BACKGROUND: Beta-2 microglobulin (ß2M) has various clinical usages and is an important prognostic marker for multiple myeloma. Although there are concerns of harmonization between assays, performance evaluation and method comparison reports are rare. Here, we evaluated the performance of Beckman-Coulter ß2M assay. METHODS: The precision and linearity of the Beckman Coulter ß2M assay were evaluated. Beckman-Coulter results were compared to DiaSorin Liaison results. The manufacturer provided reference interval was verified. RESULTS: Within-laboratory imprecision percentage coefficient of variation was 1.73% and 2.19% for low- and high-level control material, respectively. The linearity was confirmed over a range of 0.08 to 15.60 mg/L. Passing-Bablok regression analysis showed that y (Beckman-Coulter) = 0.978 x (Liaison) + 0.079 (r = 0.996) with a mean difference of 1.0%. All healthy subjects were within the range of manufacturer provided reference interval. CONCLUSIONS: The analytical performance of the Beckman-Coulter ß2M assay was clinically acceptable.

Performance of the New RAPIDPoint 500e Blood Gas Analyzer and its Comparison with Central Laboratory Biochemistry Analyzer.

BACKGROUND: Blood gas analyzers are commonly used in point of care settings in hospitals to assess and monitor critically ill patients. The blood gas analyzer, RAPIDPoint 500e (Siemens Healthcare, UK), has been newly introduced. Here, we evaluated the performance of RAPIDPoint 500e for the determination of pH, partial carbon dioxide pressure, partial oxygen pressure, sodium (Na+), potassium (K+), chloride (Cl-), ionized calcium, glucose, and lactate. METHODS: Repeatability, within-laboratory imprecision, and linearity were assessed according to CLSI guidelines. RAPIDPoint 500e results were compared to its previous version RAPIDPoint 500. For Na+, K+, and Cl-, RAPID¬Point 500e results were also compared to the central chemistry analyzer AU5822 (Beckman Coulter, Brea, CA, USA). RESULTS: The coefficients of variation of within-laboratory imprecision for all analytes were less than 5%, within the range of acceptable criteria. The linearity values for all analytes were confirmed within manufacturer claimed range. RAPIDPoint 500e correlated well with both RAPIDPoint 500 and AU5822. Correlation coefficients were over 0.95 for all analytes. CONCLUSIONS: The analytical performance of RAPIDPoint 500e was clinically acceptable.

Analytical Performance Evaluation of Goldstream Fungus (1-3)-ß-D Glucan.

BACKGROUND: (1-3)-ß-D-glucan (BDG) is a fast and simple assay to diagnose invasive fungal infection. In this study, we evaluated the performance of the Goldstream BDG assay (Beijing Gold Mountainriver Tech Development) performed on the automated analyzer, IGL-200 (Genobio Pharmaceutical). METHODS: The precision and linearity of the Goldstream BDG assay were evaluated according to Clinical and Laboratory Standards Institute procedures. BDG results performed on the IGL-200 were compared to a manual photometer, MB-80A (Genobio Pharmaceutical). The manufacturer-provided reference interval was verified. RESULTS: Within-laboratory imprecision (% Coefficient of Variation) was 9.4%. The best polynomial fit was third-order within the manufacturer's claimed linear range (32.0 - 830.0 pg/mL). The BDG assay performed on IGL-200 and MB-80A showed a total agreement of 97.6%. All healthy subjects were within range of the manufacture provided reference interval. CONCLUSIONS: The analytical performance of the Goldstream BDG assay was clinically acceptable.

Novel interaction between neurotrophic factor-α1/carboxypeptidase E and serotonin receptor, 5-HTR1E, protects human neurons against oxidative/neuroexcitotoxic stress via ß-arrestin/ERK signaling.

Protecting neurons from death during oxidative and neuroexcitotoxic stress is key for preventing cognitive dysfunction. We uncovered a novel neuroprotective mechanism involving interaction between neurotrophic factor-α1 (NF-α1/carboxypeptidase E, CPE) and human 5-HTR1E, a G protein-coupled serotonin receptor with no previously known neurological function. Co-immunoprecipitation and pull-down assays confirmed interaction between NFα1/CPE and 5-HTR1E and 125I NF-α1/CPE-binding studies demonstrated saturable, high-affinity binding to 5-HTR1E in stably transfected HEK293 cells (Kd = 13.82 nM). Treatment of 5-HTR1E stable cells with NF-α1/CPE increased pERK 1/2 and pCREB levels which prevented a decrease in pro-survival protein, BCL2, during H2O2-induced oxidative stress. Cell survival assay in ß-arrestin Knockout HEK293 cells showed that the NF-α1/CPE-5-HTR1E-mediated protection against oxidative stress was ß-arrestin-dependent. Molecular dynamics studies revealed that NF-α1/CPE interacts with 5-HTR1E via 3 salt bridges, stabilized by several hydrogen bonds, independent of the serotonin pocket. Furthermore, after phosphorylating the C-terminal tail and intracellular loop 3 (ICL3) of NF-α1/CPE-5-HTR1E, it recruited ß-arrestin1 by forming numerous salt bridges and hydrogen bonds to ICL2 and ICL3, leading to activation of ß-arrestin1. Immunofluorescence studies showed 5-HTR1E and NF-α1/CPE are highly expressed and co-localized on cell surface of human hippocampal neurons. Importantly, knock-down of 5-HTR1E in human primary neurons diminished the NF-α1/CPE-mediated protection of these neurons against oxidative stress and glutamate neurotoxicity-induced cell death. Thus, NF-α1/CPE uniquely interacts with serotonin receptor 5-HTR1E to activate the ß-arrestin/ERK/CREB/BCL2 pathway to mediate stress-induced neuroprotection.

