Martin H Kroll, Christopher R McCudden Endogenous Interferences in Clinical Laboratory Tests Patient Safety Edited by Oswald Sonntag and Mario Plebani Volume Martin H Kroll, Christopher R McCudden Endogenous Interferences in Clinical Laboratory Tests Icteric, Lipemic and Turbid Samples Authors Prof Martin H Kroll, MD Quest Diagnostics Giralda Farms Madison, NJ 07940 Park USA E-mail: martinkroll500@gmail.com Christopher R McCudden, MD University of Ottawa Faculty of Medicine Department of Pathology & Laboratory Medicine Ottawa, Ontario Canada E-mail: cmccudde@uottawa.ca The book has 28 figures and 19 tables ISBN 978-3-11-026620-7 e-ISBN 978-3-11-026622-1 Library of Congress Cataloging-in-Publication Data A CIP catalog record for this book has been applied for at the Library of Congress Bibliographic information published by the Deutsche Nationalbibliothek The Deutsche Nationalbibliothek lists this publicatioin in the Deutsche Nationalbibliografie; detailed bibliographic data are available in the Internet at http://dnb.dnb.de © 2013 Walter de Gruyter GmbH, Berlin/Boston The publisher, together with the authors and editors, has taken great pains to ensure that all information presented in this work (programs, applications, amounts, dosages, etc.) reflects the standard of knowledge and the time of publication Despite careful manuscript preparation and proof correction, errors can nevertheless occur Authors, editors and publisher disclaim all respondibility and for any errors or omissions or liability for the results obtained from use of the information, or parts thereof, contained in this work The citation of registered names, trade names, trade marks, etc in this work dies not imoly, even in the absence of a specific statement, that such names are exempt from laws and regulations preotecting trade marks etc and therefore free for general use Typesetting: PTP-Berlin Protago-TEX-Production GmbH, Berlin Printing and Binding: Hubert & Co GmbH & Co KG, Göttingen Cover image: Comstock/Getty Images ♾ Printed on acid-free paper Printed in Germany www.degruyter.com To my wife Ellen and children Allison, Lauren and Jonathan Martin H Kroll To my wife Liesje and children Katie and Sam Christopher R McCudden Preface Medicine has evolved to a new level, where not only is it expected that physicians diagnose and treat patients efficaciously, but also that all patients are protected from harm Protecting patients from harm is part of patient safety and implies that the processes used in taking care of patients are free from error Medical care depends on obtaining useful information from laboratory tests Biochemical tests provide a great deal of information at relatively low cost and usually with rapid turnaround times The achievement of low cost and rapid turnaround times depends, to a large extent, on the use of automation The dependence on automation subsequently results in a diminution of individualized attention to each individual sample To protect patient safety, laboratories need to establish detection systems to identify situations that could lead to biased results and rules to correct for the biased problems A bias occurs when the result obtained during an assay deviates from the true value of the analyte in question A systematic bias occurs when there is an inherent problem in the measurement technique, as can occur with calibration errors and reagent deterioration All samples are affected by a systematic bias Interferences cause a non-systematic bias Here, the bias occurs only for the individual sample It is important to identify common features that occur for interferences, and to identify ways of not only identifying the interferences, but also of quantifying their impact For biochemical tests, especially those using serum or plasma as a matrix, a high concentration of bilirubin and turbidity can affect biochemical tests The most common cause of turbidity is lipemia The intent of this book is to provide a foundation for those running laboratories to identify, quantify and correct for the presence of hyperbilirubinemia and lipemia (turbidity) Because most laboratories will need to perform these processes in an automated fashion, the people working in the laboratory will need to design the appropriate procedures and to manage them To establish the necessary foundation to effectively design processes to manage the interferences caused by bilirubin and lipemia (turbidity), this book contains several different perspectives The early chapters of the book provide information on the physical and chemical mechanisms involved in interferences There is considerable emphasis on the interaction of bilirubin and lipemic particles with light, the most common form of energy used to detect clinical biochemical species Additional chapters provide an emphasis on the clinical conditions where one might expect to encounter high concentrations of bilirubin or lipemia The latter half of the book discusses means of detecting bilirubin or lipemia, as well as means to quantify their presence, to allow for appropriate reporting of results Finally, the last chapter discusses means of characterizing and quantifying interferences in complex reactions, viii       Preface as frequently occurs with bilirubin, where the analyte may interact with the analyte or species directly related to the concentration of the analyte The intent of the book is to provide the laboratorian with sufficient background to deal with these interferences and protect patient safety November, 2012 Martin H Kroll, MD Contents Preface   vii 1.1 1.1.1 1.1.2 1.2 1.2.1 1.2.2 1.3 1.4  1 Accuracy Goals for Laboratory Tests  Accuracy and Precision   1 Definition   1 Imprecision as a Form of Error   2 Types of Error   2 Bias   2 Impact of Bias   4 Interference as a Type of Bias   6 References   8 2.1 2.2 2.3 2.4 2.5  11 Nature of Interferences  Definition   11 Nature of Interferences   11 Instrumentation   12 The Chemistry of the Absorbance of Light  References   20 3.1 3.2 3.3 3.3.1 3.3.2 3.3.3 3.3.4 3.4 3.4.1 3.4.2 3.5  21 The Nature of Icteric Interference  Source Information on Bilirubin Interference   21 Allen Correction as a Source of Bilirubin Interference   21 Bilirubin Interference with Oximetry   22 Co-oximetry Interference   24 Pulse Oximetry   25 Cerebral Oximetry   26 Interference with Methemoglobin   27 Chemical Reactions as a Cause of Bilirubin Interference   28 Bilirubin