ABSTRACT
BACKGROUND: In vitro diagnostic bilirubin reagents based on oxidation with bilirubin oxidase or vanadic acid for total and direct-reacting bilirubin are widely used in Japan; however, their reactivity to unconjugated and conjugated bilirubin and delta bilirubin has not been completely disclosed by manufacturers. We used artificially prepared bilirubin materials to investigate the reactivity with four in vitro diagnostic bilirubin reagents. METHODS: Porcine unconjugated bilirubin solution, chemically synthesized ditaurobilirubin solution, and chemically synthesized delta bilirubin solution were used as surrogates of naturally occurring unconjugated bilirubin, conjugated bilirubin, and delta bilirubin, respectively. The total bilirubin and direct-reacting bilirubin concentrations were measured by three bilirubin oxidase methods and one vanadic acid method, and the observed concentrations were compared with those obtained by the diazo-based reference measurement procedure. RESULTS: The unconjugated bilirubin and delta bilirubin concentrations were similar when any of the four in vitro diagnostic bilirubin reagents were used during total bilirubin measurement. This was consistent with reference measurement procedure and exhibited a converged inter-method variation. Compared with reference measurement procedure, significantly low ditaurobilirubin concentrations were observed by the in vitro diagnostic bilirubin reagents despite the converged inter-method variation. In delta bilirubin measurement, some reagents reacted doubtfully with unconjugated bilirubin, while showed lower ditaurobilirubin concentrations than its corresponding total bilirubin concentration. Reactivity with delta bilirubin was different for each method including reference measurement procedure. Some reagents were developed to react less with delta bilirubin and others to strongly react with delta bilirubin. CONCLUSIONS: We revealed the reactivity of IVD-TB and IVD-DB reagents to artificially prepared bilirubin materials, and their consistency with reference measurement procedure. The delta bilirubin data results vary depending on the reagents used.
Subject(s)
Bilirubin , Taurine , Animals , Indicators and Reagents , Japan , Oxidation-Reduction , Swine , Taurine/analysisABSTRACT
INTRODUCTION: Current production facilities for Cell-Based Health care Products (CBHPs), also referred as Advanced-Therapy Medicinal Products or Regenerative Medicine Products, are still dependent on manual work performed by skilled workers. A more robust, safer and efficient manufacturing system will be necessary to meet the expected expansion of this industrial field in the future. Thus, the 'flexible Modular Platform (fMP)' was newly designed to be a true "factory" utilizing the state-of-the-art technology to replace conventional "laboratory-like" manufacturing methods. Then, we built the Tissue Factory as the first actual entity of the fMP. METHODS: The Tissue Factory was designed based on the fMP in which several automated modules are combined to perform various culture processes. Each module has a biologically sealed chamber that can be decontaminated by hydrogen peroxide. The asepticity of the processing environment was tested according to a pharmaceutical sterility method. Then, three procedures, production of multi-layered skeletal myoblast sheets, expansion of human articular chondrocytes and passage culture of human induced pluripotent stem cells, were conducted by the system to confirm its ability to manufacture CHBPs. RESULTS: Falling or adhered microorganisms were not detected either just after decontamination or during the cell culture processes. In cell culture tests, multi-layered skeletal myoblast sheets were successfully manufactured using the method optimized for automatic processing. In addition, human articular chondrocytes and human induced-pluripotent stem cells could be propagated through three passages by the system at a yield comparable to manual operations. CONCLUSIONS: The Tissue Factory, based on the fMP, successfully reproduced three tentative manufacturing processes of CBHPs without any microbial contamination. The platform will improve the manufacturability in terms of lower production cost, improved quality variance and reduced contamination risks. Moreover, its flexibility has the potential to adapt to the modern challenges in the business environment including employment issues, low operational rates, and relocation of facilities. The fMP is expected to become the standard design basis of future manufacturing facilities for CBHPs.
ABSTRACT
BACKGROUND: The diagnosis of myocardial infraction (MI) in patients presenting to the emergency department represents a clinical challenge. It is known that creatine kinase-MB isoenzyme (CK-MB) is present in soluble cell fractions of cardiac muscle, and injury to those cells results in an increase of CK-MB in the blood. Therefore, CK-MB is a suitable clinical biomarker of myocardial infraction. METHODS: To measure CK-MB mass rapidly and easily, we developed the new reagent 'L-type Wako CK-MB mass' (L-CK-MB mass) for the latex agglutination turbidimetric immunoassay method. Using a Hitachi LABOSPECT 008, we evaluated the performance of this assay as a method for quantifying CK-MB mass, and we compared the measurement of the serum CK-MB mass concentration with this assay to that obtained using an electrochemiluminescence immunoassay (ECLIA). RESULTS: A dilution test showed linearity from 5 µg/L to 190 µg/L, and the limit of quantification of the L-CK-MB mass assay was 3.0 µg/L. The within-run CV and between-day CV were 1.0 - 4.5% and 1.8 - 4.4%, respectively. Serum CK-MB mass concentration determined using the L-CK-MB mass assay was reliably and strongly correlated with that determined using ECLIA (n = 163, r = 0.999, y = 0.977x + 0.307). CONCLUSIONS: The L-CK-MB mass assay is able to specifically determine CK-MB mass and is a very useful method for the accurate measurement of CK-MB mass for routine clinical analyses.
