Enhanced protein profiling arrays with ELISA-based amplification for high-throughput molecular changes of tumor patients' plasma.
OBJECTIVE
The purpose of this study is to develop a high-throughput approach to detect protein expression from hundreds and thousands of samples and to apply this technology to profile circulating angiogenic factor protein levels in patients with gynecological tumors.


METHODS
Analytes containing a mixture of protein are immobilized onto antibody-coated surface of support in array format. The presence of protein in analytes is detected with biotin-labeled antibody coupled with an enhanced chemiluminescence or fluorescence detection system. The exact amount of protein can be quantitatively measured. The expression levels of five angiogenic factors (angiogenin, interleukin 8, vascular endothelial growth factor, platelet-derived growth factor, and epidermal growth factor) from 157 samples were quantitatively measured using this novel protein array technology and were statistically analyzed. The expression patterns of angiogenic factors were analyzed using two-way hierarchical cluster analysis approach.


RESULTS
A novel protein array technology, which can simultaneously and quantitatively measure few protein levels from hundreds and thousands of samples was developed. Only minute amounts of sample are required for the assay. This approach also features high sensitivity and specificity. Using this novel protein array approach, we analyzed the plasma expression levels of five angiogenic factors in 137 patients diagnosed with a tumor and 20 controls. Statistical analysis reveals different expression levels of angiogenic factors between patients and controls. Cluster analysis suggests a possible classification of normal subjects from patients.


CONCLUSIONS
Enhanced protein profiling arrays provide a high-throughput and sensitive system to detect one or few protein from hundreds and thousands of samples. Such an approach should have broad application in biomedical discovery.
Huang
Ruochun
R
Department of Gynecology and Obstetrics, Emory University School of Medicine, Atlanta, Georgia 30322, USA. rhuang@emory.edu
Lin
Ying
Y
Shi
Qian
Q
Flowers
Lisa
L
Ramachandran
Sumathi
S
Horowitz
Ira R
IR
Parthasarathy
Sampath
S
Huang
Ruo-Pan
RP
D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001710:Biotin / Q000737:chemistry; D045744:Cell Line, Tumor; D016000:Cluster Analysis; D004797:Enzyme-Linked Immunosorbent Assay; D004815:Epidermal Growth Factor / Q000096:biosynthesis / Q000378:metabolism; D005260:Female; D005833:Genital Neoplasms, Female / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry; D016209:Interleukin-8 / Q000096:biosynthesis / Q000378:metabolism; D008163:Luminescent Measurements; D008856:Microscopy, Fluorescence; D008875:Middle Aged; D005810:Multigene Family; D009369:Neoplasms / Q000378:metabolism; D009389:Neovascularization, Pathologic; D020411:Oligonucleotide Array Sequence Analysis; D010982:Platelet-Derived Growth Factor / Q000096:biosynthesis / Q000378:metabolism; D040081:Protein Array Analysis / Q000379:methods*; D012259:Ribonuclease, Pancreatic / Q000096:biosynthesis; D012680:Sensitivity and Specificity; D014018:Tissue Distribution; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis / Q000378:metabolism
D007074:Immunoglobulin G; D016209:Interleukin-8; D010982:Platelet-Derived Growth Factor; D042461:Vascular Endothelial Growth Factor A; D004815:Epidermal Growth Factor; D001710:Biotin; C046366:angiogenin; D012259:Ribonuclease, Pancreatic