hello I need a complete procedure and list of regents for the...

Dear Bio-protocol user,
Thank you very much for your question. As stated in the title, our protocol is intended for the detection of ROS in Zebrafish using two specific reagents: the MitoSOX and the CellROX. While the MitoSOX is supposed to be more specific for superoxide generated in Mitochondria, CellROX is a more generic probe, labeling any kind of ROS in all the cellular districts. However, following our protocol, you should be able to use your favorite probe in order to label specific kind of ROS.
In any case, as mentioned in the protocol (“…the length of the incubation should be optimized based on the developmental stage, the staining solution and the oxidative status of the sample”), each researcher must find the optimal experimental conditions for the samples that will be analyzed. It’s impossible to prepare a protocol detailed enough to take in consideration all the possible variants (fish line, developmental stage, specific tissue/organ or cell type, oxidative status, probe concentration, ROS sub-type, etc…). Concerning the information about the Flow Cytometry approach, there is no specific knowledge that a FACS/sorter technician need to know to analyze the samples. Again, we omitted the info about that (with the exception of the instrument we used), basically because the conditions must be empirically determined and set by the researcher. Your specific conditions could vary depending for example on the zebrafish line in use, the cell type, the level of cell viability, the flow cytometer instrument in use, the power of the lasers, the threshold that you want to use, etc…
I'm sorry I can't be more helpful. If you have more specific questions, do not hesitate to contact me again and I will try to do my best to help you.
Best regards,
Alberto Rissone