Purification of human primary neuroblastomas by magnetic beads and their in vitro culture.
In the present study we have successfully isolated neuroblastoma cells from primary human neuroblastoma tissues by using a magnetic bead-mediated purification system. Since primary neuroblastoma tissues contained CD3- and CD19-positive lymphocytes, total cell suspensions were prepared and incubated with magnetic beads coated with anti-CD3 or with anti-CD19 antibody. After magnetic separation, unbound materials were recovered and analyzed by immunohistochemical staining for NB84, one of the neuroblastoma markers. Immunohistochemical and FACS analyses demonstrated that NB84-positive cells were enriched in the unbound fraction. Subsequently, unbound materials were seeded on cell culture plates and maintained at 37 degrees C overnight. After incubation, non-adherent cells were collected and stained with anti-NB84 antibody. Under our experimental conditions, a significant increase in the number of NB84-positive cells was observed. Furthermore, our purified NB84-positive cells responded to all-trans retinoic acid and nerve growth factor better than the initial primary cells. Collectively, our present results suggest that magnetic bead-mediated purification enriches neuroblastoma cells which retain their biological properties.
Nakanishi
Hiroko
H
Division of Biochemistry, Chiba Cancer Center Research Institute, Chiba 260-8717, Japan.
Ozaki
Toshinori
T
Nakamura
Youko
Y
Hashizume
Kohei
K
Iwanaka
Tadashi
T
Nakagawara
Akira
A
D018941:Antigens, CD19 / Q000032:analysis; D017252:CD3 Complex / Q000032:analysis; D018929:Cell Culture Techniques / Q000379:methods*; D002469:Cell Separation / Q000379:methods*; D002648:Child; D002675:Child, Preschool; D005260:Female; D052203:Ferrosoferric Oxide / Q000737:chemistry; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D007223:Infant; D008214:Lymphocytes / Q000737:chemistry; D008297:Male; D008863:Microspheres; D009447:Neuroblastoma / Q000473:pathology*; D010524:Peripheral Nervous System Neoplasms / Q000473:pathology*; D014407:Tumor Cells, Cultured
D018941:Antigens, CD19; D017252:CD3 Complex; D052203:Ferrosoferric Oxide