ORIGINAL ARTICLES: Basic scienceIsolation and characterization of circulating melanoma cells by size filtration and fluorescent in-situ hybridizationYanagita, Masahikoa,b; Luke, Jason J.b,c; Hodi, Frank S.b,c; Jänne, Pasi A.a,b,c; Paweletz, Cloud P.a,bAuthor Information aBelfer Center for Applied Cancer Science bDepartment of Medical Oncology, Dana-Farber Cancer Institute cDepartment of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts, USA Correspondence to Dr Cloud P. Paweletz, PhD, Dana-Farber Cancer Institute, 450 Brookline Ave, Boston, MA 02215, USA Tel: +1 617 582 7602; fax: +1 617 582 9702; e-mail: [email protected] Present address: Jason J. Luke: Department of Medicine, University of Chicago Cancer Center, Chicago, Illinois, USA Melanoma Research: April 2018 - Volume 28 - Issue 2 - p 89-95 doi: 10.1097/CMR.0000000000000431 Buy SDC Metrics Abstract Isolation of circulating tumor cells (CTCs) from blood of melanoma patients has been difficult owing to inconsistent expression of surface antigens. Here we report on the isolation, detection, and characterization of CTCs from blood of melanoma patients using microfiltration and fluorescent in-situ hybridization (FISH). Two tubes of blood from 15 patients with advanced melanoma were collected. These two tubes subsequently underwent filtration through a membrane with pore sizes of 7.5 μm. Isolated cells from one tube were analyzed by FISH for RREB1 (6p24), MYB (6q32), SE6 (D6Z1), and CCND1 (11q13) and the other paired specimen was analyzed by immunofluorescence for HMB45, melanoma-associated antigen recognized by T cells-1, tyrosinase and melanogenesis associated transcription factor. We identified CTCs in 10 out of 13 melanoma samples by immunofluorescence (2.5–99 CTCs/3 ml of blood) and in 13 specimens by FISH (7.2–76 CTCs/3 ml of blood) with more CTCs identified by FISH in 10 out 13 samples. Two filters failed. Our results show that CTCs are detectable in the majority of patients with advanced melanoma. These tools will be useful in characterizing treatment related changes of melanoma in CTCs. Copyright © 2018 Wolters Kluwer Health, Inc. All rights reserved.