2014 AAPS, SAN DIEGO, CA
Michael Tanen, Russell Weiner, Omar Laterza
Molecular Biomarkers & Diagnostics, Merck Research Laboratories, Rahway NJ 07065
Purpose:
Quanterix’ single molecule array Simoa technology is a fully automated immunoassay platform that utilizes arrays of femtoliter sized reaction chambers that can isolate and detect single molecules bound to paramagnetic beads. This method can measure proteins in the femtomolar range, up to 1000-fold improvement in sensitivity over traditional ELISA. Here we describe the development and optimization of a cytokine assay using the Quanterix platform.
Method:
The Quanterix platform is based on the use of capture antibodies coupled to paramagnetic beads and biotinylated detection antibodies, which in turn are detected by the action of a reporter enzyme: streptavidin β-galactosidase (SβG) and resorufin β-D-galactopyranoside as the fluorescence substrate. We executed a 2-variable design of experiments (DOE) strategy experiment in which multiple concentrations of detection antibody and SβG were tested. These are the 2 most critical variables to optimizing Simoa assays.
Results:
Under the optimal conditions identified (0.6 μg/mL detection antibody and 150 pM SβG), we were able to achieve an LOD of 1.4 fg/mL. This represents a 150-fold increase in sensitivity as compared to a current ultrasensitive assay developed on the Mesoscale Discovery platform.
Conclusions:
assays, with homebrew capabilities that allow for the use of proprietary reagents and control of the assay development and optimization process. The fully automated platform allows for rapid development time and efficiencies in sample analysis.
Related Posts
Neurology
Comparative Spatial Analyses of the Tumor Immune Landscape in Mouse Models of Glioblastoma Using the PhenoCode Discovery Mouse Neuro Panel and Customized Markers
Poster Background Developing effective therapies for glioblastoma (GBM) relies on robust preclinical animal models that enable detailed investigation of tumor biology, discovery of therapeutic targets, and evaluation of new treatment…
Oncology
Development of a Spatial Metabolic Map of Immunotherapy Sensitive and Resistant Cutaneous Skin Carcinoma
Poster Background Ultrahigh-Plex Single-cell Spatial Phenotyping has revealed valuable insights into the tumor immune microenvironment (TiME) for biomarker discovery and stratification of clinical responses. Metabolic reprogramming is a key hallmark…
Spatial Immunometabolic Profiling of Non-Small Cell Lung Cancer Reveals Distinct Immune Exclusion Patterns and Functional Niches
Analytical validation of a multiplex immunofluorescence 7-Biomarker Immunoprint® assay on the Akoya PhenoImager HT platform for stage I/II Melanoma
Neurology
Neurodegenerative biomarkers in different chambers of the eye relative to plasma: an agreement validation study
Alzheimer’s Research and Therapy | August 26, 2024 Sampani K, Ness S, Tuz-Zahra F, Aytan N, Spurlock EE, Alluri S, Chen X, Siegel NH, Alosco ML, Xia W, Tripodis Y,…
Neurology
Clinical validation of the LucentAD p-Tau 217 as a lab developed test (LDT) for clinical use
Poster The intersection of new therapeutic options for Alzheimer’s disease (AD), the emergence of accurate blood-based tests for AD, and the anticipated health system log jam for confirmatory diagnostic testing…
Neurology
Development of thresholds and a visualization tool for use of a blood test in routine clinical dementia practice
Alzheimer’s & Dementia | August 3, 2024 Verberk IMW, Jutte J, Kingma MY, Vigneswaran S, Gouda MMTEE, van Engelen MP, Alcolea D, Arranz J, Fortea J, Lleó A, Chevalier C,…
Neurology
Discovering novel plasma biomarkers for ischemic stroke: Lipidomic and metabolomic analyses in an aged mouse model
Journal of Lipid Research | August 2, 2024 Becktel DA, Frye JB, Le EH, Whitman SA, Schnellmann RG, Morrison HW, Doyle KP. J Lipid Res. 2024 https://doi.org/10.1016/j.jlr.2024.100614 Abstract Ischemic stroke…