Quality Control Test Procedures Research Articles

Abstract 6008: Antibody validation for clinical pharmacodynamic assay reveals cross-reactivity of commercial DNMT3B clones

Abstract Cooperativity between the maintenance DNA methyltransferase, DNMT1, and de novo methyltransferases, DNMT3A and 3B, is essential for proper maintenance and integrity of the methylome, and deregulation of these enzymes and associated transcriptional silencing of multiple genes has been shown in many types of cancer. Two DNA methyltransferase inhibitors are currently in clinical use with several others undergoing clinical evaluation, and the pharmacodynamic response of DNMT enzymes is a crucial aspect of understanding therapeutic effects in preclinical studies and clinical trials. Vigorous fit-for-purpose validation of research-grade reagents and the application of effective quality control testing procedures to assess lot-to-lot variability (and reject unacceptable lots) are essential to assure robust and reproducible pharmacodynamic assay performance. During the antibody validation process for the DNMT3B enzyme target, two commercial monoclonal antibody clones were found to exhibit significant cross-reactivity to another cellular protein and inappropriate subcellular localization via immunohistochemistry (IHC), including diffuse cytoplasmic staining and significant staining in a DNMT3B knock out (KO) cell line. Two other commercial monoclonal antibody clones showed the expected nuclear expression in the HCT116 parental cell line and lack of staining in the DNMT KO cells. Further evaluation via western blot revealed that the cross-reacting antibodies detected a band of the approximate expected molecular weight (MW) of DNMT3B but of a protein that was expressed equivalently in the parental and DNMT3B KO cell line pair, whereas the two other antibody clones only detected a significant band of correct MW in the parental but not the KO cell line. These results suggest that some commercial antibody clones are binding to a protein other than DNMT3B and that they are unsuitable for use with IHC and western blot to evaluate subcellular localization and expression levels of DNMT3B protein. We presume that these non-specific antibody clones were chosen for commercial development based on target screening via Western blot that did not include evaluation of a validated DNMT3B knock out cell line, as is not uncommon for research-grade antibody clones. This data highlights the critical importance of thorough fit-for-purpose validation and quality control for crucial reagents used for pharmacodynamic biomarker studies. Funded by NCI Contract No HHSN261200800001E Citation Format: Gabriel J. Benton, Manisha Mohandoss, James H. Doroshow, Ralph E. Parchment, Katherine V. Ferry-Galow. Antibody validation for clinical pharmacodynamic assay reveals cross-reactivity of commercial DNMT3B clones. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6008.

Assessment of Diagnostic Radiology Facilities Technical Radiation Protection Requirements in KSA

The national regulatory body in the state regulates the source of ionizing radiation such as diagnostic radiology to minimize exposure to the operators and the patients. The Saudi Food and Drug Authority (KSA-SFDA) cover all medical regulatory aspects of ionizing radiation such as diagnostic X-ray facilities for all practices and intervention requirements. The study presents an assessment and analysis of the level of technical radiation protection requirements and the status of applying the national regulatory standards for different diagnostic facilities in KSA. Based on the online scientific recent database published in the field of radiation protection regulations and dose assessment for diagnostic radiology in KSA, and from the data published by the KSA-SFDA report in 2015. About 109 diagnostic X-ray facilities were selected from 35 governorates distributed in the kingdom. More than 95% of the examined facilities were in good condition concerning the national radiological protection technical regulation. About 11.9% of the facilities had a radiation leakage or cracks in the wall of the X-ray room/entrance door of the room. 16.5% of the facilities did not have a radiation warning sign written in Arabic/English languages. About 21.9% of the operators did not use any personal radiation dosimeter such as TLD or OSL. More than 40.7% of those facilities do not keep a record of the personal dosimeter reading at their facilities. Only 11.1% of the examined facilities do not have any personal protective tools such as a lead apron or thyroid shield. About 38.2% of the examined facilities do not carry out the annual periodic maintenance for the used X-ray machines at those facilities. From the obtained results, it was concluded that the majority of the radiation protection, radiation safety requirements and physical security measures undertaken in these facilities were in good implementation of the national and international technical regulations. The study suggests a need to apply for a quality control test procedures program regularly at those facilities.