Assay Sensitivity Difference Can Induce Anti-Hepatitis A Virus IgM Non-Reactive But Total (IgM and IgG) Reactive Results in Early Acute Hepatitis A.

Although anti-hepatitis A virus (HAV) IgM non-reactive and anti-HAV total (immunoglobulin [Ig] M and IgG) reactive results are generally interpreted as immunity to HAV, some early acute hepatitis A patients show the same results. We compared IgM detection sensitivity between anti-HAV IgM and anti-HAV total assays. Acute hepatitis A patients' samples were serially diluted and tested with Elecsys anti-HAV IgM and total assay (Roche Diagnostics). This resulted in anti-HAV IgM non-reactive but anti-HAV total reactive results. Samples of two hepatitis A patients showing false-negative anti-HAV IgM at initial presentation were analyzed with Elecsys, Atellica (Siemens Healthineers), and Alinity (Abbott Laboratories) HAV assays. Elecsys, Atellica, and Alinity anti-HAV IgM converted reactive on hospital day 3, whereas Elecsys and Atellica anti-HAV total results were reactive from hospital day 1. The anti-HAV total assay had higher sensitivity in detecting IgM antibodies than the anti-HAV IgM assay.

Predicted Structure of Fully Activated Tas1R3/1R3' Homodimer Bound to G Protein and Natural Sugars: Structural Insights into G Protein Activation by a Class C Sweet Taste Homodimer with Natural Sugars.

The Tas1R3 G protein-coupled receptor constitutes the main component of sweet taste sensory response in humans via forming a heterodimer with Tas1R2 or a homodimer with Tas1R3. The Tas1R3/1R3' homodimer serves as a low-affinity sweet taste receptor, stimulating gustducin G protein (GGust) signaling in the presence of a high concentration of natural sugars. This provides an additional means to detect the taste of natural sugars, thereby differentiating the flavors between natural sugars and artificial sweeteners. We report here the predicted 3D structure of active state Tas1R3/1R3' homodimer complexed with heterotrimeric GGust and sucrose. We discovered that the GGust makes ionic anchors to intracellular loops 1 and 2 of Tas1R3 while the Gα-α5 helix engages the cytoplasmic region extensively through salt bridge and hydrophobic interactions. We show that in the activation of this complex the Venus flytrap domains of the homodimer undergo a remarkable twist up to ∼100° rotation around the vertical axis to adopt a closed-closed conformation while the intracellular region relaxes to an open-open conformation. We find that binding of sucrose to the homodimer stabilizes a preactivated conformation with a largely open intracellular region that recruits and activates the GGust. Upon activation, the Gα subunit spontaneously opens up the nucleotide-binding site, making nucleotide exchange facile for signaling. This activation of GGust promotes the interdomain twist of the Venus flytrap domains. These structures and transformations could potentially be a basis for the design of new sweeteners with higher activity and less unpleasant flavors.

Performance of Xpert MTB/RIF for the Diagnosis of Extrapulmonary Tuberculosis.

