Method Performance Verification Research Articles

Headspace Gas Chromatography Mass Spectrometry Method for Determination of class-I Residual Solvents in Several Drug Substances: Method Evaluation by Quality by Design Statistical Tool.

Class-I residual solvents such as 1,1-dichloroethene, 1,1,1-trichloroethane, carbon tetrachloride, benzene, 1,2-dichloroethane are toxic, environmental hazard and carcinogenic to human. Headspace gas chromatography mass spectrometer is the sophisticated instrument for quantification of residual solvents at lower limits. An exact, sensitive, reliable and fast method was developed to determine the 1,1-dichloroethene, 1,1,1-trichloroethane, carbon tetrachloride, benzene and 1,2-dichloroethane present in different drug substances by using a headspace gas chromatography mass spectrometer. Helium is used as a carrier gas in this method. N-Methyl-2-pyrrolidone is used as a diluent and the stationary phase is a DB-624 (60 m x 0.25 mm x 1.4 μm film thickness) column with a flow rate of 1.5 mL/min. The concentration of limit of detection for 1,1-dichloroethene, 1,1,1-trichloroethane, carbon tetrachloride, benzene and 1,2-dichloroethane were achieved with 0.24 ppm, 5 ppm, 0.12 ppm, 0.06 ppm, 0.15 ppm. Subsequently, the concentrations of limit of quantification for aforementioned impurities were achieved with 0.8 ppm, 15 ppm, 0.4 ppm, 0.2 ppm, 0.5 ppm. The linearity was covered in the range of LOQ to 120% with respect to the specification level. The current method's system suitability, precision, linearity, accuracy parameters were performed in accordance to the USP < 1225> and ICH Q2(R2) and results were lies within the acceptance criteria. No research studies are reported for the determination of class-I residual solvents in lincomycin hydrochloride, dapagliflozin, vonoprazan fumarate, telmisartan drug substances. The proposed study is to develop the common method for quantification of class-I residual solvents for drug substances. QbD concept is utilized in the method performance verification.

Investigation of select radionuclides stability in urine under various conditions for liquid scintillation counting (LSC).

Liquid Scintillation Counting (LSC) gross alpha/beta screening is a valuable tool for providing rapid laboratory response for the analysis of human clinical urine samples during a large-scale radiation incident event. Verification of method performance, as required for clinical laboratory testing, is accomplished by the evaluation of routine, periodic measurements of radioactive spiked samples for quality control, performance testing, and accuracy checks. Radionuclide stability of alpha and beta emitters in urine for LSC analysis is an important consideration. The purpose of this work is to demonstrate optimal preparations and storage conditions of samples used for method verification.

A-149 Performance Characteristics of VerifyNow PRU and ARU Tests During a Clinical Method Verification Exercise

Abstract Background Dual antiplatelet therapy with aspirin and a platelet P2Y12 receptor antagonist is indicated for the prevention of atherothrombotic events in patients with acute coronary syndromes and for those undergoing percutaneous coronary intervention procedure. Two different assays are available in the VerifyNow Test device for monitoring the anti-platelet therapy response. The aspirin assay (ARU Test) utilizes arachidonic acid as an agonist and is sensitive to acyl salicylic acid. The P2Y12 assay (PRU test) is sensitive to thienopyridines (such as clopidogrel, prasugrel, ticagrelor) and uses adenosine diphosphate (ADP) as an agonist. The PRU test is a non-waived test and ARU test is a waived test. A comprehensive method performance verification is required for the PRU test. We evaluated performance characteristics of the VerifyNow PRU Test and ARU Test for clinical use. Methods The standard precision and accuracy study using two levels (low and high) of quality control materials were performed (N = 10) for both the assays. Alternative qualitative method comparison was carried out (N = 49) using the VerifyNow PRU Test in a reference lab and in-house. The in-house inter-instrument comparisons were carried out (N = 42). The blood samples were obtained from patients taking P2Y12 inhibitors and from normal donors not on any drugs. The results of the PRU Test are expressed in P2YI2 reaction units (PRU). The VerifyNow-Aspirin assay results are expressed as aspirin reaction units (ARU). The statistical analysis to evaluate performance characteristics was carried out using the EP Evaluator. Results The VerifyNow device 1561 precision characteristics for low QC (Obs Mean = 1.5 PRU (N = 10), SD = 0.2; 95% CI for Obs Mean 1.0 to 2.0 PRU) and high QC (Obs Mean = 258.3 PRU (N = 10), SD = 8.2; 95% CI for Obs Mean 252.4 to 264.2 PRU) were within acceptable performance limits. The CV for high QC was 3.2%. The ARU test also demonstrated high precision for two levels (Low QC Obs Mean = 350ARU (N = 10), SD = 0.0); High QC Obs Mean = 583.3 (N = 10), SD = 7.2; 95% CI for Obs Mean = 578.1–588.5) with a CV of 1.2%. The PRU Test Method PRU 1561 In House vs Ref Method Verify Now PRU Test (N = 49) demonstrated overall agreement of 93.9% with positive agreement (intended use population on drugs, N = 27) of 92% and negative agreement (normal non-drug group, N = 22) of 95.8% (Cohen’s Kappa = 87.8%; Kappa &amp;gt;75% indicates “"high” agreement). The McNemar Test for Symmetry passed (P &amp;gt; 0.999, Fisher’s Exact test) in this qualitative method comparison. A value of P &amp;lt; 0.05 would suggest the methods are significantly different. The other in-house instrument 1562 also demonstrated comparable or better performance characteristics. Conclusion It is important to confirm the medical decision limit for the intended use population and determine acceptance criteria while considering biologic variability. However, the reference range verifications with drug responders and non-responders are not always feasible and cost-effective. Based on the qualitative method comparison in this study, we have transferred the medical decision limits from the peer group. The in-house multi-instrument comparison should be repeated every six months to monitor the performance of the instruments.

