Love:Protocols:Genepix analysis

Genepix Analysis

1. open slide as obtained from the scanner

2. rotate the slide

recall genepix reads from top to bottom, left to right. block 20 should be on top

3. save rotated version (multi* tif)

do it in a new folder

4. load rotated version and layout the blocks

load corresponding gal file from FILE > Load array list

start from an area where you feel comfortable in aligning the squares

replicate the successive ones; if you aligned well they should be pretty close in rotation and stretch;

this similarity may change as you go further away from the initial block

5. save settings (gps for each slide)

!!IMPORTANT make sure feature alignment options are as follows:

-find square features

-check resize feature during alignment

-min diam 85%

-max diam 400%

-check limit feat movement during align

-max transl 40

-flag feature not found

-composite pixel int thres 0

-automatic image reg 10px

ALSO: select the right ratios to be built.

!!NOTE save gps as "prealign"; this way when doing batch later we won't overwrite input & output gps

6. repeat for all rotated files

7. once all settings are saved, set up batch analysis on rotated slides

batch analysis (in new genepx version) would align features, analyze, [save with one flag if wanted]

load all tiffs

load each input gps for each tif separately

check "same as images" (do not select other folder, as linkage between image and results would be lost)

check align and configure alignment (make sure only 'align features' is checked) - this uses the settings you saved before

check analyze

[run flags if you want]

8. run batch

stay around for the first to silence potential prompt.