Hosuk:LabNotes/2013-10-24: Difference between revisions
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===Rolony count with PISA=== | ===Rolony count with PISA=== | ||
====1 tile, 2048 x 2048 image ==== | ====1 tile, 2048 x 2048 image ==== | ||
*Sample : Confocal image of S2 MatTek dish at [[Hosuk:LabNotes/2013-10-22|2013-10-22]] | |||
**Cells : about 70 cells, counted cells just by eye based on compact rolonies (assume nucleus) | |||
*Size : 581.25um x 581.25um | |||
*1st Rolony (Cy5) parameters | *1st Rolony (Cy5) parameters | ||
Line 30: | Line 31: | ||
[[File:ATTO488SegOverlapwithCy5_S2_20131022.png|600px]] | [[File:ATTO488SegOverlapwithCy5_S2_20131022.png|600px]] | ||
Line 39: | Line 36: | ||
*Crop this down to 4 x 4 (7453 x 7453 px) | *Crop this down to 4 x 4 (7453 x 7453 px) | ||
**Cells ~ about 70 x 16 = 1120 | **Cells ~ about 70 x 16 = 1120 | ||
*Size : 2.1mm x 2.1mm | |||
Line 56: | Line 54: | ||
*ACTB / 1st rolony = 363/41824 = 0.87% | *ACTB / 1st rolony = 363/41824 = 0.87% | ||
**Overlapped ACTB --> 85 (0.076 ACBT per cell) | **'''Overlapped ACTB --> 85''' (0.076 ACBT per cell) | ||
**Overlapped ACTB / 1st rolony = 85/41824 = 0.20% | **Overlapped ACTB / 1st rolony = 85/41824 = 0.20% | ||
Line 62: | Line 60: | ||
*Overlaped with ATTO488 on Cy5 | *Overlaped with ATTO488 on Cy5 | ||
**Blue : Cy5, Red : ATTO488 : [[Media:ATTO488SegOverlapwithCy5_S2_Tile_4x4_20131022.tif|Raw]] | **Blue : Cy5, Red : ATTO488 : [[Media:ATTO488SegOverlapwithCy5_S2_Tile_4x4_20131022.tif|Raw]] | ||
====Multiple 7 x 6 tiles, --> 13077 x 11334 image ==== | |||
*Size : 3.7mm x 3.2mm | |||
*processed (after rebooting PC … :( ) | |||
*7 x 6 tiles, (13077 x 11334 px), but about 7 tiles were not images well… because of miss focusing miss | |||
**Need to take more longer Z scanning! | |||
*Cells ~ about 70 x 35 = 2450 | |||
*Sample : S2_20131022_20x_2kx2k_pos1_Zstack-1.2um_MaxProjection_Cy5.tif | |||
**Area Upper : 100 px, Area lower : 2 px, | |||
**Axioratio lower 0.7, Circ Upper 1.6 Circ Lower 0.4 | |||
**Perim conn 8, bkgmulti lower 2 | |||
**'''Count --> 100424''' (41.0 rolonies per cell) | |||
*Sample : S2_20131022_20x_2kx2k_pos1_Zstack-1.2um_MaxProjection_ATTO488.tif | |||
**Area Upper : 100 px, Area lower : 2 px, | |||
**Axioratio lower 0.7, Circ Upper 1.6 Circ Lower 0.4 | |||
**Perim conn 8, bkgmulti lower 2 | |||
**'''Count --> 892''' (0. 32 ACBT per cell) | |||
*ACTB / 1st rolony = 892/100424 = 0.89% | |||
**'''Overlapped ACTB --> 213''' (0.087 ACBT per cell) | |||
**Overlapped ACTB / 1st rolony = 213/100424 = 0.21% | |||
*Overlaped with ATTO488 on Cy5 | |||
**Blue : Cy5, Red : ATTO488 : [[Media:ATTO488SegOverlapwithCy5_S2_Tile_7x6_20131022.tif|Raw]] |
Latest revision as of 21:44, 24 October 2013
