Incubator pm s1

Mouse embryonic fetal livers were dissected at stage E15.5 from timed mating females. Fetal livers were incubated on ice with fluorescently conjugated antibodies for VE-Cadherin (eBiosciences). Fetal livers were plated for imaging in 1.5% low melting agarose (Sigma) with X-VIVO media (Lonza) and 10% fetal bovine serum. Cultures were maintained at 37 °C and 5% CO₂ using a Heating Insert P Lab-Tek S1 with an Incubator PM S1 (Zeiss). Images were acquired using an Axio Observer Z1 microscope with the LSM 700 scanning module (Zeiss).

For live cell imaging of in vitro Cre transfer, B16-GFP-Cre cells were added to adherent reporter MEF that had been labeled with 5 uM of CellTracker Blue (Molecular Probes #C2110) according to the manufacturer’s protocol. The video recording was initiated after 2 hours. Images were collected every 3–4 minutes with xyzt acquisition mode using an Axio Observer.Z1 microscope with the LSM 700 scanning module (Zeiss, Jena, Germany). Cultures were maintained at 37°C, 5% CO₂ using a Heating Insert P Lab-Tek S1 with an Incubator PM S1 (Zeiss, Jena, Germany).
For imaging of in vivo Cre transfer, B16-GFP-Cre tumors were grown in reporter mice as described above. After 18–20 days, mice were sacrificed, and tumors were harvested, coated in OCT, and flash frozen in liquid nitrogen. Cryosectioning was then performed to generate tumor sections 15 μm thick, which were then imaged with a Nikon D-Eclipse C1TE2000 confocal microscope (Nikon, Tokyo, Japan).