Proteomics and Protein Markers
Editorials and Reviews are usually invited, although unsolicited manuscripts in these categories are welcome.
Reviews of Books, Software, and Web Sites inform our readers of resources relevant to their practice. They are intended to provide an evaluation of quality, contribution to the field, and relevance to other resources. Reviews are customarily commissioned; persons interested in writing reviews should contact the Book Reviews Editor. The journal does not underwrite the opinions expressed by its reviewers and does not publish rebuttals to reviews. Authors and publishers who wish to correct what they perceive as an error of fact in a published review should contact the Journal.
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Animal studies. Authors are responsible for ensuring that studies of animals are in accordance with the Guide for the Care and Use of Laboratory Animals (2).
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Use wide (2 cm) margins and double spacing throughout the abstract, text, references, tables, figure legends, and footnotes. Place references, table, and figure legends on separate pages, in that order. Do not use headers or footers, but include page numbering. For guidance on manuscript preparation and style, consult the CBE Style Manual (3) or, as needed, The ACS Style Guide: A Manual for Authors and Editors (4). Use the International System of Units (SI) (5) consistently throughout. Values expressed in conventional units may be added in parentheses after the value in SI units. Space in the journal is limited, and authors are expected to present their work concisely. Special limits apply to Technical Briefs and Letters to the Editor (see Journal categories). Additional data, images, or method details may be submitted for archival electronic posting with the published paper. (See “Data Supplements” below.)
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Abbreviations. Define all nonstandard abbreviations the first time they are used in the text and (if more than two) in a single footnote after the abstract. Avoid unnecessary new abbreviations. For terms used fewer than four times in a report, do not introduce an abbreviation. A list of abbreviations that may be used without definitions is available with the Information for Authors at http://www.aacc.org/ccj/authors.stm.
Materials and methods. Identify the apparatus used and provide the name of the manufacturer and the model number if multiple versions exist. For modified or newly created equipment, provide enough detail for the reader to duplicate the construction. Describe the roles, if any, of funding organizations in (a) the design of the study, (b) the data collection, analysis and interpretation, and (c) the preparation of the manuscript, and their rights to approve, delay, or disapprove of publication of the work.
Acknowledgments. In a paragraph preceding the references, acknowledge financial support, gifts, technical help, or other assistance, including that rendered by a company in evaluation of a product. (See the Conflict of interest section above.)
Figures. Acceptable image file formats for print publication are TIFF and EPS, at resolutions dictated by our print publication vendor at http://cpc.cadmus.com/da/guidelines.asp, and must be submitted as independent files, not embedded within a word processing document Microsoft PowerPoint (.PPT) files are also acceptable, but the graphics contained within them must also meet the vendor’s resolution requirements. GIF, JPEG, PDF, and embedded figures may be used for initial submission and review; however, authors are advised to create and keep print-quality files on hand, as they will be required upon first revision of the manuscript. In the case that publication quality electronic figure files cannot be provided, acceptable publication prints are: for halftones, glossy prints; for line drawings, glossy prints or laser prints on coated (nonabsorbent) laser-printer paper or drawings in black India ink on tracing paper. Figures printed with dot-matrix printers are not acceptable. Figures, electronic or otherwise, should be redesigned or recreated if they do not appear sharp and clear on paper. Authors are advised to test print electronic files before submitting them for publication. Verify that symbols and lettering will be legible when the figure is reduced to one column (85 mm) or two columns (176 mm) wide. Letters should be 8\10 points when reduced, subscripts at least 6 points. Use a medium (not bold) sans serif (Swiss, Helvetica, Arial) font. Lowercase lettering with an initial capital is preferred. Use abbreviations appropriate for journal style (e.g., mL and mmol/L, not ml and mM). Use a period (full stop), not a comma, to indicate decimal position.
Color illustrations will be considered for publication. The author will be required to bear the full cost of their preparation and publication, invited contributions excepted. The charge for the first color figure is $1500. Subsequent color figures or parts of figures are $500 each. Authors may contact the Editor to discuss undue hardship. Every effort is made to ensure that no meritorious manuscript is declined for lack of funds. Color versions of figures may be provided for publication in the on-line Journal at no extra charge, with the figures produced in black and white in the print journal.
