Functional Proteomics Reverse Phase Protein Array Core Facility
Functional Proteomics
Functional proteomics is the accumulation of genetic lesions results in tumor development and progression. However, the genetic lesions integrate at the level of protein function to result in the cellular correlates of the hallmarks of cancer. Indeed, the correlation between DNA, RNA and protein changes is only about 40% requiring an ability to assess the functional proteome. Thus, a cell-based functional proteomics approach is required to determine the consequence of genetic aberrations in cancer cells.
Targeted therapeutics interfere with the function of cellular proteins, pathways and networks. A functional proteomics approach is needed for the validation of targets, demonstration of an off-target activity of drugs and evaluating pharmacodynamics.
Advantages of Reverse Phase Protein Arrays (RPPA)
A Cell-Based Functional Proteomics Approach
- Inexpensive, high-throughput method that can concurrently test hundreds of different proteins and pathways at expression levels and protein modifications, such as phosphorylation or cleavage
- Applicable to very small sample sizes (nanograms of protein lysates, detecting attomoles of a specific protein), less than 10-cell equivalents
- Recombinant protein/ phosphopeptides enables absolute quantification of protein and modified protein levels on over 1000 samples on a single slide
- Does not require direct labeling of the lysate sample
Getting Started
- Design your experiment based on biological question.
- Prepare protein lysates from cultured cells or tissue samples based on the procedure provided by the RPPA Shared Resource.
- Adjust protein concentration around 1 µg/µl. A minimum of 20 µl of protein lysates is required for RPPA analysis.
- Biological replicates are encouraged.
- For customer antibody validation, please provide your positive and negative cell lysates along with your western blot result and densitometer reading on each band.
Core Grant Citation
This facility is funded by NCI # CA16672. Publications should cite the Core grant in the acknowledgment section, if publications use data generated by the Core facility. Two copies of the publication acknowledging the Core grant should also be submitted to the facility at Unit 950.