Chinese expert consensus on clinical practice of quantitative real-time polymerase chain reaction techniques in molecular pathological diagnostics of tumor(2023 edition)

Quantitative real-time polymerase chain reaction (qPCR) is one of the most widely used molecular pathological diagnostic techniques in China due to its advantages of the simple operation, short turnaround time, high sensitivity, and standardizable result analysis. However, in clinical practice, there is not yet an expert consensus to guide molecular pathological diagnostics of tumor using qPCR techniques in terms of validation and verification of method performance, quality management and interpretation of complex results. Therefore, this expert consensus aims to provide standardized opinions on the practical application of qPCR techniques, and suggestions on how to deal with common problems and abnormal results, and reach a Chinese expert consensus on the clinical practice of qPCR techniques in molecular pathological diagnostics of tumor, in order to standardize the testing process, improve the accuracy of results, and promote the clinical applications of qPCR techniques.

Quality by design tool-evaluated stability-indicating ultra-performance liquid chromatography method for the determination of drugs (ritonavir and darunavir) used to treat the human immunodeficiency virus/acquired immunodeficiency syndrome.

Ritonavir and darunavir were examined using a ultra-performance liquid chromatography (UPLC) approach in pharmaceutical dosage forms. The small number of analytical studies that are currently available do not demonstrate the method's stability or nature. The study sought to assess both chemicals using a stability-indicating approach with a relatively short run time. The HSS C18 (100 × 2.1 mm), 2-mm column was used for the chromatographic separation, and isocratic elution was used to achieve this. In the mobile phase, methanol and 0.01 M phosphate buffer (pH 4.0) were included in a 60:40 (v/v) ratio. Throughout the analysis, the flow rate was kept at 0.2 mL min-1 , and a photodiode array detector set to 266 nm was used to find the major components. The proposed method showed a linear response (r2 > 0.999), and the accuracy was between 98.0% and 102.0%. The precision data showed relative standard deviation ≤1.0%. The UPLC method for quantification of ritonavir and darunavir in pharmaceutical dosage forms using a very short run time of under a minute is the subject of the proposed article. To meet current regulatory criteria, the quality by design idea was used in the method performance verification.

Estimation of the Detection Limit of Fecal Coliforms and &lt;i&gt;Staphylococcus aureus&lt;/i&gt; Qualitative Methods

ISO/IEC 17025 : 2017 requires laboratories to verify the performance of methods before introducing them. ISO 16140-3 : 2021 employs LOD50 (the level of detection for which 50% of tests give a positive result) as an index for evaluating the performance of microbial qualitative methods. While fecal coliforms and Staphylococcus aureus qualitative methods are widely implemented in Japan, their LOD50 values have not been established, making performance evaluation of these methods impossible at the time of introduction. As a result, we looked into estimating the LOD50 values of fecal coliforms and S. aureus qualitative methods to evaluate the index of method performance. The fecal coliforms method had an LOD50 value of 19–31 CFU/mL and the S. aureus method had an LOD50 value of 29–49 CFU/mL by laboratory personnels using samples without food matrix. We estimated LOD50 of six food samples using each method in a single laboratory. The LOD50 values of all foods calculated using the fecal coliforms method were in the range of 14–27 CFU/g and those calculated using the S. aureus method were in the range of 25–48 CFU/g, indicating that these methods did not have low levels of sensitivity detection compared with skill variation. LOD50 values for food samples with the highest or lowest LOD50 in each method were compared among five laboratories. Because the estimated LOD50 values of both the methods were within four times of each laboratory result, which is the acceptable limit specified in ISO 16140-3 : 2021, they were assumed to be suitable values. This study is expected to be utilized as a benchmark in the verification of method performance in each laboratory.