Provide figure captions and subcaptions on a separate page. If a figure has been published previously, acknowledge the original source and submit written permission from the copyright holder to reproduce the material.
Tables. Place Tables after figure captions. Most common word processing formats are acceptable. Spreadsheet generated tables should be placed in a word-processing document. Graphic formats in the above Figures section are also acceptable. Type each table and its brief caption on its own page. Indicate explanatory footnotes with superscript lowercase italic letters in alphabetical order, reading across the table. Number tables consecutively with Arabic numerals. Give every column a heading with clearly defined units as appropriate.
References. Number references in the order of appearance and list them in numerical order in a section titled "References" placed after the text and any acknowledgements. Do not alphabetize. Numbers for reference citations in the text should be typed on the line, in parentheses, and underlined; our typesetting program will convert them into the style in which they will appear in published text. In the reference list, name all authors of a paper unless there are more than seven, in which case list the first six plus "et al." Indicate any references that are Editorials, Abstracts, Letters, Technical Briefs, or Reviews. Abbreviations for journal names are those used by the National Library of Medicine; for journals not listed there, follow Chemical Abstracts or Biological Abstracts. Do not use italic or boldface type in the reference citations. The on-line submission and reviewing system uses a reference parser for displaying references in hyperlinked HTML format as an aid for reviewers and editors. This also provides a check on the format and accuracy of reference citations. Correct linking of references depends on strict adherence to Journal style. Label the reference section with "References" and make sure that your manuscript does not contain headers, or footers. Journal abbreviations found at http://www.ncbi.nlm.nih.gov/entrez/journals/noprov/loftext_full_noprov.html will yield the best results. Examples of reference format are as follows:
- Fiechtner M, Ramp J, England B, Knudson MA, Little RR, England JD, et al. Affinity binding assay of glycohemoglobin by two-dimensional centrifugation referenced to hemoglobin Alc. Clin Chem 1992;38:2372–9.
- Demers LM. New biochemical marker for bone disease: is it a breakthrough? [Editorial]. Clin Chem 1992;38:2169–70.
- Davey L, Naidoo L. Urinary screen for acetaminophen (paracetamol) in the presence of N-acetylcysteine [Letter]. Clin Chem 1993;39:2348–9.
Books and monographs (except serial volumes, which are treated as journals)
- Siminovitch KA. Molecular characterization of human anti-DNA antibodies. In: Farid NR, Bona CA, eds. The molecular aspects of autoimmunity. San Diego: Academic Press, 1991:59–72.
- Bailar JC III, Mosteller F, eds. Medical uses of statistics, 2nd ed. Boston: NEJM Books, 1992:449pp.
- Harley JB, Gaither KK. Autoantibodies. In: Klippel JH, ed. Systemic lupus erythematosus. Rheumatic disease clinics of North America, Vol. 14. Philadelphia: WB Saunders, 1988:43–56.
- Haughton MA. Immunonephelometric measurement of vitamin D binding protein [MAppSci thesis]. Sydney, Australia: University of Technology, 1989:87pp.
Internet sources should include the author (if any), the title of the site, the URL (address), and the date accessed.
- American Association for Clinical Chemistry. AACC continuing education. http://www.aacc.org/services/default.stm (Accessed November 1998).
Reference to unpublished work. Authors citing manuscripts in press, personal communications, or other unpublished work related to the manuscript must submit with their manuscript a letter from the individuals concerned, confirming the work and giving permission for the citation. If any of the authors’ unpublished manuscripts submitted to other journals are cited in the manuscript submitted to Clinical Chemistry or may be critical to its evaluation, a copy of each unpublished manuscript must be included with the original submission. Personal communications, unpublished work, and manuscripts that have not been accepted must be cited parenthetically in the text and not as numbered references.
Data supplements. Supplemental data too large for print publication or exceeding the bounds of the manuscript type may be submitted with a manuscript for inclusion in the on-line version of Clinical Chemistry. Files intended for online publication only should be uploaded to the submission system at http://submit.clinchem.org as type "Supplemental Files", and referred to as online supplements in the text. Such supplemental information will be reviewed as part of the manuscript and will be evaluated for its importance and relevance. If accepted, the supplemental data will be referenced in the text of the article, directing readers to the URL of the Web site.
