Double Analysis – Studying RNA after Protein Analysis
CellCover is the only reagent in the market which allows the parallel storage of proteins, RNA and DNA within it’s cellular context while maintaining the integrity of cellular shape without chemical crosslinking. This means for example, that you can take a heterogeneous cell population, discriminate different cells types by antigen mining (immunolabelling), sort them by flow cytometry and then isolate RNA, DNA or proteins for further analyses.
Cells grown on almost any substrate could be treated with CellCover (liquid freezing) to maintain in vivo morphology without chemical crosslinking of biomolecules during cell fixation. RNA degradation is inhibited leading to great results in RNA sequencing or single cell sequencing experiments. Protein analyses benefit greatly from a sample’s treatment with CellCover too – compared to conventional formaldehyde fixation, formalin sensitive epitopes are readily accessible for antibodies in immunohistochemical experiments.
Some examples of protein analyses that can be performed with CellCover include protein expression, protein modification, protein sequencing and immunostaining.
Protein expression studies how protein is produced, modified and regulated. Proteins are isolated and often analyzed using tandem mass spectrometry. Proteins are produced by translation from mRNA, which is transcribed from DNA, so it can also be useful to study RNA and DNA to learn more about protein expression.
Protein modification, often referred to as post-translational protein modification, refers to changes that happen to a protein once it has already been translated from mRNA. Mass spectrometry is often used to study these changes.
Immunostaining is a technique using antibodies to detect the presence of a specific protein in a sample.
If your original sample is treated with CellCover first for any of these methods, you can go back to the sample days later to do follow up analysis.
Let’s take the example of immunostaining. By following our Basic Protocol 2 (Basic immunostaining protocol for suspension cells), if you use CellCover as an antibody diluent and washing buffer in your immunostaining experiments, you will be ready to process those same samples for successive RNA analysis, such as RNA sequencing or RNA FISH.