Rolony count with PISA[edit]
1 tile, 2048 x 2048 image[edit]
- Sample : Confocal image of S2 MatTek dish at 2013-10-22
- Cells : about 70 cells, counted cells just by eye based on compact rolonies (assume nucleus)
- Size : 581.25um x 581.25um
- 1st Rolony (Cy5) parameters
- Area Upper : 100 px, Area lower : 2 px,
- Axioratio lower 0.7, Circ Upper 1.6 Circ Lower 0.4
- Perim conn 8, bkgmulti lower 2
- Count --> 5210 (about 74.4 cells per cell)
- ACTB (ATTO488) parameters
- Area Upper : 100 px, Area lower : 2 px,
- Axioratio lower 0.7, Circ Upper 1.6, Circ Lower 0.4
- Perim conn 8, bkgmulti lower 2
- Count --> 69 (about 1 cells per cell)
- Overlapped ACTB --> 40 (about 0.57 cells per cell)
- Overlapped ACTB / 1st rolony = 40/5210 = 0.77%
- Overlaped with ATTO488 on Cy5
- Blue : Cy5, Red : ATTO488 : Raw
File:ATTO488SegOverlapwithCy5 S2 20131022.png
Multiple 7 x 6 tiles, --> Cropped to 4 x 4 --> 7453 x 7453 image[edit]
- Crop this down to 4 x 4 (7453 x 7453 px)
- Cells ~ about 70 x 16 = 1120
- Size : 2.1mm x 2.1mm
- Sample : S2_20131022_20x_2kx2k_pos1_Zstack-1.2um_MaxProjection_Crop4x4_Cy5.tif
- Area Upper : 100 px, Area lower : 2 px,
- Axioratio lower 0.7, Circ Upper 1.6 Circ Lower 0.4
- Perim conn 8, bkgmulti lower 2
- Count --> 41824 (37.3 rolonies per cell)
- Sample : S2_20131022_20x_2kx2k_pos1_Zstack-1.2um_MaxProjection_Crop4x4_ATTO488.tif
- Area Upper : 100 px, Area lower : 2 px,
- Axioratio lower 0.7, Circ Upper 1.6, Circ Lower 0.4
- Perim conn 8, bkgmulti lower 2
- Count --> 363 (0.32 ACBT per cell)
- ACTB / 1st rolony = 363/41824 = 0.87%
- Overlapped ACTB --> 85 (0.076 ACBT per cell)
- Overlapped ACTB / 1st rolony = 85/41824 = 0.20%
- Overlaped with ATTO488 on Cy5
- Blue : Cy5, Red : ATTO488 : Raw
Multiple 7 x 6 tiles, --> 13077 x 11334 image[edit]
- Size : 3.7mm x 3.2mm
- processed (after rebooting PC … :( )
- 7 x 6 tiles, (13077 x 11334 px), but about 7 tiles were not images well… because of miss focusing miss
- Need to take more longer Z scanning!
- Cells ~ about 70 x 35 = 2450
- Sample : S2_20131022_20x_2kx2k_pos1_Zstack-1.2um_MaxProjection_Cy5.tif
- Area Upper : 100 px, Area lower : 2 px,
- Axioratio lower 0.7, Circ Upper 1.6 Circ Lower 0.4
- Perim conn 8, bkgmulti lower 2
- Count --> 100424 (41.0 rolonies per cell)
- Sample : S2_20131022_20x_2kx2k_pos1_Zstack-1.2um_MaxProjection_ATTO488.tif
- Area Upper : 100 px, Area lower : 2 px,
- Axioratio lower 0.7, Circ Upper 1.6 Circ Lower 0.4
- Perim conn 8, bkgmulti lower 2
- Count --> 892 (0. 32 ACBT per cell)
- ACTB / 1st rolony = 892/100424 = 0.89%
- Overlapped ACTB --> 213 (0.087 ACBT per cell)
- Overlapped ACTB / 1st rolony = 213/100424 = 0.21%
- Overlaped with ATTO488 on Cy5
- Blue : Cy5, Red : ATTO488 : Raw