Methyl-binding domain protein-based DNA isolation from human blood serum combines DNA analyses and serum-autoantibody testing

Serum or plasma samples from 98 control individuals and 54 breast cancer patients were evaluated upon silica membrane- or MBD affinity-based DNA isolation via qPCR targeting potential DNA methylation markers as well as by protein-microarrays for tumor-autoantibody testing.In control individuals, an average DNA level of 22.8 ± 25.7 ng/ml was detected applying the silica membrane based protocol and 8.5 ± 7.5 ng/ml using the MBD-approach, both values strongly dependent on the serum sample preparation methods used. In contrast to malignant and benign tumor serum samples, cell free DNA concentrations were significantly elevated in sera of metastasizing breast cancer patients. Technical evaluation revealed that serum upon MBD-based DNA isolation is suitable for protein-array analyses when data are consistent to untreated serum samples.MBD affinity purification allows DNA isolations under native conditions retaining the protein function, thus for example enabling combined analyses of DNA methylation and autoantigene-profiles from the same serum sample and thereby improving minimal invasive diagnostics.The prognostic potential of cell free nucleic acids in plasma and serum caused a lot of attention in current efforts to improve diagnoses of several cancerous diseases [1,2]. During the last years various approaches in this field, like the determination of genetic aberrations, the methylation status of the DNA [3], DNA level [4] and DNA-integrity [5] as well as miscellaneous proteomic approaches [6] have been reported to have clinical relevance. Despite this, breast cancer diagnosis still relies on physical examinations and mammography, because no accurate and reliable approach is currently available to be implemented into an effective diagnostic test. Therefore, the need for a simple non invasive blood test to perform routine pre-symptomatic screenings that improve early diagnosis is high.Serum markers for breast cancer detection may be divided into two areas. On the one han