BACKGROUND: Mycobacterial burden is low in extrapulmonary specimens, making diagnosis and treatment difficult. Xpert MTB/RIF is a real-time PCR assay for the detection of Mycobacterium tuberculosis and rifampin resistance. This study evaluated the performance of the Xpert MTB/RIF assay in extrapulmonary specimens. METHODS: Acid-Fast Bacilli (AFB) smear, culture, and Xpert MTB/RIF were performed on extrapulmonary specimens. Mycobacterial culture was performed on BACTEC MGIT liquid for 6 weeks and 2% Ogawa medium for 8 weeks. Overall sensitivity and specificity of Xpert MTB/RIF was estimated using culture as a gold standard. Xpert MTB/RIF sensitivity and cycle-threshold (Ct) values according to AFB smear grade were evaluated. The sensitivity, specificity, and concordance of rifampin resistance compared to the phenotypic drug sensitivity test were evaluated. RESULTS: A total of 1,289 specimens were included in the study. The overall sensitivity and specificity of the Xpert MTB/RIF assay were 59.4% (41/69, 95% CI 46.9 - 70.9%) and 99.3% (1,212/1,220, 95% CI 98.7 - 99.7), respectively. Positive predictive value of Xpert MTB/RIF was 83.7% (41/49, 95% CI 69.8 - 92.2) and negative predictive value was 97.7% (1,212/1,240, 95% CI 96.7 - 98.5%). Xpert MTB/RIF assay sensitivity significantly increased with increases in AFB smear grade (p < 0.001). AFB smear grades and Xpert MTB/RIF Ct values were negatively correlated. Rifampin resistance results of Xpert MTB/RIF and culture showed a concordance rate of 97.2%. CONCLUSIONS: The Xpert MTB/RIF assay could be used to replace the AFB smear for the diagnosis of extrapulmonary tuberculosis, and has high specificity for the detection of rifampin resistance.

Sample size and rejection limits for detecting reagent lot variability: analysis of the applicability of the Clinical and Laboratory Standards Institute (CLSI) EP26-A protocol to real-world clinical chemistry data.

Objectives: To maintain the consistency of laboratory test results, between-reagent lot variation should be verified before using new reagent lots in clinical laboratory. Although the Clinical and Laboratory Standards Institute (CLSI) document EP26-A deals with this issue, evaluation of reagent lot-to-lot difference is challenging in reality. We aim to investigate a practical way for determining between-reagent lot variation using real-world data in clinical chemistry. Methods: The CLSI EP26-A protocol was applied to 83 chemistry tests in three clinical labs. Three criteria were used to define the critical difference (CD) of each test as follows: reference change value and total allowable error, which are based on biological variation, and acceptable limits by external quality assurance agencies. The sample size and rejection limits that could detect CD between-reagent lots were determined. Results: For more than half of chemistry tests, reagent lot-to-lot differences could be evaluated using only one patient sample per decision level. In many cases, the rejection limit that could detect reagent lot-to-lot difference with ≥90% probability was 0.6 times CD. However, the sample size and rejection limits vary depending on how the CD is defined. In some cases, impractical sample size or rejection limits were obtained. In some cases, information on sample size and rejection limit that met intended statistical power was not found in EP26-A. Conclusions: The CLSI EP26-A did not provide all necessary answers. Alternative practical approaches are suggested when CLSI EP26-A does not provide guidance.

Performance Evaluation of the Third-Generation Elecsys FT4 III Assay for Free Thyroxine.

BACKGROUND: Roche Diagnostics has recently released the Elecsys FT4 III, Roche's third-generation reagent for free thyroxine. We aim to evaluate the analytical performance of the Elecsys FT4 III. METHODS: The precision and linearity of the Elecsys FT4 III assay were evaluated according to Clinical and Laboratory Standards Institute procedures. The results of the third-generation assay were compared with that of Roche's second-generation assay (Elecsys FT II) on a Roche cobas e801 modular analyzer. RESULTS: The coefficient of variation of within-laboratory variation was less than 2.6% for the control materials and patient samples. The linearity was confirmed over a range of 0.41 to 7.52 ng/dL. The mean difference between the second- and third-generation reagents was 0.0026 ng/dL (95% confidence interval, -0.0172 to +0.0224 ng/dL). CONCLUSIONS: The analytical performance of the Elecsys FT4 III assay for free thyroxine is clinically acceptable.

Performance Evaluation of the Elecsys Growth Hormone and Insulin-Like Growth Factor 1 Assay.

BACKGROUND: Growth hormone (GH) and insulin-like growth factor 1 (IGF-1) are important biomarkers for diagnosis and follow-up of growth-related disorders. Performance of Elecsys hGH and Elecsys IGF-1 (Roche Diagnostics) was evaluated. METHODS: The performance of the Elecsys hGH and Elecsys IGF-1 assays on a Roche Cobas e 801 modular analyzer were evaluated. Repeatability, within-laboratory imprecision, and linearity were assessed. Elecsys hGH was compared to hGH [I-125] IRMA (Izotop), and Elecsys IGF-1 was compared to IRMA IGF-1 (Immunotech). RESULTS: The coefficient of variation of within-laboratory imprecision for Elecsys hGH and IGF-1 was less than 1.7%. The linearity values for Elecsys hGH and IGF-1 were confirmed over a range of 0.05 - 24.70 ng/mL and 40.41 - 687.53 ng/mL, respectively. The mean difference between Elecsys hGH and IRMA, and Elecsys IGF-1 and IRMA was 0.284 ng/mL and 14.21 ng/mL, respectively. CONCLUSIONS: The analytical performance of Elecsys hGH and Elecsys IGF-1 was clinically acceptable.