Historical Biomarker Assays
DCTD-supported Pharmacodynamic Biomarkers Program offers historical biomarker assays. Unlike other validated biomarker assays, these published assays do not have a formal training program or qualified reagent supply; however, consultation may be requested.
γH2AX, pNBS1, and β-Catenin Multiplex Immunofluorescence Assay
The γH2AX, pNBS1 IFA with β-Catenin segmentation is an immunofluorescence-based assay developed to quantify γH2AX and pNBS1 using β-Catenin staining to segment tumor from non-tumor to precisely limit quantitation of these DNA damage response markers to the tumor cells within a tumor biopsy section.
Assay SOPs
Specimen Collection:
- Sample Preparation
- Assay SOP
- Image Capture
- Image and Data Analysis
NCI Publications
Analysis of Circulating Tumor Cells Using the CellSearch System
Induction of γH2AX can be detected in topotecan-treated tumor cell lines and whole blood samples spiked with treated tumor cell lines using the γH2AX-AF488 antibody. A, Photomicrographs of control and 1 µM topotecan-treated MCF7 cells stained with γH2AX-AF488 (green) and DAPI (blue). 200x. B, MCF7 or PC-3 cells were treated with 1 µM topotecan for 2 h, spiked into whole blood from healthy donors, and stained using antibodies against CK-PE, CD45-APC, γH2AX-AF488, and DAPI. Check boxes adjacent to images: a checked box next to the composite image (CK-PE/DAPI) indicates a CTC; next to the CD45-APC image indicates the presence of a contaminating leukocyte; and next to an image for γH2AX-AF488 staining indicates a γH2AX-positive cell. Four representative fields of cells isolated using the CellTracks AutoPrep and detected with the CellTracks Analyzer II are presented for each cell line (i–iv). Image from Wang LH et al. Clin Cancer Res. 2010 February 1; 16(3): 1073
Biomarker Assessment
- CellSave Tube Blood Collection and Shipping Instructions
Antibody Qualification and Laboratory Proficiency Testing SOPs:
- Assay SOP
NCI Publications
- A phase I study of veliparib in combination with metronomic cyclophosphamide in adults with refractory solid tumors and lymphomas.
- Phase I study of PARP inhibitor ABT-888 in combination with topotecan in adults with refractory solid tumors and lymphomas.
- Histone γH2AX and poly(ADP-ribose) as clinical pharmacodynamic biomarkers.
- Monitoring drug-induced γH2AX as a pharmacodynamic biomarker in individual circulating tumor cells.
- Use of circulating tumor cells to monitor pharmacodynamic responses of patients in early stage clinical trials.
Immunoassays (ELISA)
Hypoxia Inducible Factor 1-alpha, ELISA
Solid Tumor Assessment
- Specimen Collection
- Sample Preparation
- Immunoassay SOP
- Data Analysis, Quality Control, and Reporting SOP
NCI Publications
- Validation of a hypoxia-inducible factor-1 alpha specimen collection procedure and quantitative enzyme-linked immunosorbent assay in solid tumor tissues.
- Pilot trial of EZN-2968, an antisense oligonucleotide inhibitor of hypoxia-inducible factor-1 alpha (HIF-1α), in patients with refractory solid tumors.
- Weekly EZN-2208 (PEGylated SN-38) in combination with bevacizumab in patients with refractory solid tumors.
Topoisomerase I (Top1) Immunoassay
Solid Tumor Assessment
- Specimen Collection
- Sample Preparation
- Immunoassay SOP
- Data Analysis, Quality Control, and Reporting SOP
Liquid Tissue Assessment
- Specimen Collection
- Sample Preparation
- Immunoassay SOP
- Data Analysis, Quality Control, and Reporting SOP
NCI Publications
- Development and validation of an immunoassay for quantification of topoisomerase I in solid tumor tissues.
- Development of a validated immunofluorescence assay for γH2AX as a pharmacodynamic marker of topoisomerase I inhibitor activity.
- Topoisomerase I levels in the NCI-60 cancer cell line panel determined by validated ELISA and microarray analysis and correlation with indenoisoquinoline sensitivity.
Contact
PDinfo@mail.nih.gov (questions, training requests, or comments)