Daniel:Notebook/RNAFACS/2015-10-28
RNA Scope Trial Run 5: Pretreat III[edit]
Microscopy[edit]
Positive Control (20X)[edit]
- 20151028 PositiveControl.lif Series006Snapshot1 ch00.jpg
Field 1
- 20151028 PositiveControl.lif Series007Snapshot1 ch00.jpg
Field 2
- 20151028 PositiveControl.lif Series009Snapshot1 ch00.jpg
Field 3
- 20151028 PositiveControl.lif Series011Snapshot1 ch00.jpg
Field 4
It's possible that these are signal, but I can't tell because they overlap. I checked with the positive controls (POLR2A, PPIB, UBC) and they are present (according to gene cards), but I can't tell if RNA should be in the nucleus or not. Technically, most genes being transcribed probably are, but they need to be present in the nucleus for this to work.
Experiment (20X)[edit]
- 20151028 Experiment.lif Series003Snapshot1 ch00.jpg
Field 1
- 20151028 Experiment.lif Series005Snapshot1 ch00.jpg
Field 2
- 20151028 Experiment.lif Series007Snapshot1 ch00.jpg
Field 3
These, I believe, are true signal. They get close to overlap in some points, but the dots do appear distinct, so I believe this sample does probably contain signal. This means it should be visible in the 10X objective, which will be necessary if we wish to use the Cell Raft array.
Experiment (10X)[edit]
- 20151028 Experiment.lif Series010Snapshot1 ch00.jpg
Field 1
- 20151028 Experiment.lif Series015Snapshot1 ch00.jpg
Field 2
- 20151028 Experiment.lif Series018Snapshot1 ch00.jpg
Field 3
- 20151028 Experiment.lif Series021Snapshot1 ch00.jpg
Field 4
I don't believe you can see much with these samples. I may not be using the right settings, since using the confocal the gain and laser intensity are often different depending on the objective. The fact that the 10X uses air may also be a problem.