Exact(5)
Using the same setting, experimental slides were imaged.
Unstained control slides and PBS control samples were also used to distinguish background and establish appropriate laser intensity for visualizing experimental slides.
Rabbit IgG (Upstate), Chicken IgY (Aves) and Mouse IgG (Upstate) were used on Control slides with the same secondary as the experimental slides to test for background levels of staining or immunofluorescence.
Control (no primary antibody) and experimental slides were incubated overnight at 4°C followed by 30 minutes at room temperature, respectively, in 5% Serum block goat alone (Dako X0907), or with H2AX antibody (abcam ab2893) at 1 500 dilution.
After three washes in Tris Buffered Saline (TBS) for 3 minutes, control (no primary antibody) and experimental slides were incubated for 50 minutes at room temperature, respectively, in Dual Block (Dako S2003) alone, or with lamin A/C antibody (cell signalling 2032) at 1 50 dilution.
Similar(55)
The file for the control image was similarly generated: the control slide is acquired and treated identically as the experimental slide.
Digital images from the experimental glass slides were obtained using ScanScope Digital Slide Scanner at a 20× magnification and archived on the devoted Spectrum Server V10.2.2.2315.
Digital images from the experimental glass slides were obtained using ScanScope Digital Slide Scanner (Aperio, Vista, CA, USA) at a 20× magnification and archived on the devoted Spectrum Server V10.2.2.2315 (Aperio).
In addition, Tables S2 and S3 contain supplementary data which list the 100 genes from our genome-scale analysis which have the highest relative gene activity on any of the 9 experimental array slides (Table S2), or the 100 genes in the human genome with the lowest relative gene activity at any time point in shake 2 (Table S3).
At experimental endpoint slides were fixed with 100% methanol and stained with 10% Giemsa in water.
The lower reproducibility of the among-slide group may be due to the experimental variations among slides, such as hybridization temperature fluctuation, washing, and scanning.
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