First morning voided urinary gonadotropins in children: verification of method performance and establishment of reference intervals.

Urinary luteinizing hormone (uLH) and urinary follicle-stimulating hormone (uFSH) have been shown to be useful screening and management tools for children with central precocious puberty. However, studies on uLH and uFSH reference intervals are scarce. Therefore, we aimed to establish reference intervals for uLH and uFSH, according to age, sex, and pubertal status in apparently healthy children aged 6-11years. We performed detection capability, precision, accuracy by recovery, linearity, agreement analysis, and stability testing to analyze the method performance of uLH and uFSH. The Clinical Laboratory Standards Institute's C28-A3 criteria was used to establish the reference intervals. Both uLH and uFSH were stable at 4°C for 52.6h and 64.8days, respectively. The total imprecision of uFSH is within the manufacturer's claim, while the total imprecision of uLH remained within tolerable bias. Both uLH and uFSH could be measured with acceptable detection capability. The recovery rates of the hormones were 87.6-98.8% and 102.8-103.4%, respectively, and therefore within acceptable limits. There were significant correlations between the serum and urine concentrations (LH: r=0.91, p<0.001; FSH: r=0.90, p<0.001). The reference intervals of uLH and uFSH were established according to age, sex, and pubertal status. We established reference intervals for uLH and uFSH based on age, sex and pubertal status to provide a non-invasive clinical screening tool for precocious puberty in children aged 6-11years.

Method Performance Verification of Anti-GAD65 and Anti-Insulin antibody Assays.

The aim of this study was to evaluate the performance of chemiluminescence immunoassays for anti-GAD65 and anti-insulin antibodies following user verification guidelines. The analytical performance of anti-GAD65 and anti-insulin antibodies using a MAGLUMI 2000 analyzer was verified following user verification guidelines by the Clinical and Laboratory Standards Institute. Performance specifications including precision, linearity, carry-over, cutoffs for positive results, reference intervals, and comparability with pre-existing commercially available radioimmunoassays using patient specimens and certified reference material were verified (coefficients of variation for precision of anti-GAD65 and anti-insulin antibodies were 2.6% and 3.4%, respectively). Comparability assessed using clinical serum specimens showed overall agreement with radioimmunoassay of 87.2% (95% confidence interval 74.8% - 94.0%) for the anti-GAD65 antibody assay and 85.4% (95% confidence interval 71.6% - 93.1%) for the anti-insulin antibody assay. The results of this study verified the analytical performance of MAGLUMI anti-GAD65 and anti-insulin antibody assays for clinical use.

Method performance verification for the combined detection and quantitation of the marine neurotoxins cyclic imines and brevetoxin shellfish metabolites in mussels (Mytilus edulis) and oysters (Crassostrea gigas) by UHPLC-MS/MS

A single laboratory method performance verification is reported for a rapid sensitive UHPLC-MS/MS method for the quantification of eight cyclic imine and two brevetoxin analogues in two bivalve shellfish matrices: mussel (Mytilus edulis) and Pacific oyster (Crassostrea gigas). Targeted cyclic imine analogues were from the spirolide, gymnodimine and pinnatoxin groups, namely 20-Me-SPX-C, 13-desMe-SPX-C, 13,19-didesMe-SPX-C, GYM-A, 12-Me-GYM, PnTx-E, PnTx-F and PnTx-G. Brevetoxin analogues consisted of the shellfish metabolites BTX-B5 and S-desoxy-BTX-B2. A rapid dispersive extraction was used as well as a fast six-minute UHPLC-MS/MS analysis. Mobile phase prepared using ammonium fluoride and methanol was optimised for both chromatographic separation and MS/MS response to suit all analytes. Method performance verification checks for both matrices were carried out. Matrix influence was acceptable for the majority of analogues with the MS response for all analogues being linear across an appropriate range of concentrations. In terms of limits of detection and quantitation the method was shown to be highly sensitive when compared with other methods. Acceptable recoveries were found with most analogues, with laboratory precision in terms of intra- and inter-batch precision deemed appropriate. The method was applied to environmental shellfish samples with results showing low concentrations of cyclic imines to be present. The method is fast and highly sensitive for the detection and quantification of all targeted analogues, in both mussel and oyster matrices. Consequently, the method has been shown to provide a useful tool for simultaneous monitoring for the presence or future emergence of these two toxin groups in shellfish.