DESCRIPTION OF ANALYTICAL METHODS AND RESULTS
Manuscripts describing the development and evaluation of the performance of methods and instruments should discuss linearity, imprecision, analytical specificity, recovery, lower limit of detection, comparability with other analytical methods, lower limit of quantification and reference interval(s); some clinical data are usually needed. Document the analytical advantages of the new or modified method over existing methods.
Calibration curves and linearity. Data for these studies should be subjected to linear regression analysis (if a linear response is obtained) and should include the slope, intercept, standard error of estimate (standard deviation about the regression line), and the standard deviations of the slope and intercept. Standard deviations of repeated points may be included. In preparing radioimmunoassay calibration curves, authors may use any objective, statistically valid method but must specify the method used [see, e.g., Ref. (6)].
Imprecision. Studies must include estimates of "within-run" and "total" standard deviations. Each should be determined at low, normal, and above-normal concentrations with use of specimens that are in an appropriate biological matrix. One method of estimating both within-run and total standard deviations is the analysis of variance experiment described in NCCLS EP5-T (7), which calls for two replicates per specimen per run and two runs per day for 20 days. This permits separate estimation of between-day and between-run, within-day standard deviations, as well as within-run and total standard deviations. For acceptable alternatives that include only one run per day, see the cited document.
Indicators of Accuracy ("Trueness"). (a) Analytical recovery studiesinvolve analyses after known amounts of analyte are added to the biological fluid on which the determination will be performed. Recovery of added analyte should be calculated. (b) Interference studies should be performed to assess the effects of common interferents, e.g., lipids, hemoglobin, bilirubin, and components of uremic plasma. Exogenous materials, such as commonly used or commonly coadministered drugs that might interfere with the determination, should also be tested for interferences. (c) Comparison-of-methods studies should compare results by the new or proposed method with those by a reference-quality method or other generally accepted analytical method for which assay performance is documented (8,9). It is desirable to test 100 to 200 different samples from patients who have been selected to include a wide variety of pathologic conditions and to present a range of values for the analyte that includes those likely to be encountered in routine application. For a table of the required number of samples, see Linnet (10). If regression analysis is used for statistical evaluation of the data, authors must supply slopes and intercepts (and their standard deviations) and standard deviations of residuals (Sy|x, often called standard errors of estimates). Unbiased (e.g., Deming) regression is typically required (11). A program to perform Deming regression is available on-line as a supplement from this journal (12). The correlation coefficient has limited utility. Residuals plots [e.g., Bland-Altman (13)] are often useful. (d) Analyses of reference materials are (along with comparisons with a Reference Method) indicators of accuracy (trueness).
Analytical sensitivity and detection limit. These terms are commonly confused. The International Union of Pure and Applied Chemistry defines analytical sensitivity as the ability of an analytical procedure to produce a change in signal for a defined change of the quantity (e.g., the slope of the calibration curve). Detection limit (or limit of detection) is defined as the lowest concentration or quantity of an analyte that can be detected with a stated reasonable uncertainty for a given analytical procedure. The operational definition of this limit must be supplied by the author: e.g., the concentration at a signal-to-noise ratio of 10 or the concentration corresponding to a signal 3 SD above the mean for a calibrator that is free of analyte.
Analytical quality. Results obtained for the performance characteristics should be compared objectively with well-documented quality specifications, e.g., published data on the state of the art, performance required by regulatory bodies such as CLIA ¢88, or recommendations documented by expert professional groups (14).
Reference interval (normal range). Depending on the conclusions of the accuracy studies, modification of an accepted reference interval may be indicated. Description of the reference interval study should include details about sampling; selection of subjects, including their number, age, and sex distribution; the statistical method for summarizing the results (15); and other factors that would influence the values obtained.
Chromatograms. Chromatograms from gas-liquid and liquid chromatography should usually be presented so that readers can see the efficiency of the separation and observe the resolution from interferents in the matrix. Similar images are often needed for electrophoretic separations.
Enzyme activities. Enzyme activities may be expressed in international units (U) or katals. Temperature and other key assay features must be described in the text or by reference to a published method. When first mentioned in the text, enzymes (whether measured by activity or mass assays) must be numbered (EC no.) in accordance with the recommendations of the Nomenclature Committee of the International Union of Biochemistry and Molecular Biology on the Nomenclature and Classification of Enzymes (16).
