Invitrogen™ FAK[pY397] Human ProcartaPlex™ Simplex Kit

For preparation of cell lysates, we recommend the use of either ProcartaPlex Cell Lysis Buffer (Cat. No. EPX-99999-000) or Cell Extraction Buffer (Cat. No. FNN0011) supplemented with 0.5 M EDTA and Halt Protease and Phosphatase Inhibitor Single to Use Cocktail 100X (Cat. No. 78440 or 78442). These buffers are not included in the kit. Depending on your cell system, other buffers can be suitable as well.

The ProcartaPlex Human FAK[pY397] Simplex Kit is part of the ProcartaPlex Human MAPK Pathway Panel (phospho) 6plex (Cat. No. EPX060-97117-PHO), which complements the ProcartaPlex Human MAPK Pathway Panel (total) 6plex (Cat. No. EPX060-97017-901).

About ProcartaPlex Assays for the Luminex Platform

ProcartaPlex immunoassays are based on the principles of a sandwich ELISA, using two highly specific antibodies binding to different epitopes of one protein to quantitate all protein targets simultaneously using a Luminex instrument. ProcartaPlex multiplex assays require as little as 25 μL of plasma or serum, or 50 μL of cell culture supernatant, and just four hours to obtain analyzed results.

• More results per sample - measure multiple protein targets simultaneously in a single 25 to 50 μL sample
• Well-established Luminex technology - highly referenced multiplexing platform for protein detection and quantitation

ProcartaPlex assays utilize Luminex xMAP (multianalyte profiling) technology for the simultaneous detection and quantitation of up to 80 protein targets in a single 25 to 50 μL sample from plasma, serum, cell culture supernatants, and other bodily fluids.

The Luminex beads in the ProcartaPlex assay are internally dyed with precise proportions of red and infrared fluorophores to create spectrally unique signatures that can be identified by the Luminex xMAP detection systems (e.g., Luminex 200, FLEXMAP 3D, and MAGPIX). Similar to a sandwich ELISA, the ProcartaPlex assay uses matched antibody pairs to identify the protein of interest. In a multiplexed assay, each spectrally unique bead is labeled with antibodies specific for a single target protein, and bound proteins are identified with biotinylated antibodies and streptavidin to R-phycoerythrin (RPE). The conjugation of protein-specific antibodies to a distinct bead allows for analysis of multiple targets in a single well.

The most significant difference between a ProcartaPlex assay and ELISA is that the capture antibody in the ProcartaPlex assay is conjugated to a bead and not adsorbed to the microplate well, so the ProcartaPlex assay reagents are free-floating in the solution. For detection, the Luminex 200 instrument, for example, contains two lasers, one to distinguish the spectral signature of each bead and the second to quantify the amount of RPE fluorescence, which is proportional to the amount of protein present in the sample. ProcartaPlex multiplex assays can profile more target proteins using significantly less sample in the same time that it takes to perform a traditional sandwich ELISA.