Irdye 800cw 2 dg optical probe

N-methyl-D-Aspartate (NMDA), NMDA receptor inhibitor, MK801 (Tocris), PKC inhibitor, Staurosporine (Selleckchem), CaMKII inhibitor, KN-93 (Merck-Millipore) were purchased from manufacturers. Fluo-4 AM, AF594-conjugated Transferrin (Thermoscientific), FITC-ChromPure Mouse Transferrin (Jackson Immunoresearch), and IRDye 800CW 2-DG Optical Probe (Li-Cor) were purchased from manufacturers. Anti-GFAP (Cat# 12389S), NeuN (Cat# 24307S), Caveolin-1 (Cat# 3238) antibodies, Rab5 (Cat# 3547S) were purchased from Cell Signaling Technology. Anti-CD31 antibody (Cat# 550,274, BD Bioscience), Anti-Glucose transporter 1 (GLUT1, Cat# MA1-37,783, Invitrogen), Clathrin Heavy Chain antibodies (Cat# MA1-065, Invitrogen), Anti CD13 (Cat# ab7417, Abcam), and LAMP1 (Cat# sc20011, Santa Cruz Biotechnology) were purchased from respective manufaturer. Dilutions for all antibodies were 1:500. Human primary brain endothelial cells were purchased from Cell Systems (Cat # ACBRI 376). Cells were isolated from microvessels from brains of normal, healthy donor and are positive for CD31, vWF. Experiments were done cells with passage number less than 8 for the maintenance of their properties.

All experimental procedures were conducted in accordance with the Federation of Laboratory Animal Science Associations (FELASA) and were approved by the Bar-Ilan University animal care and use committee in Israel #27-04-2015. A total of 1 × 10⁶ Huh7.5 cells infected with HCV HJ3-5 virus or non-infected control cells were administered into 6-week-old NOD-SCID-gamma (NSG) male mice by intrahepatic injection into the left lobes of the liver. At four weeks following tumor cell injection, an IRDye 800CW 2-DG optical probe (LI-COR Biosciences, Lincoln, NE, USA) was injected into the tail vein of tumor-bearing mice. Mice were imaged using the Pearl Trilogy small animal imaging system (LI-COR Biosciences). Fluorescent signals from acquired images were analyzed using the Image Studio Lite software (LI-COR Biosciences). At 2 and 4 weeks following injection, the left lobe and right lobes of the liver as well as lungs from each mouse were harvested, and DNA or RNA was extracted for further analyses.

A substitute for glucose, 2-DG, is taken up by inflammatory tissue and can be quantitated using a near-infrared fluorescence imaging system (Pearl, Li-COR Biosciences, Lincoln, NE, USA). One hour after the intraperitoneal injection of LPS in mice, a 10 nmol IRDye 800CW 2-DG optical probe (C71103-07, LI-COR Biosciences, Lincoln, NE, USA) was injected into the tail vein. Twenty-four hours later, a small animal live imaging system was utilized to monitor inflammation in the mice [61 (link)]. The results of this section are displayed in the Supplementary Materials Figure S2.