STUDIES OF DIAGNOSTIC ACCURACY
For studies of diagnostic accuracy of tests, complete and return the STARD Checklist for Evaluations of Diagnostic Accuracy (17). The STARD statement (17) and explanatory document (18) provide guidance. Provide literature reference(s) describing the evaluated test(s) and criterion ("gold") standard test(s) or include detailed descriptions of them. Follow accepted methodologic standards including the following: (a) Specify spectrum of evaluated patients (age and sex distributions, eligibility criteria, and summary of symptoms or disease stage). (b) Analyze pertinent subgroups of subjects (e.g., symptomatic and asymptomatic patients). (c) Avoid verification bias (usually by application of a "gold-standard" test to all subjects rather than to a clinically selected subset). (d) Categorize test results and patients independently to avoid reviewer bias (usually by performance of tests with blinding to patient information and vice versa). (e) Provide confidence intervals (or SE) for indices of diagnostic accuracy such as sensitivity/specificity, likelihood ratios, and areas under receiver-operating characteristic (ROC) curves (19). (f) Indicate the number of indeterminate test results and their use (if any) in further data analysis. (g) Provide laboratory data on analytical imprecision of the test (usually day-to-day CV at two or more concentrations) or reproducibility of observer interpretation [e.g., for a visually read, dichotomous (e.g., positive/negative) test]. See below for statistical treatment of data. A flow diagram is strongly recommended (17,18).
Describe statistical methods with enough detail to enable a knowledgeable reader with access to the original data to verify the reported results. When possible, quantify findings and present them with appropriate indicators of measurement error or uncertainty. Avoid sole reliance on statistical hypothesis testing, such as the use of P values, which fails to convey important quantitative information. When appropriate, confidence intervals should be presented; see, e.g., Harris (20), Henderson (21), and references therein.
Evaluation of diagnostic accuracy. (See also “Studies of Diagnostic Accuracy” above). In clinical studies, simple testing of the significance of differences between mean values of patient groups (e.g., by Student’s t-test) provides insufficient information to assess diagnostic accuracy. Scatter plots of data, calculations of diagnostic sensitivities and specificities and their confidence intervals (19), and use of approaches such as ROC curves (22), cumulative distribution analyses (23), likelihood ratios (24), and discriminant analysis (25) provide information that is appropriate to specific situations. Confidence intervals should be provided (17). Discussions of predictive values in illustrative settings may be useful additions to assess the potential clinical utility of tests. Analysis of serial measurements requires special attention (26).
Outcomes studies. The design and reporting of outcomes studies can be complex. The CONSORT statement (27), although designed for randomized controlled trials and used for therapeutic rather than diagnostic interventions, is recommended as an often-useful guide. For other guidance, contact the Editor.
Permissions and Reprints
Clinical Chemistry is copyrighted by the AACC. Permission to reproduce copyrighted material for scholarly or commercial use can be arranged by contacting the Permissions Coordinator, AACC, 2101 L Street, N.W., Suite 202, Washington, DC.20037-1558 or by email to firstname.lastname@example.org.
Reprints of papers published in Clinical Chemistry can be obtained at any time in multiples of 100. To order, contact the Cadmus Reprint Department at 1-800-407-9190. Authors of invited Editorials and Reviews receive 100 reprints free of charge.
- International Committee of Medical Journal Editors. Uniform requirements for manuscripts submitted to biomedical journals. Ann Intern Med 1997;126:36–47. (Also available at http://www.acponline.org/journals/annals/01jan97/unifreqr.htm. Updated May 1999).
- Committee on Care and Use of Laboratory Animals. Guide for the care and use of laboratory animals. Washington, DC: Institute of Laboratory Animal Resources, National Research Council, 1985:83pp.
- Scientific style and format (the CBE manual for authors, editors, and publishers, 6th ed). Northbrook, IL: Council of Biology Editors, 1994:825pp. (Address: 60 Revere Dr., Suite 500, Northbrook, IL 60062.)
- Dodd JS, ed. The ACS style guide: a manual for authors and editors, 2nd ed. Washington, DC: American Chemical Society, 1997:460pp. (Address: 1155 16th St., NW, Washington, DC 20036.)
