Axio imager a2 upright microscope

To provide an immunocompetent model for inhibition of HCV protein expression, a mouse strain harbouring an HCV transgene was generated via a Cre/loxP switching system37 (link). We bred CN2-29 transgenic mice, which carry an HCV transgene (nt. 294–3435), with Mx1-Cre transgenic mice, which express Cre recombinase in response to interferon (IFN)-α or a chemical inducer of IFN-α, poly(I:C). Following poly(I:C) injection, the HCV transgene was rearranged, and HCV sequences were expressed in the livers. In this model, HCV structure proteins are expressed in the liver within 7 days after poly(I:C) injection. Male CN2-29 trasgegenic mice (8–9 week-old) were injected intraperitoneally with 0.3 mL of 1 mg/mL poly(I:C) solution [in PBS (-)]. At 6 months after the poly(I:C) injection, the CN2-29 mice were administrated intravenously twice with the siRNA-MEND complex solution [1 mg/mL in PBS(-)] via orbital sinus at day 0 and day 2. The mice were sacrifice under anesthesia with ketamine/xylazine 2 days after the second siRNA-MEND administration. Livers were removed, fixed in 10% buffered formalin, and embedded in paraffin. Section (4 μm) were stained with hematoxylin and eosin, and observed using ZEISS Axio Imager A2 upright microscope (Carl Zeiss MicroImaging, Inc, Germany).

Arabidopsis seedling roots were stained with monodansylcadaverine (MDC) as described previously (Contento et al., 2005 (link)). MDC-stained seedlings were observed with a Zeiss Axio Imager.A2 upright microscope (Zeiss) equipped with Zeiss Axiocam BW/color digital cameras using a DAPI-specific filter at the Iowa State University Microscopy and Nanoimaging Facility. GFP-ATG8e transgenic seedlings were observed and photographed with the same microscope system with a GFP-specific filter. Cells within the root elongation zone were photographed and the number of autophagosomes in each image was counted and averaged from at least 10 images per sample. Confocal microscopy images of autophagosomes in root cells and leaf protoplasts were taken using a Leica SP5 × MP confocal/multiphoton microscope system (Leica) with a 63x/1.4 oil immersion objective at the Iowa State University Roy J. Carver High Resolution Microscopy Facility (Pu and Bassham, 2016 (link)).

Cells were filmed with a Axiocam MRm camera coupled to a Zeiss AxioImager A2 upright microscope fitted with a Zeiss 40×/0.75 Plan-Neofluar objective and X-Cite 120Q Metal Halide lamp, controlled by Axio Vision SE64 Rel 4.9.1. The resulting images were converted to black and white and analyzed using ImageJ 1.50i (Fig 1). Intensity of immunofluorescence was quantified by corrected total cell fluorescence (CTCF), by substracting background signal (average signal per pixel for a region selected just beside the cell) from whole cell signal (sum of the intensity of the pixels for one cell) [29 (link)]. For each sample, five CTCFs were measured and the average was calculated for statistical analysis.