Abstract
Liposomes containing the photosensitive dye sulphonated aluminium phthalocyanine (AlSPc) were coupled to polyclonal sheep anti-mouse-Ig antibody and bound to cells coated with specific mouse monoclonal antibody. When illuminated with red light, the AlSPc in the liposomes was activated to produce singlet oxygen and the antibody and liposome targeted cells were destroyed. DW-BCL cells (an Epstein Barr virus immortalised B-cell line) were targeted with an anti-B-cell antibody (8A) and killed specifically, both alone and in the presence of bone marrow mononuclear cells (BM-cells), without phototoxic effects on the untargeted bone marrow CFU-GM progenitor cells. The presence of an excess of non-target cells did not interfere with antibody and liposome binding, or light access to target cells. Similar results were obtained with T-lymphocytes as target cells using anti-CD3 antibody. Specific targeting to the B-cells was demonstrated in the cell mixtures by use of fluorescent microscopy combined with a sensitive technique to detect low levels of AlSPc fluorescence, a cooled charge couple device (CCD) camera. This was also able to show low levels of non-specific background binding of AlSPc to BM-cells and a small population of cells that took up AlSPc in the absence of antibody. The latter were shown to be monocytes by flow cytometry.
Highlights
DW-BCL is an in vitro Epstein Barr virus (EBV) immortalised polyclonal B-cell line (Azim, 1988)
DW-BCL cells were seeded at 1 x 105 cells per ml and cultured in RPMI 1640 medium supplemented with 10% foetal calf serum
Bone marrow was harvested from normal donors, or patients with non-Hodgkin's lymphoma in remission undergoing harvest for autologous bone marrow transplantation
Summary
DW-BCL is an in vitro Epstein Barr virus (EBV) immortalised polyclonal B-cell line (Azim, 1988). DW-BCL cells were seeded at 1 x 105 cells per ml and cultured in RPMI 1640 medium supplemented with 10% foetal calf serum. Peripheral blood was collected from normal donors. The mononuclear cells of bone marrow (BM-cells), or peripheral blood (PBM) were obtained by layering the samples over Lymphoprep (Nycomed, Norway), centrifuging, and collecting and washing the cells at the interface. T-lymphocytes were purified from PBM by incubating in 20% FCS, in a plastic petri-dish at 37°C for 4 h to allow monocytes to adhere. Non adherent cells were removed and were found to be greater than 95% positive for the CD3 antigen (a marker of T-lymphocytes) using UCHTI antibody and flow cytometry
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