- Taylor BN, US ed. The international system of units (SI). NIST Special Publication 330, Gaithersburg, MD: National Institute of Standards and Technology, 1991:62pp. (Periodically revised; for sale by Supt. of Documents, Code No. NSPUE2, US Government Printing Office, Washington, DC 20402-9325.)
- Rodbard D. Data processing for radioimmunoassays: an overview. In: Natelson S, Pesce AJ, Dietz AA, eds. Clinical chemistry and immunochemistry; chemical and cellular bases and applications in disease. Washington, DC: AACC, 1978:477–94.
- NCCLS Tentative Guideline EP5-T. User evaluation of precision performance of clinical chemistry devices. Wayne, PA: National Committee for Clinical Laboratory Standards, June 1984.
- Carey RN, Garber CC. Evaluation of methods. In: Kaplan LA, Pesce AJ, eds. Clinical chemistry. Theory, practice and correlation, 2nd ed. St. Louis: CV Mosby, 1989:290–310.
- Koch DO, Peters T Jr. Selection and evaluation of methods. In: Burtis CA, Ashwood ER, eds. Tietz textbook of clinical chemistry, 2nd ed. Philadelphia: WB Saunders, 1994:508–25.
- Linnet K. Necessary sample size for method comparison studies based on regression analysis. Clin Chem 1999;45:882–94.
- Linnet K. Evaluation of regression procedures for methods comparison studies. Clin Chem 1993;39:424–32.
- Martin RF. General Deming regression for estimating systematic bias and its confidence interval in method-comparison studies. Data supplement. GDR: Executable program for general Deming regression calculations and graphics. http://www.clinchem.org/content/vol46/issue1/.
- Bland JM, Altman DG. Statistical methods for assessing agreement between two methods of clinical measurement. Lancet 1986;i:307–10.
- Fraser CG, Petersen PH. Analytical performance characteristics should be judged against objective quality specifications. Clin Chem 1999;45:321–3.
- Solberg HE. Establishment and use of reference values. In: Burtis CA, Ashwood ER, eds. Tietz textbook of clinical chemistry, 2nd ed. Philadelphia: WB Saunders, 1994:454–84.
- International Union of Biochemistry and Molecular Biology, Nomenclature Committee. Enzyme nomenclature 1992. San Diego: Academic Press, 1992:862pp.
- STARD Group. Towards complete and accurate reporting of studies of diagnostic accuracy: the STARD statement. Clin Chem 2003;49:1–6.
- Bossuyt PM, Reitsma JB, Bruns DE, Gatsonis CA, Glasziou PP, Irwig LM, Moher D, Rennie D, de Vet HCW, Lijmer J. The STARD statement for reporting studies of diagnostic accuracy: explanation and elaboration. Clin Chem 2003;49:7–18.
- Harper R, Reeves B. Reporting of precision of estimates for diagnostic accuracy: a review. BMJ 1999;318:1322–3.
- Harris EK. On P values and confidence intervals (why can't we P with more confidence?) [Editorial]. Clin Chem 1993;39:927–8.
- Henderson AR. Chemistry with confidence: should Clinical Chemistry require confidence intervals for analytical and other data? [Opinion]. Clin Chem 1993;39:929–35.
- Zweig MH, Campbell G. Receiver-operating characteristic (ROC) plots: a fundamental evaluation tool in clinical medicine [Review]. Clin Chem 1993;39:561–77. Note that in Figs. 4–12 in this paper, the labels for the x-axis at the top and bottom are reversed. The (correct) dual labeling of the x-axis solves the problem of whether to plot specificity or 1 – specificity on the x-axis.
- Krouwer JS. Cumulative distribution analysis graphs—an alternative to ROC curves [Technical Brief]. Clin Chem 1987;33:2305–6.
- Albert A. On the use and computation of likelihood ratios in clinical chemistry. Clin Chem 1982;28:1113–9.
- Solberg HE. Discriminant analysis. Crit Rev Clin Lab Sci 1978;9:209–42.
- Matthews JNS, Altman DG, Campbell MJ, Royston P. Analysis of serial measurements in medical research. Br Med J 1990;300:230–5.
- Moher D, Schulz KF, Altman DG for the CONSORT Group. The CONSORT statement: revised recommendations for improving the quality of reports of parallel group randomized trials. www.consort-statement.org.