Dual-color proteomic profiling of complex samples with a microarray of 810 cancer-related antibodies.
Antibody microarrays have the potential to enable comprehensive proteomic analysis of small amounts of sample material. Here, protocols are presented for the production, quality assessment, and reproducible application of antibody microarrays in a two-color mode with an array of 1,800 features, representing 810 antibodies that were directed at 741 cancer-related proteins. In addition to measures of array quality, we implemented indicators for the accuracy and significance of dual-color detection. Dual-color measurements outperform a single-color approach concerning assay reproducibility and discriminative power. In the analysis of serum samples, depletion of high-abundance proteins did not improve technical assay quality. On the contrary, depletion introduced a strong bias in protein representation. In an initial study, we demonstrated the applicability of the protocols to proteins derived from urine samples. We identified differences between urine samples from pancreatic cancer patients and healthy subjects and between sexes. This study demonstrates that biomedically relevant data can be produced. As demonstrated by the thorough quality analysis, the dual-color antibody array approach proved to be competitive with other proteomic techniques and comparable in performance to transcriptional microarray analyses.
Schröder
Christoph
C
Division of Functional Genome Analysis, Deutsches Krebsforschungszentrum, 69120 Heidelberg, Germany. christoph.schroeder@dkfz.de
Jacob
Anette
A
Tonack
Sarah
S
Radon
Tomasz P
TP
Sill
Martin
M
Zucknick
Manuela
M
Rüffer
Sven
S
Costello
Eithne
E
Neoptolemos
John P
JP
Crnogorac-Jurcevic
Tatjana
T
Bauer
Andrea
A
Fellenberg
Kurt
K
Hoheisel
Jörg D
JD
D000912:Antibodies, Neoplasm / Q000097:blood / Q000276:immunology* / Q000652:urine; D001681:Biological Assay; D016022:Case-Control Studies; D003116:Color; D006801:Humans; D009369:Neoplasms / Q000097:blood / Q000276:immunology*; D010190:Pancreatic Neoplasms / Q000097:blood / Q000276:immunology / Q000652:urine; D040081:Protein Array Analysis / Q000379:methods*; D020543:Proteome / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D011786:Quality Control
D000912:Antibodies, Neoplasm; D020543:Proteome