DIA has been acquired by scanning the entire slide through histology slide scanner (Aperio ScanScope) at resolution 1x.
Raw images were obtained by scanning the microscope slides with Agilent Scan G2505B and Agilent Scan Control software (version 9.5).
The three fields were first identified by scanning the entire slide at × 100 magnification.
We also tested photo bleaching of the 4 dyes by scanning the microarray slide up to 5 times with the same scanner settings for all 4 dyes, and plotted the mean signal intensities as percentage of the signal intensity at the first scan.
For each of the 44 cases (36 tumors and 8 normal prostate tissues), digital slides were acquired by scanning the H&E stained slide with AT Turbo (Aperio technologies, Vista, CA).. Every 2.1 × 1.3 mm area of the micrograph was then saved into a.tiff file at a resolution of 72 pixels per inch using the ImageScope software (Aperio technologies, Vista, CA).
Cantilevers and parameters were proved for proper work by scanning the surface of a cover slide.
Mirax Viewer Image Software Ver (1, 6) was used for scanning the Hematoxylin-eosine slides by a ZEISS-MIRAX Midi Slide scanning system (Zeiss Microimaging, Oberkochen, Germany and 3DTech, Budapest, Hungary).
The extraction of data from TIFF image files generated by scanning microarray slides was performed using Feature Extraction version 9.5.3.1 software (Agilent Technologies , Inc using the linear Lowess algorithm modeling method to obtain background subtracted and normalized intensity values.
After labeling cells ex vivo and injecting them into tumor-bearing mice, the distribution of cells in tumor and organs could be quantified in tissue sections with high throughput by scanning many slides at once.
Differential cell counts were measured by scanning cytospin slides with high power (×500) magnification.
