Digital camera dxm 1200c

To evaluate the effects of α-Klotho on NF-κB activation by LPS, after purification, astrocytes were treated for 24 h with vehicle (control) (PBS) or 1 nM α-Klotho. Inflammatory stimulation with LPS 1 μg/ml was then performed and the cells were fixed with methanol (10 min) 4 h later and washed with PBS three times for 5 min. The fixed cells were incubated with serum blocking (5% normal donkey serum in triton X-100 0.01%) for one hour and incubated overnight with primary antibodies GFAP (1:300) (3670; Cell Signaling, and RelA (p65 (1:100) (ab7970; ABCAM, USA). The primary antibody was removed, and the plate was washed with serum blocking three times for 10 min. The cells were incubated with secondary antibody (rabbit or mouse anti-donkey- Alexa 594 or 488, Thermo Fischer Scientific, 1:1000), diluted in PBS with Triton X-100 0.01% for 2 h, and protected from the light. The coverslips were washed five times with PBS for 5 min and incubated with DAPI (4'-6-Diamidino-2-phenylindole; Sigma) for 1 min in dilution 1:10.000. Slices were transferred to glass slides and analyzed in Fluorescence microscope Nikon Eclipse 80i (Nikon, Tokyo, Japan) with a camera system, Nikon Digital Camera DXM 1200C.

Young panicles (80–90 mm), including both wild-type and Osfignl1 spikelets in meiosis were fixed with 3:1 ethanol:acetic acid (EAA), and stored at 4°C until observation. Microsporocytes undergoing meiotic division were squashed on glass slides and stained with an acetocarmine solution. Slides with chromosomes were frozen in liquid nitrogen. After removing the coverslips, the slides were dehydrated through an ethanol series (70, 90, and 100%). Male meiotic chromosomes were counterstained with 4′,6-diamidino-phenylindole (DAPI). Chromosome images were captured using Nikon Eclipse 90i microscope with Nikon Digital Camera DXM1200C and Nikon C-HGFI.