Multiplex serum biomarker assessments: technical and biostatistical issues

University of Pittsburgh Cancer Institute, Pittsburgh, PA 15213, USA.
Journal of Translational Medicine (Impact Factor: 3.99). 10/2011; 9:173. DOI: 10.1186/1479-5876-9-173
Source: PubMed

ABSTRACT Identification of predictive and prognostic biomarkers for patients with disease and undergoing different therapeutic options is a very active area of investigation. Many of these studies seek biomarkers among circulating proteins accessed in blood. Many levels of standardization in materials and procedures have been identified which can impact the resulting data.
Here, we have observed unexpected variability in levels of commonly tested analytes in serum which were processed and stored under standardized conditions. We have identified apparent changes in cytokine, chemokine and growth factor levels detected by multiplex Luminex assay in melanoma patient and healthy donor serum samples, over storage time at -80°C. Controls included Luminex kit standards, multiplexed cytokine standards and WHO cytokine controls. Data were analyzed by Wilcoxon rank-sum testing and Spearman's test for correlations.
The interpretation of these changes is confounded by lot-to-lot kit standard curve reagent changes made by a single manufacturer of Luminex kits.
This study identifies previously unknown sources of variation in a commonly used biomarker assay, and suggests additional levels of controls needed for identification of true changes in circulating protein levels.


Available from: John M Kirkwood, May 16, 2015
  • [Show abstract] [Hide abstract]
    ABSTRACT: Cytokines are bioactive proteins produced by many different cells of the immune system. Due to their role in different inflammatory disease states and maintaining homeostasis, there is enormous clinical interest in the quantitation of cytokines. The typical standard methods for quantitation of cytokines are immunoassay-based techniques including enzyme-linked immusorbent assays (ELISA) and bead-based immunoassays read by either standard or modified flow cytometers. A review of recent developments in analytical methods for measurements of cytokine proteins is provided. This review briefly covers cytokine biology and the analysis challenges associated with measurement of these biomarker proteins for understanding both health and disease. New techniques applied to immunoassay-based assays are presented along with the uses of aptamers, electrochemistry, mass spectrometry, optical resonator-based methods. Methods used for elucidating the release of cytokines from single cells as well as in vivo collection methods are described. Copyright © 2014 Elsevier B.V. All rights reserved.
    Analytica Chimica Acta 01/2015; 853C:95-115. DOI:10.1016/j.aca.2014.10.009 · 4.52 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The quest for a well-validated, non-invasive biomarker to aid in clinical decision making has remained elusive in the cancer space over the last 30 years. Much promise has been attached to circulating tumor cells (CTCs) as prognostic, predictive, and pharmacodynamic biomarkers with the potential to eliminate the need for invasive tumor biopsies and improve on the clinical value of other circulating tumor markers. The CellSearch® system (Veridex, LLC., Raritan, NJ, USA) cleared the U.S. Food and Drug Administration when the presence of CTCs was shown to have prognostic significance in patients with breast, prostate, and colorectal cancer. However, CTCs are not, at present, routinely being used in the clinic to guide treatment decisions. This paper discusses key attributes that a biomarker must possess, the status of other potential cancer biomarkers, and advancements in the capture and characterization of CTCs that will enable actualization of their potential as a reliable and efficient biomarker of disease diagnosis, progression, and response to therapy in the clinic.
    Drug Development Research 03/2013; 74(2). DOI:10.1002/ddr.21072 · 0.73 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Chronic wounds, including diabetic foot ulcers, pressure ulcers and venous leg ulcers, impact the lives of millions of people worldwide. These types of wounds represent a significant physical, social and financial burden to both patients and health care systems. Wound care has made great progress in recent years as a result of the critical research performed in academic, clinical and industrial settings. However, there has been relatively little translation of basic research discoveries into novel and effective treatments. One underlying reason for this paucity may be inconsistency in the methods of wound analysis and sample collection, resulting in the inability of researchers to accurately characterise the healing process and compare results from different studies. This review examines the various types of analytical methods being used in wound research today with emphasis on sampling techniques, processing and storage, and the findings call forth the wound care research community to standardise its approach to wound analysis in order to yield more robust and comparable data sets.
    International Wound Journal 01/2015; DOI:10.1111/iwj.12399 · 2.02 Impact Factor