Reaction with Creatinine Methods   29 Bilirubin Reactions with Peroxidase Methods   31 References   32 4.1 4.2 4.3 4.3.1 4.3.2 4.3.3 4.3.4 The Nature of Lipemic and Turbidity Interferences  Types of Interferences   35 Lipemia Causes Turbidity   36 Lipemia Interference Mechanisms   37 Light Scattering   37 Lipoprotein Particles   40 Intralipid® and Lipemia Simulation   42 Empirical Studies in Lipemia Turbidity   43  15  35 12.2 Statistical Testing for Significance    Result = −0.0002[Bilirubin ]2 + 0.299[Bilirubin ] + 279    129 (12.4) Notice that the first term for Bilirubin is squared These results indicate that the interference mechanisms are complicated If one is not aware of the nonlinear nature of the process one might miss important details 12.2 Statistical Testing for Significance To test for statistical significance one can use a t-test using the following formula t = |b1 − 0| SE b1 , (12.5) where SE represents the standard error of the coefficient (the slope in standard linear regression) In this case the subscript ‘1’ indicates that the coefficient and the standard error are for the first term in the formula, which is because it serves as the coefficient for the concentration of the analyte, and indicates the recovery of the signal for the analytical method Usually the b1 coefficient will have a value very close to 1.00 The standard error of the coefficient is calculated when one performs the multiple regression analysis To test for bias in the recovery of the method, one would calculate assuming that the coefficient should be equal to 1.00 One would calculate t as t= |b1 − 1| SE b1 , (12.6) if the value for t, based on the number of standard deviations, is low enough not to reject the null hypothesis, e.g., t = 1.5 for degrees of freedom, then there is no bias for the method due to some sort of calibration error On the other hand, if t = 3.0 for degrees of freedom, then, for a two-tailed test (a two-tailed test implies that the values can be higher or lower), then p 0.05, then one can accept the null hypothesis and claim that there is no interference; however, if the result for the t-test yields a p 0.05, then one can accept the null hypothesis and claim that there is no interaction leading to interference; however, if the result for the t-test yields a p [...]... and nephrons in the kidney In physiologic studies, the glomerular filtration rate is assessed by injecting and measuring inulin, which provides the inulin clearance The inulin clearance is not a practical way to assess renal function Instead, clinicians use the creatinine clearance Creatinine is an endogenous compound, the breakdown product of creatine produced by skeletal muscle Its main advantage... creatinine to assess renal function is a standard practice In the past, many clinicians used the creatinine clearance to assess renal function Calculating a creatinine clearance requires the collection of a 24-hour urine sample to measure the creatinine in it The collection must be done at the same time that the serum or plasma sample is obtained for creatinine The clearance is calculated by dividing... by dividing the urine creatinine production rate by the serum or plasma creatinine concentration [8] Because there is a discrepancy in the orders of magnitude of the creatinine concentration in urine and serum (plasma) and there is some degree of error in the collection of the 24-hour urine specimen, there is considerable error in calculating a creatinine clearance Today, most clinicians depend on... Analytical interferences and analytical quality Clin Chim Acta The Netherlands, 2009,404,37–40 [6] Owen LJ, Keevil BR Does bilirubin cause interference in Roche creatinine method? Clin Chem USA 2007,53,370–371 [7] Kaplan LA, Pesce AJ Clinical Chemistry: Theory, Analysis, Correlation St Louis, MO, USA, Mosby, 2010 [8] Noe DA, Rock RC Laboratory Medicine: The Selection and Interpretation of Clinical Laboratory. .. for Clinical Laboratory Standards, 1986 Büttner J Unspecificity and interference in analytical systems: concepts and theoretical aspects DG Klin Chem Mitteilungen Germany 1991,22,3–11 Glick MR, Ryder KW, Glick SJ Interferographs User’s Guide to Interferences in Clinical Chemistry Instruments 2nd ed Indianapolis, IN, USA, Science Enterprises, Inc, 1991 Willard HH Merritt Jr LL, Dean JA, Settle Jr FA Instrumental... chemistry tests, including ALT, albumin, alkaline phosphatase, calcium, creatinine, sodium, glucose, lipase, total protein, creatine kinase, magnesium, phosphorus, urea and uric acid Because bilirubin interference varies by peer group, platform, test methodology and reagent formulation, each laboratory should evaluate the methods used in their laboratory The College of American Pathologists Interfering Substance... Medications   89  89 HIV Infection  Renal Disease   90 References   91    xi  89  93 Impact of Lipemia/Turbidity  Introduction   93 Estimated Impacts Based on Interference Studies  Interference by Light Scattering   95 Interference by Volume Displacement   96 Interference by Lipid Partitioning   99 Summary   99 References   99  95 10 Endogenous Interferences in Clinical Laboratory Tests: Icteric, Lipemic... potential interferents, such as bilirubin 20       2 Nature of Interferences 2.5 References [1] [2] [3] [4] [5] [6] [7] [8] [9] IFCC, IFCC provisional recommendation on quality control in clinical chemistry J Clin Chem Clin Biochem Germany 1976,14,270 Kroll MH, Elin RJ Interference with clinical laboratory analysis Clin Chem USA 1994,40, 1996–2005 chemistry; proposed guidelines NCCLS Document EP7-P Villanova,... from the sensor Testing and assigning values in the laboratory are separated into three phases: the pre-analytic, analytic and post-analytic phase The pre-analytic phase includes preparing the patient to obtain the specimen, collecting the specimen into an appropriate container (often with an anti-coagulant for blood), labeling and transporting the specimen to the laboratory and processing of the specimen... would have been obtained in the absence of the agent [2] Defining the term interference in this broad fashion would allow the definition to cover such agents as bilirubin, lipemia and turbidity The definition for interference can include the effect of an agent in the detection or determination of concentration or activity of an analyte occurring at any step along the path to providing a result [3] For