Concatenation does not add times together

[cbligaard]

Hi premessa team,

I have a question related to the concatenation of files. I have run my barcoded samples in separate runs on the CyTOF, and therefore I would like to concatenate these files before normalizing.
However, I have noticed that the concatenated files generated with premessa does not have an altered time variable. Does this not lead to problems in bead normalization?

Since time intervals are used to normalize, I think it is a problem that the time variable is not corrected (the time parameter in each subfile starts with 0). CATALYST for example does a correction...

Best, Christina

[pfgherardini]

Hi,

No it's not a problem in the sense that the normalization procedure does not look at the time column it just assumes that the events in the FCS are ordered (i.e. the first event happened before the second event etc.).

It is up to you to concatenate the files in the right order though. If the time order is A.fcs, B.fcs, C.fcs, then you have to concatenate the files in that order as the normalization will not check the time and just assume the order is correct