Quantifying the freeze-thaw performance of air-entrained concrete using the time to reach critical saturation modelling approach

Many State Highway Agencies have been working to develop performance-based specifications for concrete pavements and concrete bridge decks in freeze-thaw environments. A time to reach critical saturation (TTRCS) model has been proposed to estimate the freeze-thaw performance of concrete. This study evaluates the TTRCS model for thirty different concrete mixtures with varying w/c, air volumes, and quality of air void (size and spacing). Simple quality control test procedures are used to determine the input parameters for the TTRCS model. The estimated time to reach critical saturation is compared with the measured durability factor using ASTM C 666–15. Results indicate that 86% of the mixtures with air volume above 4.5% and a Sequential Air Method (SAM) Number below 0.30 have a normalized time to reach critical saturation of greater than 20, and a durability factor that is greater than 75%. The mixtures with a high range water reducer require a higher volume fraction of entrained air to satisfy the recommended limit for the durability factor. This appears to be due to an interaction between the high range water reducer and air entraining admixture which results in greater air void spacing (i.e., larger air voids). However, the addition of high range water reducer was also found to increase the time to reach critical saturation in the mixtures with a low w/c due to a refined pore structure with a reduction in the connectivity of the matrix pores. Reducing the w/c improves the freeze-thaw performance due to a reduction in the pore volume, connectivity, and absorption rate of the concrete. A relationship is developed to estimate the time to reach critical saturation based on SAM Number and apparent formation factor. In addition, a relationship is proposed to estimate the critical degree of saturation based on air void content and quality.

Simultaneous quantification of the viral antigens hemagglutinin and neuraminidase in influenza vaccines by LC–MSE

Current methods for quality control of inactivated influenza vaccines prior to regulatory approval include determining the hemagglutinin (HA) content by single radial immunodiffusion (SRID), verifying neuraminidase (NA) enzymatic activity, and demonstrating that the levels of the contaminant protein ovalbumin are below a set threshold of 1μg/dose. The SRID assays require the availability of strain-specific reference HA antigens and antibodies, the production of which is a potential rate-limiting step in vaccine development and release, particularly during a pandemic. Immune responses induced by neuraminidase also contribute to protection from infection; however, the amounts of NA antigen in influenza vaccines are currently not quantified or standardized. Here, we report a method for vaccine analysis that yields simultaneous quantification of HA and NA levels much more rapidly than conventional HA quantification techniques, while providing additional valuable information on the total protein content. Enzymatically digested vaccine proteins were analyzed by LC–MSE, a mass spectrometric technology that allows absolute quantification of analytes, including the HA and NA antigens, other structural influenza proteins and chicken egg proteins associated with the manufacturing process. This method has potential application for increasing the accuracy of reference antigen standards and for validating label claims for HA content in formulated vaccines. It can also be used to monitor NA and chicken egg protein content in order to monitor manufacturing consistency. While this is a useful methodology with potential for broad application, we also discuss herein some of the inherent limitations of this approach and the care and caution that must be taken in its use as a tool for absolute protein quantification. The variations in HA, NA and chicken egg protein concentrations in the vaccines analyzed in this study are indicative of the challenges associated with the current manufacturing and quality control testing procedures.

CRS/AAPS Joint Workshop on Critical Variables in the In Vitro and In Vivo Performance of Parenteral Sustained-Release Products

The Controlled Release Society (CRS) and the American Association of Pharmaceutical Scientists (AAPS) recently co-sponsored a workshop on the critical variables impacting the in vivo performance of parenteral controlled-release formulations. This workshop was a continuation of discussions initiated during the 2007 CRS Educational Workshop entitled “In Vitro and In Vivo and Considerations Associated with Parenteral Sustained Release Products,” the outcome of which was summarized in the Journal of Controlled Release (1). The 2008 workshop focused on the use of design-space concepts as a mechanism to identify the critical quality attributes associated with in vivo product performance and to facilitate the development and use of in vitro test methods for (1) setting batch release characteristics, (2) ensuring the equivalence of product performance when there are changes in formulations, excipients, or manufacturing methods, and (3) modifying batch release specifications. To address these complex issues, the workshop format allowed experts to present their perspectives on challenges associated with assuring the quality and performance of a range of state-of-the art parenteral delivery systems and provided an opportunity to hear and discuss a range of topics critical to parenteral product performance. The ten questions that served as the focus of breakout group discussions are provided in Table 1. It is also noteworthy that this workshop was the first opportunity for CRS membership to consider these questions and provide a response electronically by logging on to the CRS website (2). Feedback from the workshop participants included the following points: • Regardless of the type of parenteral product, some methodology should be employed to ensure that it is functioning as intended before batch release. • In some cases, quality control testing procedures necessitate the use of multiple tests to demonstrate adequate function of these complex delivery systems. • Host response may not be due solely to the bioactive in the product or to its chemical components, but may be related to dosage form size, shape, age, or surface characteristics. • It is important that product development is not hindered by the process of defining test requirements and that economic and time constraints are considered when developing in vitro release specifications. • Nontraditional tests can be developed, but relevant release specifications must be set if they are developed. That said, existing USP methods are underutilized, and it would be preferable (prudent) to develop tests that rely on USP apparatus since the specifications for the physical and experimental conditions associated with the use of USP apparatus are well-defined. • The establishment of any set of product specifications needs to be an iterative process. • There could be a difference between quality control methods and a method that is used to support a biowaiver. However, if a test is developed solely for quality control purposes and has been designed specifically to monitor critical quality attributes, then it cannot be used to predict biological performance. • If the release specification is intended to ensure therapeutic activity, it will be necessary to ascertain the biological relevance of the test results and the release specifications through studies that establish the in vivo–in vitro relationship. • For more complex dosage forms, safety concerns may extend beyond the rate and extent of drug exposure. When there are additional safety or efficacy concerns that cannot be addressed with bioequivalence trials, it may not be feasible to develop generic formulations of these products (e.g., products with safety and activity profiles based on ligand-based targeted delivery systems). • When employing the design-space concept, extremes of the proposed specifications should be tested both in vitro and in vivo (where in vivo studies can include the use of animal models). The in vivo data are needed to ensure that the in vitro test is reliable and discriminating. When using design space, aged materials need

