A comparison of methods for the analysis of low abundance proteins in desmoid tumor cells.
The desmoids are a group of rare clinically diverse, deep-seated fibrous neoplasms. The exact etiology is unknown, but several factors are considered to be positively correlated with their development and growth, i.e., genetic and hormonal factors and trauma. These tumors may be sporadic or associated with a genetic disease such as familial adenomatous polyposis (FAP). Devoid of metastatic potential, they tend to form large, infiltrative masses which, if not completely excised, recur repeatedly. Although surgery is widely accepted as the first-line treatment for extra-abdominal and abdominal wall desmoids, a proportion of cases are successfully palliated with either estrogen antagonists (tamoxifen, toremifene, and raloxifene) or nonsteroidal anti-inflammatory drugs. We describe and compare four methods for evaluating the expression of estrogen receptors alpha/beta and COX-1 and COX-2 in desmoid tumor-derived cells and tissues: immunocytochemistry, immunohistochemistry, RT-PCR, and two-color Western blot detection with the Odyssey infrared imaging system. Through this comparative analysis, Western blot with Odyssey was recognized as the best method to analyze the expression particularly of low expressed proteins in desmoid-derived cells. The use of a specific and reliable assessment method becomes fundamental in the evaluation of the presence and modulation of proteins which are important but weakly expressed in these rare tumors.
Picariello
L
L
Department of Clinical Physiopathology, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy.
Carbonell Sala
S
S
Martineti
V
V
Gozzini
A
A
Aragona
P
P
Tognarini
I
I
Paglierani
M
M
Nesi
G
G
Brandi
M L
ML
Tonelli
F
F
D001483:Base Sequence; D015153:Blotting, Western; D051545:Cyclooxygenase 1 / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D051546:Cyclooxygenase 2 / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D047628:Estrogen Receptor alpha / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D047629:Estrogen Receptor beta / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D018222:Desmoid Tumors / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D007150:Immunohistochemistry; D008565:Membrane Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D009363:Neoplasm Proteins / Q000032:analysis* / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012334:RNA, Neoplasm / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
D047628:Estrogen Receptor alpha; D047629:Estrogen Receptor beta; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; C496511:PTGS1 protein, human; C496540:PTGS2 protein, human