zienpiggie

Hi science noob,

you are right, I made a mistake recalling what I did. It was fixed in 1% paraformaldehyde on ice for 1 hour, and then after washing with PBS twice was then placed in 70% ethanol and then stored at -20C overnight.

Your flow tubes are glass? Mine looks like it's made of polystyrene. My manual suggests to do my cell prep in the 12 x 75 mm test tubes as the 'polystyrene' tubes would have built up static so cells my adhere to the walls or something. But it does not specifically suggest 'glass'. In that case may be I should prep my samples in the polypropylene microfuge tubes instead then and only move it to the flow tubes on the last stage. Thanks for the suggestion.

There are a number of different probes that you can use depending on the type of free radicals you are targeting. Here's a list I found from invitrogen website http://www.invitroge...en-species.html .

For your BSA/milk blocking system, if there is a high background then may be you can try optimize the primary/secondary dilutions. I just had this problem a few weeks ago, so what I did was run a gel of the exact same sample, and then cut off each lane and then try out different combinations of primary and secondary dilutions, and see if it gets better.