Western blot quantification: lane regions, background correction and HSP90 normalization

Video chapters

1. 0:00Open “条带灰度定量” (Densitometry).
2. 0:02Select “以内参归一化” (Normalize to reference).
3. 0:05Choose HSP90 as the reference for GSDMD at 30 kDa.
4. 0:11Turn off equal-width locking.
5. 0:14Drag the lane divider to avoid neighboring lanes.
6. 0:17Enable equal width again when uniform widths are needed.
7. 0:20Move one lane region vertically onto the 30 kDa signal.
8. 0:25Drag the blue edge to adjust the region height.
9. 0:29Enable equal height for all six lane regions.
10. 0:33Include the signal and a little background. Avoid the 53 kDa band above.
11. 0:39Click “确认泳道选区” (Confirm lane regions) for GSDMD.
12. 0:42Then click “确认泳道选区” for HSP90.
13. 0:47Click “计算灰度比值” (Calculate ratios) to measure original pixels.
14. 0:53The result table lists the target, reference and ratio for all six lanes.
15. 0:58Each ratio is GSDMD 30 kDa integral density divided by its HSP90 density.
16. 1:07Click “导出 Excel” (Export Excel) to save all six results and regions.
17. 1:11Open the actual exported workbook.
18. 1:14The six lane labels, integral densities and ratios match the result table.