We have been actively evaluating, developing and implementing cutting edge immunoassays and innovative proteomic analysis technologies, and offer our expertise on high performance functional proteomic studies, such as: 1) Comprehensive and quantitative cell signaling event dissection. 2) Nanoscale quantitative proteomics assessment. 3) Biomarker & therapeutic target identification and validation. 4) On and off target drug activity assessment, pharmacodynamics evaluation. 5) Clinical applicable assay development and implementation. The capillary nanoimmunoassay (CNIA) (Simple WesternTM) and Luminex multiplexing immunoassay systems have been established and successfully supported various research projects from different CCR/NCI laboratories/branches/programs. The technologies address the challenges of conventional proteomic approaches, such as limited sample size, poor assay reproducibility, unreliable data quantification, and low successful rate on assay transfer from bench to bedside etc. High performance assays and analysis strategies are developed, research data were presented at national and international meetings, and multiple manuscripts have been published. Recently, a single-cell western system was acquired to accommodate the emerging demand in the field of single cell research. Collaborations with technology vendors are also formulated, which allows for continued technology improvement and accessibility of top-in-class methodologies to the CCR/NCI investigators. Implementation of the CNIA analysis platform provides high performance assays for comprehensive and quantitative signaling molecule profiling at protein level, esp. transferring those assays from discovery research into preclinical / clinical practice. A panel of about 200 CNIA assays, covering key signaling pathways from receptor activation, down-stream signaling transduction, transcriptional regulation, cell cycle control to apoptosis etc., has been developed / established. This allows us to provide CCR investigators a signaling network array for protein activity characterization, therapeutic target identification and drug selectivity determination, as well as defining the regulatory mechanisms. We have supported functional proteomic researches on a variety of projects including cell signaling profiling; biomarker development; targeted therapy evaluations and investigation on drug functioning mechanisms etc. Highly reproducible and sensitive assays with good analysis dynamic range and small sample consumption make the CNIA assays extremely appealing to clinical trial and drug development programs. High performance assays and analysis strategies have been developed and successfully implemented to monitor signaling molecule responses to drug treatment. This allows clinical investigators to 1) incorporate protein level companion diagnostic analysis and correlate proteomic with genomic data; 2) perform targeted therapy evaluation (both on-target and potential off-target effects); 3) determine pharmacodynamics and biologically relevant dose; and 4) determine prognosis and further modify treatment strategies etc. Recently, a panel of about 100 CNIA assays have been established in PBMC samples, allowing measuring of signaling molecule activities in specimens collected in a non-invasive manner. This enables samples from more treatment time points to be analyzed and provides more detail information of protein level responses to drug treatment. The Luminex assays provide multiplex in-solution ELISA analysis, based on the Luminex xMAP beads technology. The system is the most widely cited multiplex immunoassay platform in life science research. Implementation of this technology allows us to provide another high-performance analysis platform with small sample consumption for cytokine, metabolite, immune response, and serum/plasma biomarker etc. measurement. The single-cell western system performs western analysis on 1000-2000 single cells in parallel. The system measures proteins hard-to-detect by conventional single cell analysis platforms (e.g. FACS), such as isoforms, phosphorylated proteins, transcription factors, intracellular proteins etc. The technology is more affordable and less complicated to process than other single cell separation and protein analysis systems (e.g. CyTOF, Fluidium C1, and DEPArray systems etc.). It offers a cost effective tool for single cell signaling study and population heterogeneity dissection at the protein level. The technology is named #1 on The Scientist's Top 10 Innovations of 2016. Working with the CCR Office of Science and Technology Resource (OSTR), we have been continuously exploring/evaluating innovative proteomic analysis technologies to facilitate CCR research, such as 1) the Nanostring 3D Biology technology, using molecular barcodes and single molecule imaging for multiplex, simultaneous DNA, RNA, and protein measurement in a single reaction; 2) the ZellKraftwerk chipcytometry technology, an imaging cytometry system for quantitative analysis of 90 biomarkers on same cell or tissue samples while maintaining cell and biomarker integrity over a storage period of 20+ months; and 3) novel single cell proteomics analysis technologies etc. We actively offer our expertise through all project stages, such as project feasibility discussion, experiment design, method/analysis strategy development, sample preparation/analysis, data evaluation/summary, further project advancement advice and manuscript preparation assistance etc. Education and training have also been provided on technology applications and data analysis. The GLP concept is introduced in core operation to ensure data accuracy & reliability and as an effort to facilitate high quality/reproducible research at the CCR community. A web-based interface (https://cptr.cancer.gov) is established for better accessibility of our technologies and protocols, information & resource sharing, as well as more efficient project management.
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