TU-D-230C-01: Digital Radiography QC Workshop I and II

Computed radiography and digital radiography systems now comprise a large segment of radiography work. It is important that appropriate performance testing and quality control (QC) tools and procedures be made available for these systems. The vendors and/or manufacturers of digital imaging systems may provide‐or make available‐test tools, software and procedures for performance evaluation and routine quality controltesting of their digital systems. Currently, such tools and programs vary significantly from manufacturer to manufacturer. This two‐part workshop will include both formal presentations and hands‐on demonstrations. During the first half of the workshop, each participating vendor or manufacturer will present a brief (12–15 minute) overview of tools and procedures for performance testing and QC of their systems. The second half of the workshop is dedicated to hands‐on demonstrations at table‐top exhibits hosted by each participating vendor. Exhibits will include test tools, workstations and samples of performance testing and QC test procedures. Companies participating in this workshop will include Agfa, Fuji, General Electric, Kodak, Konica, IDC, LoDox, Philips and potentially others. Educational Objectives: 1. To understand performance test procedures used by various DR/CR systems. 2. To see types QC test tools and procedures made available with DR/CR systems. 3. To obtain hands‐on demonstration of DR/CR ttesting and QC procedures.

TU‐C‐W‐608‐01: Ultrasound QC Workshop

This workshop will provide attendees an opportunity to observe ultrasound scanner operations and quality assurance procedures. A portable ultrasound unit and various ultrasound QC phantoms will be provided and used to demonstrate the QC procedures. A set of QC test procedures described in AAPM Ultrasound Task Group 1 Report (Medical Physics 1998;Vol 25: 1385–1406) will be presented, including display monitor fidelity, image uniformity, depth of visualization, horizontal and vertical distance accuracy, axial and lateral resolution, slice thickness, dead zone measurement. Demonstrations of test procedures and equipment will be performed by the instructors and industry representatives.Educational Objectives:1. To learn the effect of parameter settings on various aspects of ultrasound imaging.2. To observe the ultrasound quality control test procedures for real‐time B‐mode units.3. To become acquainted with various ultrasound QC phantoms.

Real-time B-mode ultrasound quality control test procedures. Report of AAPM Ultrasound Task Group No. 1.

Real-time B-mode ultrasound quality control test procedures. Report of AAPM Ultrasound Task Group No. 1.

Geomembrane seam nondestructive tests: Construction quality control (CQC) perspective

This paper is intended to present and compare the various commonly used geomembrane field seam nondestructive field testing methods from a construction quality control perspective. The testing methods addressed include: vacuum; air lance; pressure; electrical resistivity; electric spark; and mechanical point stress. The viewpoint presented is one of an installation contractor's quality control testing procedures and the associated limitations, results, data documentation and costs.

Use of glacial till and outwash silty sand as construction material for dam cores at Long Spruce Generating Station

This paper documents experience gained during the construction of the earthworks at the Long Spruce Generating Station using impervious core materials of different geologic origins. Testfills of glacial till made to compare compaction equipment are described. The results are discussed and the performance of self-propelled sheepsfoot and pneumatic 50 ton (45 Mg) rollers is compared. Problems encountered in the use of the two types of material are discussed. Problems with glacial outwash silty sand stemmed primarily from the variability of the material in the borrow pits. The till proved to be more homogeneous but certain care in excavation and conditioning was required. Construction control and quality control testing procedures are reviewed. Both nuclear and conventional density tests were utilized and the results are compared.

Buoyancy materials for deep submergence

High strengtj buoyancy materials, such as syntactic foam, are required to provide buoyancy for a wide variety of deep submergence vehicles and deep ocean installations. Syntactic foam of 22pcf net buoyancy is currently variable for these applications, together with suitable quality control testing procedures. Materials of 34pcf net buoyancy are achievable based on the development of composites such as aggregates of small hollow spheres embedded in a matrix of syntactic foam.