Background: The ductal/alveolar system of the female breast constantly secretes and reabsorbs fluid in nonpregnant/nonlactating women. This fluid, referred to as nipple aspirate fluid (NAF), can be obtained by a noninvasive procedure and it is part of the microenvironment where more than 95% of breast cancers arise.
Methods: Using an Orbitrap mass analyzer coupled to a linear ion trap, we performed an in-depth proteomic analysis of NAF samples obtained from 3 healthy individuals and 3 patients with breast cancer. Multiple fractionation methods such as size-exclusion and anion-exchange chromatography were applied for protein separation before mass spectrometric analysis.
Results: We identified more than 800 unique proteins in total, generating the most extensive NAF proteome thus far. Using gene ontology, we classified the identified proteins by their subcellular localization and found that more than 50% were extracellular or plasma membrane proteins. By searching against the Plasma Proteome Database, we confirmed that 40% of the proteins were also found in the plasma. Unigene database searching for transcripts of the proteins not found in the plasma revealed that the vast majority were expressed in the mammary gland.
Conclusions: Our extensive proteome database for NAF may be helpful in the identification of novel cancer biomarkers.