Hello,
I am trying to purify intact nuclei from termite cells (Cryptotermes secundus to be precise) without any success. My procedure involved dissociation of the tissues using papain followed by lysis of the cells to release the nuclei. After nuclei release, I stain the nuclei using DAPI/tryptophan blue. Using either of the two dyes, the nuclei are not visible under a phase-contrast microscope which convinces me that the extraction was not successful.
Has anyone ever been successful in isolating intact nuclei from fixed termite cells? I would also appreciate it if you could share termites, honeybee, or ants cells/nuclei stained with DAPI or tryptophan blue.
The end goal is to use the nuclei for ChIP-seq analysis in a technique that requires me to isolate intact nuclei and fragment the DNA before lysing the nuclear membrane to release the nuclei (not the traditional sonication method).
Thank you,
You failed because the microscope did not record any data. In fact, when you contrast the nucleus measurment with a microscope, light passes through the nucleus and you have no reflected data. I suggest using a suitable label on the core so that the data is recorded by sending a beam under a microscope so that you can calculate the size of the core. It is best to use a Hilbert phase microscope or digital holography to measure the termite nucleus, as you no longer need to add color and worry about the nucleus information not being recorded.
Mahyar Radak Thank you for your response. However, the technique I was using has been used previously in mammalian cells (I guess they are a bit larger).
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