Hello Everyone! I work in a wet lab but I obtained my degree in biotechnology. Every so often, I will get a challenge to try integrating bioinformatics in my lab. It gives me a good chance to develop my bioinformatician skills.
My challenge is this: a colleague is going to give me a dataset they generated from a 96 well plate. Essentially, they are looking at protein expression along each well. Using florescent, a quantitative value for particular proteins will be accounted for. Protein fluorescence will include the colors Red, Blue, Green, and Yellow. After the quantitative data is received, they want to use this want to use R as a means to analyze and visualize this data.
The thing really confusing me is that this experiment will have thresholds generated by our positive and negative controls. They want to use these to validate results, and while that does make sense , I don’t have the knowhow to actually set this parameter.
When I asked for an example of how the data set would look I got this:
I know that there are tons of resources for Flow Cytometry and protein expression but I am having a hard time grasping which method I should focus on for the sake of this challenge. I also don’t have a true data set to show because I am trying to learn how to utilize R in this fashion before I get the actual assignment. Any help would be greatly appreciated.
Their controls are just baselines? They will need subtracting I assume but I don't really understand from the question.
If you want to visualise/analyse, i don't think you'd need any specific packages for R. You might like to look at
ggplotfor visualisation though - a personal favourite.To clarify: I think the they want to use the values from the controls as a way to automatically include and exclude data points or as a way to reduce background. It seems like this is visually going to look a lot like Flow Cytometry data. However until I can obtain further information or the finished data set I wont know. I guess what I should really be asking is:
a) What is the best way to graph this data in a way which automatically uses the Positive and Negative Controls as a threshold?
b)For generally learning in R Which are the most user friendly packages for analyzing Protein expression and/or Flow Cytometry data?
Regarding flow cytometry and R, have a look a the Bioconductor flowCore package to see if it suits your needs. If the experiments are run in 96-well plates, you could look into RNAi screens to get ideas on how people analyze plate data, see this paper as a starting point. In this respect, you may find the Bioconductor package cellHTS2 useful.
I don't understand what question a) means. The closest thing I can think of is the Z factor (which you'll find described in the paper I mentioned above).
Thank you for your responses. I think when more information and my dataset are available, I will post a new question with much more clarity and definition.