I have one bulk soil sample which i will extract by two different methods. Both methods have around 6-7 steps before the purified protein is ready for mass spectrometery identification of the different proteins extracted by each method. The bulk sample is split in two A and B , sample A is subjected to a boiling treatment, some sonication and then is split into 5 parts for overnight protein precipitation . The five subsamples are cleaned and precipitated with methanol etc . Three different samples of these are separated on an SDS page gel. Each lane is in-gel digested and run through a mass spectrometer.
The second sample B is treated in a different manner for two processes and then is split into 8-9 for the overnight protein precipitation using a different chemistry. The precipitated protein is then centrifuged the next day and cleaned with solvent and prepared for mass spectrometry. Three different samples from B are then run on an SDS-page gel, in-gel digested and run on a mass spectrometer, giving MS and MS /MSprofiles.
I want to compare the groups of proteins I have extracted by both extraction methods A and B and make inferances from the differences. Perhaps to say that A method extracts proteins which belong more to membrane type protein families than method B . To do this I think I need triplicate observations so that I can make some statistical inferences. Can I call these triplicates of observations biological replicates rather than technical replicates ?
I would say you cannot call them biological replicates. In my opinion, you have one biological replicate with n = 3 for that sample. I personally would not call the n = 3 within your biological replicate a technical replicate, but that is likely to vary with the field you work in. From an MS perspective, a technical replicate is literally the same sample run two or more times under identical conditions on the MS.
To get n = 3 biological replicates, you need to repeat the entire process 3 times. Within each biological I would have n = 3. Others may have varying opinions on this. It should be a straightforward idea, but it's not always the case.
I hope that helps.
I would say you cannot call them biological replicates. In my opinion, you have one biological replicate with n = 3 for that sample. I personally would not call the n = 3 within your biological replicate a technical replicate, but that is likely to vary with the field you work in. From an MS perspective, a technical replicate is literally the same sample run two or more times under identical conditions on the MS.
To get n = 3 biological replicates, you need to repeat the entire process 3 times. Within each biological I would have n = 3. Others may have varying opinions on this. It should be a straightforward idea, but it's not always the case.
I hope that helps.
In general, biological replicates infers to the conduct of experiment on 3 different days or 3 different occasion, whereas, technical replicates implies to conduct of the experiment in replicates. I hope this helps.
So you start from a sample, then subdivided in two: A goes with a purification path, get split again and the split samples analyzed, B goes with a different purification path, get split again and analyzed.
Now a technical sample is, in this case, simply the sample split after the two different procedures, so you have 3 technical replicates of A and 3 technical replicates of B. Generally a technical replicate is defined as a sample at the end of a given procedure who get split before the actual analysis to account for internal variability (like measurement errors) whereas in a biological sample in a different sample from the same origin sampled in the same (or close as possible) condition and this account for biological variability.
So I don't think you can call the three split samples from each procedure biological sample.
The splitting of samples cab be called an experimental or a biochemical replicate.
Such an experiment can be very informative, especially when part of a multi step experimental procedure.
A biological experiment (defined well above) would incorporate the variability at the biological, experimental and (mass spectrometry) technical level.
For this reason actually biological replication is strongly recommended over experimental/technical replication.
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