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      Steroid Assays in Paediatric Endocrinology

      review-article
      1 ,
      Journal of Clinical Research in Pediatric Endocrinology
      Galenos Publishing
      Steroid, adrenal, adrenal tests assay

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          Abstract

          Most steroid disorders of the adrenal cortex come to clinical attention in childhood and in order to investigate these problems, there are many challenges to the laboratory which need to be appreciated to a certain extent by clinicians. The analysis of sex steroids in biological fluids from neonates, over adrenarche and puberty present challenges of specificities and concentrations often in small sample sizes. Different reference ranges are also needed for interpretations. For around 40 years, quantitative assays for the steroids and their regulatory peptide hormones have been possible using immunoassay techniques. Problems are recognised and this review aims to summarise the benefits and failings of immunoassays and introduce where tandem mass spectrometry is anticipated to meet the clinical needs for steroid analysis in paediatric endocrine investigations. It is important to keep a dialogue between clinicians and the laboratory, especially when any laboratory result does not make sense in the clinical investigation.

          Conflict of interest:None declared.

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          Most cited references185

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          Strategies for the assessment of matrix effect in quantitative bioanalytical methods based on HPLC-MS/MS.

          In recent years, high-performance liquid chromatography (HPLC) with tandem mass spectrometric (MS/MS) detection has been demonstrated to be a powerful technique for the quantitative determination of drugs and metabolites in biological fluids. However, the common and early perception that utilization of HPLC-MS/MS practically guarantees selectivity is being challenged by a number of reported examples of lack of selectivity due to ion suppression or enhancement caused by the sample matrix and interferences from metabolites. In light of these serious method liabilities, questions about how to develop and validate reliable HPLC-MS/MS methods, especially for supporting long-term human pharmacokinetic studies, are being raised. The central issue is what experiments, in addition to the validation data usually provided for the conventional bioanalytical methods, need to be conducted to confirm HPLC-MS/MS assay selectivity and reliability. The current regulatory requirements include the need for the assessment and elimination of the matrix effect in the bioanalytical methods, but the experimental procedures necessary to assess the matrix effect are not detailed. Practical, experimental approaches for studying, identifying, and eliminating the effect of matrix on the results of quantitative analyses by HPLC-MS/MS are described in this paper. Using as an example a set of validation experiments performed for one of our investigational new drug candidates, the concepts of the quantitative assessment of the "absolute" versus "relative" matrix effect are introduced. In addition, experiments for the determination of, the "true" recovery of analytes using HPLC-MS/MS are described eliminating the uncertainty about the effect of matrix on the determination of this commonly measured method parameter. Determination of the matrix effect allows the assessment of the reliability and selectivity of an existing HPLC-MS/MS method. If the results of these studies are not satisfactory, the parameters determined may provide a guide to what changes in the method need to be made to improve assay selectivity. In addition, a direct comparison of the extent of the matrix effect using two different interfaces (a heated nebulizer, HN, and ion spray, ISP) under otherwise the same sample preparation and chromatographic conditions was made. It was demonstrated that, for the investigational drug under study, the matrix effect was clearly observed when ISP interface was utilized but it was absent when the HN interface was employed.
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            Position statement: Utility, limitations, and pitfalls in measuring testosterone: an Endocrine Society position statement.

            The objective of the study was to evaluate the current state of clinical assays for total and free testosterone. The five participants were appointed by the Council of The Endocrine Society and charged with attaining the objective using published data and expert opinion. Data were gleaned from published sources via online databases (principally PubMed, Ovid MEDLINE, Google Scholar), the College of American Pathologists, and the clinical and laboratory experiences of the participants. The statement was an effort of the committee and was reviewed in detail by each member. The Council of The Endocrine Society reviewed a late draft and made specific recommendations. Laboratory proficiency testing should be based on the ability to measure accurately and precisely samples containing known concentrations of testosterone, not only on agreement with others using the same method. When such standardization is in place, normative values for total and free testosterone should be established for both genders and children, taking into account the many variables that influence serum testosterone concentration.
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              Interferences and contaminants encountered in modern mass spectrometry.

              With the invention of electrospray ionization and matrix-assisted laser desorption/ionization, scientists employing modern mass spectrometry naturally face new challenges with respect to background interferences and contaminants that might not play a significant role in traditional or other analytical techniques. Efforts to continuously minimize sample volumes and measurable concentrations increase the need to understand where these interferences come from, how they can be identified, and if they can be eliminated. Knowledge of identity enables their use as internal calibrants for accurate mass measurements. This review/tutorial summarizes current literature on reported contaminants and introduces a number of novel interferences that have been observed and identified in our laboratories over the past decade. These include both compounds of proteinaceous and non-proteinaceous nature. In the supplemental data a spreadsheet is provided that contains a searchable ion list of all compounds identified to date.
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                Author and article information

                Journal
                J Clin Res Pediatr Endocrinol
                JCRPE
                Journal of Clinical Research in Pediatric Endocrinology
                Galenos Publishing
                1308-5727
                1308-5735
                March 2010
                8 December 2010
                : 2
                : 1
                : 1-16
                Affiliations
                [1 ] University College London Hospitals, London, England
                +44 771 090 1800 john.honour@ 123456uclh.nhs.uk Clinical Biochemistry, University College London Hospitals, London, W1T 4EU, England
                Article
                45
                10.4274/jcrpe.v2i1.1
                3005666
                21274330
                1c4d9208-eb86-42ca-a3eb-073321ff8b9f
                © Journal of Clinical Research in Pediatric Endocrinology, Published by Galenos Publishing.

                This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 3 February 2010
                : 25 February 2010
                Categories
                Review

                Pediatrics
                adrenal,steroid,adrenal tests assay
                Pediatrics
                adrenal, steroid, adrenal tests assay

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