Analytical Method Lifecycle Management in Pharmaceutical Industry: a Review.

The adoption of Quality by Design (QbD) and Analytical Method Lifecycle Management (AMLM) concepts to ensure the quality of pharmaceutical products has been applied and proposed over the last few years. These concepts are based on knowledge gained from the application of scientific and quality risk management approaches, throughout method lifecycle to assure continuous improvement and high reliability of analytical results. The overall AMLM starts with the definition of the method's intended use through the Analytical Target Profile definition, including three stages: (1) Method Design, taking advantage of the well-known concept of QbD; (2) Method Performance Qualification; (3) Continued Method Performance Verification. This is intended to holistically align method variability with product requirements, increasing confidence in the data generated, a regulatory requirement that the pharmaceutical industry must follow. This approach views all method-related activities, such as development, validation, transfer, and routine use as a continuum and interrelated process, where knowledge and risk management are the key enablers. An increase in method robustness, cost reduction, and decreased risk failures are some of the intrinsic benefits from this lifecycle management. This approach is clearly acknowledged both by regulators and industry. The roadmap of the regulatory and industry events that mark the evolution of these concepts helps to capture the current and future expectation of the pharmaceutical framework.

Statistically Designed, Targeted Profile UPLC Method Development for Assay and Purity of Haloperidol in Haloperidol Drug Substance and Haloperidol 1 mg Tablets

In view of the US pharmacopeia (USP)-proposed general chapter $$\left\langle {1220} \right\rangle$$ “The Analytical Procedure Lifecycle” and ICH-proposed ICHQ14 guideline “Analytical Procedure Development”, application of quality by design approach during analytical method development gains a significant importance. Methods developed by QbD approach will be more robust and thereby decrease efforts for method performance verification and post approval changes. Additionally, these methods will be more QC friendly as out of trend or out of specification results due to incompetent methods will be reduced and it will support the approach of life cycle management for analytical procedures. The present study deals with the development of an analytical targeted profile (ATP)-based, rapid, specific, precise, and mass-compatible RP-UPLC method with proven design space (DS) for the quantitative determination of Haloperidol and its degradation products in Haloperidol drug substance and Haloperidol 1 mg tablets. Using 5 different column chemistries, 4 different pH ranges, oven temperatures, and flow rates as variables experiments were carried out and statistically evaluated to finalize the chromatographic conditions. The significance and interaction effects of variables on the critical attributes (tailing factor and resolution) were evaluated through statistical analysis like analysis of variance (ANOVA). DOE study was also applied for the sample preparation to prove robustness of the sample preparation procedure. Forced degradation study was carried to demonstrate the stability-indicating nature of the method. Validation for active and available impurities was performed as per ICH and ANVISA guidelines and the method was found to be linear, accurate, specific, precise, and robust. Specificity of the method was confirmed orthogonally using QDa (mass detector) along with PDA detector. The method can be employed for release and stability testing of Haloperidol drug substance as well as Haloperidol 1 mg tablets.

Method performance verification of Alinity c and i systems in Karachi, Pakistan

Method performance verification of Alinity c and i systems in Karachi, Pakistan

A practical guide to validation and verification of analytical methods in the clinical laboratory.

Although the measures to improve quality in the clinical laboratory have been enormous in the past years, not least of all due to the introduction of the ISO standards 15189 and 17025, the handling of validation and verification of method performance often still differs widely from laboratory to laboratory. Much of what is published on the topic contains complex statistics and is difficult to implement in routine laboratories. The result is, that this point is often neglected or implemented incorrectly, which in turn can lead to false conclusions about method performances, potentially compromising patient safety or contributing to incorrect diagnoses. As it has long become a standard requirement for accredited laboratories to evaluate and document the analytical performance of all methods not only prior to their first implementation, but also during ongoing operation, there is a need for clear, standardized and practical guidelines on the subject. This review summarizes the current literature on the topic, focusing on the requirements for method validations, or as the case may be, verifications and describes when to validate, when to verify and which statistical tests are appropriate for each. Proper interpretation of statistical test results and acceptance criteria for each procedure are alluded to. Specific topics, which are addressed, are precision and bias verification of quantitative, qualitative and semi-quantitative procedures, method comparisons with Bland-Altman Plots, Passing-Bablok regression analysis, 2×2 contingency tables and bubble charts, linearity studies, analytical sensitivity and specificity, performing carry-over studies and establishing and confirming reference ranges.

