Profiling the expression of mitogen-induced T-cell proteins by using multi-membrane dot-blotting.
High throughput technologies are standard methods for analysis of the proteome. Multi-layer multi-well plate dot-blotting system (MLDot) technology is a high-throughput dot blotting system that provides a simple, cost-effective approach for protein expression profiling in multiple samples. In contrast to traditional dot blot, MLDot uses a layered stack of thin, sieve-like membranes in place of a single nitrocellulose membrane. Therefore, up to 10 membranes can be prepared from the samples arrayed in a single 96-well plate. We describe the ability of MLDot to detect the predicted changes in protein expression following multiple mitogen treatment of T-cells. We compare the levels of the phopshorylated forms of CREB, Jun, and Akt in Jurkat T-cells as detected by MLDot to those measured by a gel-based assay. We also describe the ability of MLDot to detect differences in the levels of phosphorylated Akt in Jurkat cells as compared to primary lymphocytes.
Traicoff
June L
JL
20/20 Gene Systems, Inc., Rockville, MD, USA.
Galperin
Mikhail M
MM
Ramesh
Arun
A
Freebern
Wendy J
WJ
Haggerty
Cynthia M
CM
Gardner
Kevin
K
Knezevic
Vladimir
V
D019921:Biotinylation; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D015195:Drug Design; D006801:Humans; D019169:Jurkat Cells; D008214:Lymphocytes / Q000378:metabolism; D008934:Mitogens; D010766:Phosphorylation; D040081:Protein Array Analysis / Q000379:methods*; D025941:Protein Interaction Mapping; D020543:Proteome; D015398:Signal Transduction; D013050:Spectrometry, Fluorescence; D013601:T-Lymphocytes / Q000378:metabolism
D008934:Mitogens; D020543:Proteome