<p> SMILE Johns Hopkins University Baltimore, MD USA</p><p>SMILE Linearity Requirements-Chemistry</p><p>Author: Validation Committee Document Number: Equ35-A-05</p><p>Effective (or Post) 23-Mar-11 Date:</p><p>Review History Date of last review: NA</p><p>Reviewed by: Heidi Hanes</p><p>SMILE Comments: This document is provided as an example only. It must be revised to accurately reflect your lab’s specific processes and/or specific protocol requirements. Users are directed to countercheck facts when considering their use in other applications. If you have any questions contact your SMILE representative.</p><p>SMILE Linearity Requirements-Chemistry Document Number 230</p><p>Effective Date 19 Dec 2008</p><p>Subject Page 1 of 1</p><p>Guidelines for performing linearity testing on a chemistry analyzer Supersedes New</p><p>Author(s) Name, Title Date Mark Swartz, Heidi Hanes, Jo Shim, Penny Dec 2008 Stevens, Anne Sholander Approved by Name, Title Date SMILE Validation Committee Dec 2008</p><p>Review History Date of last review: 11 Mar 2011</p><p>Reviewed by: Anne Sholander</p><p>Version # [0.0] Revision Date Description (notes) [dd/mm/yy] Revision 1.1 03/11/11 Updated to acceptability criteria to remove reference to “pass or fail”. History</p><p>08991e838589556df5868a855ca597ce.doc Page 1 of 4 SMILE Johns Hopkins University Baltimore, MD USA</p><p>SMILE Chemistry Linearity Guidelines</p><p>A quantitative analytical method is said to be LINEAR when measured results from a series of sample solutions are directly proportional to the concentration or activity in the test specimens. This means that a straight line can be used to characterize the relationship between measured results and the concentrations or activity levels of an analyte for some stated range of analyte values.</p><p>I. Linearity</p><p>A. Sample Criteria</p><p>1. A minimum of 5 samples that cover the reportable range of the method.</p><p>2. When plotted, the values should ideally be equidistant from each other.</p><p>3. Quality control, calibrators or commercial linearity standards should be used.</p><p>B. Testing </p><p>1. At a minimum, run each sample in duplicate.</p><p>2. Data should be plotted immediately to identify and correct any outliers.</p><p>C. Evaluation of data:</p><p>1. Plot the data in regression analysis program. The SMILE Linearity spreadsheet (Appendix 1) performs all necessary calculations. </p><p> a. Plot the known values of the standards on the X-axis</p><p> b. Plot the mean of the measured values on the Y-axis.</p><p> c. Calculate slope and intercept using linear regression. This can be done using the Excel SLOPE and INTERCEPT functions or from the website http://tools.westgard.com/cgi- bin/westgard/comp_calc.cgi? header=http:/www.westgard.com/images/headtoolkit.gif</p><p> d. Using slope and intercept, calculate a predicted Y value for each X value. </p><p>08991e838589556df5868a855ca597ce.doc Page 2 of 4 SMILE Johns Hopkins University Baltimore, MD USA</p><p> e. Plot the predicted Y values versus the corresponding known X values on the same graph mentioned in a and b above. Draw a straight line to connect all the predicted Y points on the graph.</p><p> f. Subtract each measured Y value from the associated predicted Y value. This difference is the systematic error due to non-linearity. </p><p> g. Compare that systematic error to 50% of the total error. The systematic error must be less than 50% of the total error.</p><p>2. Alternatively, plot the data in EP Evaluator Linearity Module or similar regression analysis program. If using EP follow the steps below. (See Appendix 2 for an example EP Linearity Report).</p><p> a. Under the Linearity parameters select Confirm Linearity.</p><p> b. Enter the Allowable Total Error (TEa) concentration and percent. Refer to SMILE Chemistry TE Limits table (Appendix 3).</p><p> c. Enter 50% for Systematic Error.</p><p> d. Select edit and enter the assigned values of your standards.</p><p> e. Enter each replicate of your test data.</p><p>D. Acceptability criteria</p><p>The method is linear if the difference between the predicted Y value and the measured Y value is less than the allowable error for each specimen point.</p><p>II. References</p><p>A. GCLP Workshop and Workbook18-20 May 2008, Verification of Performance Specifications, pages 1-33.</p><p>B. Clinical and Laboratory Standards Institute (CLSI). Evaluation of the Linearity of Quantitative Measurement Approved Guideline-Second Edition, CLSI document EP6-A (ISBN 1-56238-498-8) Clinical and Laboratory Standards Institutes, 940 West Valley Road, Suite 100, Wayne, Pennsylvania 19098- 1898 USA, 2005.</p><p>C. EP Evaluator Release 8, David G. Rhoads Associates Inc., www.dgrhoads.com.</p><p>08991e838589556df5868a855ca597ce.doc Page 3 of 4 SMILE Johns Hopkins University Baltimore, MD USA</p><p>D. James O. Westgard, Online Validation Training, Westgard QC, Inc. www.westgard.com, Sections 9-Determining Reportable Range.</p><p>08991e838589556df5868a855ca597ce.doc Page 4 of 4 </p>