Infrared-based protein detection arrays for quantitative proteomics.
The advancement of efficient technologies to comply with the needs of systems biology and drug discovery has so far not received adequate attention. A substantial bottleneck for the time-resolved quantitative description of signaling networks is the limited throughput and the inadequate sensitivity of currently established methods. Here, we present an improved protein microarray-based approach towards the sensitive detection of proteins in the fg-range which is based on signal detection in the near-infrared range. The high sensitivity of the assay permits the specific quantification of proteins derived from as little as only 20,000 cells with an error rate of only 5%. The capacity is limited to the analysis of up to 500 different samples per microarray. Protein abundance is determined qualitatively, and quantitatively, if recombinant protein is available. This novel approach was called IPAQ (infrared-based protein arrays with quantitative readout). IPAQ offers a highly sensitive experimental approach superior to the established standard protein quantification technologies, and is suitable for quantitative proteomics. Employing the IPAQ approach, a detailed analysis of activated signaling networks in biopsy samples and of crosstalk between signaling modules as required in drug discovery strategies can easily be performed.
Loebke
Christian
C
Division of Molecular Genome Analysis, German Cancer Research Center, Heidelberg, Germany.
Sueltmann
Holger
H
Division of Molecular Genome Analysis, German Cancer Research Center, Heidelberg, Germany.
Schmidt
Christian
C
Division of Molecular Genome Analysis, German Cancer Research Center, Heidelberg, Germany.
Henjes
Frauke
F
Division of Molecular Genome Analysis, German Cancer Research Center, Heidelberg, Germany.
Wiemann
Stefan
S
Division of Molecular Genome Analysis, German Cancer Research Center, Heidelberg, Germany.
Poustka
Annemarie
A
Division of Molecular Genome Analysis, German Cancer Research Center, Heidelberg, Germany.
Korf
Ulrike
U
Division of Molecular Genome Analysis, German Cancer Research Center, Heidelberg, Germany.
D045744:Cell Line, Tumor; D048049:Extracellular Signal-Regulated MAP Kinases / Q000032:analysis; D005260:Female; D006801:Humans; D007259:Infrared Rays; D040081:Protein Array Analysis / Q000379:methods*; D040901:Proteomics; D011994:Recombinant Proteins / Q000032:analysis*; D012015:Reference Standards; D012680:Sensitivity and Specificity
D011994:Recombinant Proteins; D048049:Extracellular Signal-Regulated MAP Kinases