Abstract
Purpose: :
To employ surface plasmon resonance to target and kill endothelial cells by means of IgG–conjugated quantum dots (gold nanospheres).
Methods: :
6 confluent monolayers of mouse vascular endothelial cells grown in 2% gelatin–coated 4–well LabTek glass chambered slides were each incubated with 1 µg/ml anti–mouse CD31 (PECAM–1) monoclonal rat IgG for 30 minutes at 37º. Three 500 µl 1% BSA washes were performed in all wells. Monolayers in wells #1, 3, 5 were incubated with 30nm quantum dots (gold nanospheres) conjugated with 1 µg/ml anti–rat IgG goat IgG at 37º for 30 minutes. One 500 µl 1% BSA wash was performed in all six wells. Endothelial monolayers were covered with 500 µl of media per well and wells #1–2 and 3–4 were irradiated with 50 or 100 pulses, respectively, of a 532–nm, 0.5 J/cm2, Q–switched Nd:YAG laser (6 mm spot). Media was removed and Trypan blue exclusion testing was performed to assay cell viability. Viable and non–viable cells were counted within the 6 mm irradiated spot for wells #1–4 and within a corresponding area for wells #5–6 by using an Axiovert 25 microscope (Carl Zeiss, Inc., Thornwood, NY) with a CWHK grid ocular (Olympus America Inc., Melville, NY) (ob 10×).
Results: :
Unirradiated cells (wells # 5 and 6) demonstrated only viable cells (0% cell death) regardless of the presence (well 5: 84 viable, 0 non–viable cells) or absence (well 6: 84 viable, 0 non–viable cells) of quantum dots. Similarly, irradiated cells without quantum dots demonstrated only viable cells regardless of exposure to 50 pulses (well 2: 77 viable, 0 non–viable cells) or 100 pulses (well 4: 73 viable, 0 non–viable cells). In dramatic contrast, endothelial cells labeled with quantum dots and exposed to 50 laser pulses demonstrated 100% non–viable cells (well 1: 0 viable, 63 non–viable cells). Endothelial cells labeled with quantum dots and exposed to 100 laser pulses demonstrated complete disintegration with few identifiable cells remaining (well 3: 0 viable, 1 non–viable cell).
Conclusions: :
Antibody conjugated quantum dot mediated surface plasmon resonance can achieve 100% vascular endothelial cell lethality in vitro. Application of this technology in vivo holds promise for treatment of angiogenesis as in proliferative diabetic retinopathy and choroidal neovascularization.
Keywords: neovascularization • growth factors/growth factor receptors • phagocytosis and killing