Hosuk:LabNotes/2013-11-6: Difference between revisions
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*2 pixel off, 281.25nm x 2 = 562.5 nm off to the right direction | *2 pixel off, 281.25nm x 2 = 562.5 nm off to the right direction | ||
[[File:Exp1_MovementTest_Before_AfterWashing_ACTB.png|600px]] | [[File:Exp1_MovementTest_Before_AfterWashing_ACTB.png|600px]] | ||
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*#Step7-0 : Strip | *#Step7-0 : Strip | ||
*#Step7 : ACTB + RAB7A only | *#Step7 : ACTB + RAB7A only | ||
====Result==== | ====Result==== | ||
Line 57: | Line 56: | ||
| w/i 1st Rolony | | w/i 1st Rolony | ||
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| 74 | | 74 [[Media:Exp2_DyeTest.lif_Step5_1stRolony_ACTB_20x_2k_Cy5_ATTO488_ch01_MIP.tif|Raw]] | ||
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| 1.45 | | 1.45 | ||
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| 114 | | 114 [[Media:Exp2_DyeTest.lif_Step3_1stRolony_RAB7A_20x_2k_Cy5_ATTO550_ch01_MIP.tif|Raw]] | ||
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| 2.25 | | 2.25 | ||
Line 67: | Line 66: | ||
| 1 probe only | | 1 probe only | ||
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| 157 | | 157 [[Media:Exp2_DyeTest.lif_Step6_AddACTBOnly_20x_2k_Cy5_ATTO488_ch01_MIP.tif|Raw]] | ||
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| 3.08 | | 3.08 | ||
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| 260 | | 260 [[Media:Exp2_DyeTest.lif_Step4_AddRAB7AOnly_20x_2k_Cy5_ATTO550_ch01_MIP.tif|Raw]] | ||
|| | || | ||
| 5.13 | | 5.13 | ||
Line 77: | Line 76: | ||
| ACTB+RAB7A | | ACTB+RAB7A | ||
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| 130 | | 130 [[Media:Exp2_DyeTest.lif_Step7_AddACTB_RAB7A_20x_2k_ATTO488_550_ch00_MIP.tif|Raw]] | ||
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| 2.55 | | 2.55 | ||
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| 268 | | 268 [[Media:Exp2_DyeTest.lif_Step7_AddACTB_RAB7A_20x_2k_ATTO488_550_ch01_MIP.tif|Raw]] | ||
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| 5.25 | | 5.25 |
Revision as of 17:34, 7 November 2013
Movement test before after washing, stripping
Procedure
- Image - ACTB (ATTO550)
- Aspirate, Add Formamide, Incubate, Aspirate, Washing with PBS
- Image again - ACTB (ATTO550)
Result
- 2 pixel off, 281.25nm x 2 = 562.5 nm off to the right direction
File:Exp1 MovementTest Before AfterWashing ACTB.png
ACTB, RAB7A probing test
- 1st Rolony probe (Cy5) + ACTB (ATTO488) + RAB7A (ATTO550) ACTB and RAB7A showed always weak signal.
- Previous experiment (10/29/13) ACTB were seen more when added ACTB only.
- So, let’s compare between RAB7A or ACTB and 1st Rolony probe(Cy5) together.
- And compare to ACTB + RAB7A mix
Procedure
- Step1 : RAB7A only
- Step2 : Strip
- Step3 : 1st Rolony + RAB7A
- Step4-0 : Strip
- Step4 : RAB7A only
- Step5-0 : Strip
- Step5 : 1st Rolony + ACTB
- Step6-0 : Strip
- Step6 : ACTB only
- Step7-0 : Strip
- Step7 : ACTB + RAB7A only
Result
- ACTB only had more than 1st Rolony+ACTB
- RAB7A only had more than 1st Rolony+RAB7A
- ACTB+RAB7A was similar to ACTB only or RAB7A only.
ACTB | (%) | RAB7A | (%) | |||||
w/i 1st Rolony | 74 Raw | 1.45 | 114 Raw | 2.25 | ||||
1 probe only | 157 Raw | 3.08 | 260 Raw | 5.13 | ||||
ACTB+RAB7A | 130 Raw | 2.55 | 268 Raw | 5.25 |
- Step1 : RAB7A only,
- RAB7A, Count = 514, Overlapped = 160
- Step3 : 1st Rolony + RAB7A,
- 1st Rolony Count = 5066
- RAB7A, Count = 186, Overlapped = 114
- Step4 : RAB7A only
- RAB7A, Count = 591, Overlapped = 260
- Step5 : 1st Rolony + ACTB
- 1st Rolony Count = 5105
- ACTB, Count = 137, Overlapped = 74
- Step6 : ACTB only
- ACTB, Count = 246, Overlapped = 157
- Step7 : ACTB + RAB7A only
- ACTB, Count = 234, Overlapped = 130
- RAB7A, Count = 463, Overlapped = 268