Twelve metastatic melanoma samples were obtained from patients that were not undergoing therapy at the time of sample collection. Patients had undergone a wide range of prior therapies, including surgery, chemotherapy, radiotherapy, immunotherapy, or none of the above. PBLs were obtained by either leukapheresis or venipuncture, prepared over Ficoll-Hypaque gradient (LSM; ICN Biomedicals Inc.), and cryopreserved until analysis. After surgical resection, tumor specimens were processed as previously described (18 (link)). Briefly, tumor specimens were minced, enzymatically digested overnight at room temperature or for several hours at 37°C (RPMI-1640 with l-glutamine [Lonza], 1 mg/ml collagenase IV [Sigma-Aldrich], 30 U/ml DNAse [Genentech], and antibiotics) and the tissue was separated mechanically using gentleMACS (Miltenyi Biotech). Tumor single-cell suspensions were cryopreserved.
Protocol detail
Metastatic Melanoma Sample Preparation
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Antibiotics
Cell
Chemotherapy
Collagenase
Dnase
Ficoll
Glutamine
Hypaque
Immunotherapy
Leukapheresis
Melanoma
Ml iv
Patients
Radiotherapy
Sample collection
Surgical
Tissue
Tumor
Venipuncture
Corresponding Organization : National Cancer Institute
Other organizations : National Institute of Allergy and Infectious Diseases, Adaptive Biotechnologies (United States), Fred Hutch Cancer Center
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Variable analysis
independent variables
- Prior therapies (surgery, chemotherapy, radiotherapy, immunotherapy, or none of the above)
- Metastatic melanoma samples
- Patients were not undergoing therapy at the time of sample collection
- PBLs were obtained by either leukapheresis or venipuncture, prepared over Ficoll-Hypaque gradient (LSM; ICN Biomedicals Inc.), and cryopreserved until analysis
- Tumor specimens were processed as previously described (18 (link)): minced, enzymatically digested overnight at room temperature or for several hours at 37°C (RPMI-1640 with l-glutamine [Lonza], 1 mg/ml collagenase IV [Sigma-Aldrich], 30 U/ml DNAse [Genentech], and antibiotics), and the tissue was separated mechanically using gentleMACS (Miltenyi Biotech)
- The study does not explicitly mention any positive or negative controls.
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