Multiplex Immunoassay Profiling of Hormones Involved in Metabolic Regulation.

Multiplex immunoassays are used for rapid profiling of biomarker proteins and small molecules in biological fluids. The advantages over single immunoassays include lower sample consumption, cost, and labor. This chapter details a protocol to develop a 5-plex assay for glucagon-like peptide 1, growth hormone, insulin, leptin, and thyroid-stimulating hormone on the Luminex platform.

Necroptotic debris including damaged mitochondria elicits sepsis-like syndrome during late-phase tularemia.

Infection with ssp. () strain SchuS4 causes an often lethal disease known as tularemia in rodents, non-human primates, and humans. subverts host cell death programs to facilitate their exponential replication within macrophages and other cell types during early respiratory infection (⩽72 h).

Recommendations for clinical biomarker specimen preservation and stability assessments.

With the wide use of biomarkers to enable critical drug-development decisions, there is a growing concern from scientific community on the need for a 'standardized process' for ensuring biomarker specimen stability and hence, a strong desire to share best practices on preserving the integrity of biomarker specimens in clinical trials and the design of studies to evaluate analyte stability. By leveraging representative industry experience, we have attempted to provide an overview of critical aspects of biomarker specimen stability commonly encountered during clinical development, including: planning of clinical sample collection procedures, clinical site training, selection of sample preservation buffers, shipping logistics, fit-for-purpose stability assessments in the analytical laboratory and presentation of case studies covering widely utilized biomarker specimen types.

Multiplex Immunoassay Profiling of Serum in Psychiatric Disorders.

Multiplex immunoassays allow for the rapid profiling of biomarker proteins in biological fluids, using less sample and labour than in single immunoassays. This chapter details the methods to develop and manufacture a 5-plex immunoassay for the Luminex® platform. Although assay development is not included here, the same methods can be used to covalently couple antibodies to the Luminex beads and to label antibodies for the screening of sandwich pairs, if needed.

Multiplex Immunoassay Profiling.

Multiplex immunoassays allow for the rapid profiling of biomarker proteins in biological fluids, using less sample and labor than single immunoassays. This chapter details the methods to develop and manufacture multiplex assays for the Luminex platform. Although assay development is not included here, the same methods can be used to covalently couple antibodies to the Luminex beads and to label antibodies for the screening of sandwich pairs, if needed.

Effect of a Smac Mimetic (TL32711, Birinapant) on the Apoptotic Program and Apoptosis Biomarkers Examined with Validated Multiplex Immunoassays Fit for Clinical Use.

Purpose: To support clinical pharmacodynamic evaluation of the Smac mimetic TL32711 (birinapant) and other apoptosis-targeting drugs, we describe the development, validation, and application of novel immunoassays for 15 cytosolic and membrane-associated proteins indicative of the induction, onset, and commitment to apoptosis in human tumors.

Experimental Design: The multiplex immunoassays were constructed on the Luminex platform with apoptosis biomarkers grouped into three panels. Panel 1 contains Bak, Bax, total caspase-3, total lamin-B (intact and 45 kDa fragment), and Smac; panel 2 contains Bad, Bax-Bcl-2 heterodimer, Bcl-xL, Bim, and Mcl1; and panel 3 contains active (cleaved) caspase-3, Bcl-xL-Bak heterodimer, Mcl1-Bak heterodimer, pS99-Bad, and survivin.

Recommendations for Use and Fit-for-Purpose Validation of Biomarker Multiplex Ligand Binding Assays in Drug Development.

Multiplex ligand binding assays (LBAs) are increasingly being used to support many stages of drug development. The complexity of multiplex assays creates many unique challenges in comparison to single-plexed assays leading to various adjustments for validation and potentially during sample analysis to accommodate all of the analytes being measured. This often requires a compromise in decision making with respect to choosing final assay conditions and acceptance criteria of some key assay parameters, depending on the intended use of the assay.

Recommendations for adaptation and validation of commercial kits for biomarker quantification in drug development.

Increasingly, commercial immunoassay kits are used to support drug discovery and development. Longitudinally consistent kit performance is crucial, but the degree to which kits and reagents are characterized by manufacturers is not standardized, nor are the approaches by users to adapt them and evaluate their performance through validation prior to use. These factors can negatively impact data quality.

SMA-MAP: a plasma protein panel for spinal muscular atrophy.

Objectives: Spinal Muscular Atrophy (SMA) presents challenges in (i) monitoring disease activity and predicting progression, (ii) designing trials that allow rapid assessment of candidate therapies, and (iii) understanding molecular causes and consequences of the disease. Validated biomarkers of SMA motor and non-motor function would offer utility in addressing these challenges. Our objectives were (i) to discover additional markers from the Biomarkers for SMA (BforSMA) study using an immunoassay platform, and (ii) to validate the putative biomarkers in an independent cohort of SMA patients collected from a multi-site natural history study (NHS).

Comprehensive serum profiling for the discovery of epithelial ovarian cancer biomarkers.

FDA-cleared ovarian cancer biomarkers are limited to CA-125 and HE4 for monitoring and recurrence and OVA1, a multivariate panel consisting of CA-125 and four additional biomarkers, for referring patients to a specialist. Due to relatively poor performance of these tests, more accurate and broadly applicable biomarkers are needed. We evaluated the dysregulation of 259 candidate cancer markers in serum samples from 499 patients.