Cultrex® In Vitro Angiogenesis Assay Tube Formation Kit

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	Cells are supplemented with 2% FBS containing SH-SY5Y conditioned media alone or supplemented with 150 ng/ml anti-VEGF or melatonin 1 mM and after incubation for 5 h,tube-like formation was observed by microscopy.

Protocol tips
Cells are supplemented with 2% FBS containing SH-SY5Y conditioned media alone or supplemented with 150 ng/ml anti-VEGF or melatonin 1 mM and after incubation for 5 h,tube-like formation was observed by microscopy.

Publication protocol

To study the effects of SH-SY5Y conditioned media on the formation of tubular structures in HUVECs we used the In vitro Angiogenesis Assay Tube Formation kit (Cultrex; Trevigen Inc., Gaithersburg, MD, USA) according to the manufacturer's instructions. First, reduced growth factor basement membrane extract (BME) was pippetted into a 24-well plate and polymerized for 1 h at 37̊C. HUVECs (1×105/well) were then seeded in VCBM supplemented with 2% FBS containing SH-SY5Y conditioned media alone or supplemented with 150 ng/ml anti-VEGF or melatonin 1 mM. After 5 h of incubation, 4 random fields were selected/well and tubular structures were photographed using a Nikon Sight DS-SML1 camera (Sendai Nikon Corporation, Miyagi, Japan) attached to a Nikon Eclipse Ti-S fluorescence microscope at a magnification of ×10. The formation of the tubular network was assessed using ImageJ 1.45S software. Each assay was performed three times.


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Manufacturer protocol

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