Daniel:Notebook/HiResChrPaint/2014-8-6

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Revision as of 23:22, 6 August 2014 by >Djacobse (→‎Sample A)
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No Wash FISH

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Microscopy

First I'll test whether the samples can be taken directly to the microscope, with the hybridization buffer still on. This will produce a higher background noise, but will hopefully still result in strong areas of signal distinguishable from the background. Since this is just a test, I'll only use the positive control (sample D, Cot1-488) for now, and I'll investigate the others pending successful trial.

Sample D

As expected, the background is definitely higher, but the signal is also stronger. It will be interesting to do a wash on these samples, and then examine them after. Given the success, I'm going to return the positive control to the incubator and I'll take snapshots of the other 3 samples.

Sample A

Sample A is the Dystrophin gene labeled with Alexa 488. The sample has not been washed yet, but was prepared in the same manner I normally prepare samples.

Most cells showed no signal above the high background. However, several samples did show possible hybridization. Most photos shown here do show positive samples, although plenty of cells without signal can be seen also. From the Beliveau protocol I read that the success should be closer to 90%.

Sample B

Sample B is the Dystrophin gene labeled with Alexa 546. The sample has not been washed yet, but was prepared in the same manner I normally prepare samples.

Most cells showed no signal above the high background (ex. field 3). However, several samples (field 1 and 2) did show possible hybridization. From the Beliveau protocol I read that the success should be closer to 90%.

Sample C

Sample C is the dystrophin gene labeled with Alexa 488. The sample was not vacuum centrifuged, but rather 5 uL sample was added directly to the hybridization buffer.