Performance Characteristics of AOAC Method 2005.06 for the Determination of Paralytic Shellfish Toxins in Manila Clams, European Otter Clams, Grooved Carpet Shell Clams, Surf Clams, and Processed King Scallops.

An approach was developed for the verification of method performance of the AOAC 2005.06 LC-fluorescence detector (FLD) method for determination of paralytic shellfish poisoning (PSP) toxins in bivalve shellfish. This was developed following advice published by the Analytical Laboratory Accreditation Criteria Committee and applied to shellfish species that had not been previously subjected to a full single-laboratory validation scheme. The refined approach was developed following the need to assess performance in a number of shellfish species infrequently monitored through the UK statutory monitoring program, while reducing the impact and cost of the studies, most notably in terms of the use of valuable reference standards. The species assessed were manila clams (Ruditapes philippinarum), European otter clams (Lutraria lutraria), grooved carpet shell clams (R. decussatus), surf clams (Spisula solida), and king scallops (Pecten maximus) presented as adductor only or adductor plus roe. The method was assessed for sensitivity in terms of LOD and LOQ, toxin recovery, and method precision in each species. It incorporated the PSP toxins deemed toxic and/or prevalent in UK samples and commercially available as certified reference standards. The toxins studied included GTX1-5, dcSTX, STX, C1&2, and NEO. The toxins dcGTX2&3 were included for surf clams due to the prevalence of these toxins in this species as a result of toxin decarbamoylation. Method performance targets were met for each of the characteristics investigated. Consequently, the method was deemed fit for purpose for the screening and quantification of these clam and scallop species for PSP toxins by AOAC Method 2005.06 LC-FLD.

Dampfsterilisation Folge 5: Prozessvalidierung – Definition, Gesetze und Normen, Durchführung der Validierung

Although the measures to improve quality in the clinical laboratory have been enormous in the past years, not least of all due to the introduction of the ISO standards 15189 and 17025, the handling of validation and verification of method performance often still differs widely from laboratory to laboratory. Much of what is published on the topic contains complex statistics and is difficult to implement in routine laboratories. The result is, that this point is often neglected or implemented incorrectly, which in turn can lead to false conclusions about method performances, potentially compromising patient safety or contributing to incorrect diagnoses. As it has long become a standard requirement for accredited laboratories to evaluate and document the analytical performance of all methods not only prior to their first implementation, but also during ongoing operation, there is a need for clear, standardized and practical guidelines on the subject. This review summarizes the current literature on the topic, focusing on the requirements for method validations, or as the case may be, verifications and describes when to validate, when to verify and which statistical tests are appropriate for each. Proper interpretation of statistical test results and acceptance criteria for each procedure are alluded to. Specific topics, which are addressed, are precision and bias verification of quantitative, qualitative and semi-quantitative procedures, method comparisons with Bland-Altman Plots, Passing-Bablok regression analysis, 2 × 2 contingency tables and bubble charts, linearity studies, analytical sensitivity and specificity, performing carry-over studies and establishing and confirming reference ranges.

Developing new reference materials for effective veterinary drug-residue testing in food-producing animals

Reliable analysis of veterinary drug residues in animal-derived foodstuffs represents an important measure to ensure consumer protection. European legislation addresses this issue (e.g., by defining maximum residue limits, specifying sampling and monitoring plans, and stipulating performance and validation criteria for analytical methods). Certified reference materials (CRMs) constitute an important tool for method validation and method-performance verification. This paper addresses the current status of legislation, describes characteristics of analytical methods in veterinary drug-residue testing and then focuses on the challenges and considerations in developing new RMs in this field.

CLIA Final Rules for Quality Systems: Quality Assessment Issues and Answers. James O. Westgard, Sharon S. Ehrmeyer, and Teresa P. Darcy. Madison, WI: Westgard QC, Inc., 2004, 225 pp., softcover, $60.00 ($50.00 AACC members). ISBN 1-886958-20-3.

Laboratories in the United States were first regulated at the federal level in 1967 under the Medicare Act and the Clinical Laboratory Improvement Act of 1967 (CLIA’67). Government agencies have modified the regulations to these statutes in an attempt to adapt to changes in testing sites (increase in near-patient testing), technologic advances, and evolving approaches to maintaining quality systems. The last revision, published in the Federal Register on January 24, 2003, does not change the scope or intent for achieving quality in laboratories, but it does contain important changes in the prescriptive aspects of the regulation, e.g., verification of method performance and frequency of performing quality control (QC). The authors provide a practical, informative roadmap for interpretation and insight from their long experiences with CLIA. There are also helpful discussions of where College of American Pathologists (CAP) accreditation and CLIA requirements might differ. References to source material, such as the CMS State Operations Manual …