Plasma was stored as 250-1000 μl aliquots at -80°C
until assayed. Ovarian tumor classification was based on the FIGO staging NVP-BSK805 research buy system, however, no stage IV tumors were identified for inclusion in this study. Study Design The study was a retrospective, case-control design (i.e. a phase 2 biomarker trial  involving 107 plasma samples (see Table 1) obtained from 61 controls and 46 cases (i.e. women previously diagnosed with ovarian cancer). Inclusion and exclusion criteria are presented in Table 2. The primary outcomes of the study were: quantification of plasma concentrations of ir MDK and evaluation of its diagnostic performance (as defined by AUC); and comparison with AGR2 and CA125 concentrations measured in the same cohort. In addition, the contribution for these biomarkers to multi-analyte classification models was determined. Table 1 Distribution of Ovarian Tumor Types and Stages of ovarian cancer patients used for plasma AGR2 and CA125 measurements. All Tumors Stage I Stage II Stage III Unstaged Serous 29 3 17 9 Mucinous 5 1 3 1 Endometrioid 4 2 2 Clear Cell 2 1 1 Mixed Mullerian 3 1 2 Untyped 3 2 1 Total 46 9 26 10 1 Table 2 Inclusion and exclusion criteria for inclusion of patient samples in the Erismodegib research buy study. Inclusion Criteria Exclusion Criteria Age 18-80 Chemotherapy, biologic therapy or any other investigational drug for any reason within 28 days prior to sampling.
Newly diagnosed, histologically or pathologically confirmed diagnosis of
epithelial carcinoma of the ovary. Except for cancer-related abnormalities, patients should not have unstable or pre-existing major medical conditions. No NSAID or CP-690550 molecular weight prednisone use within 14 days of sampling. Major surgical procedure, open biopsy, or significant traumatic injury within 28 days prior to sampling No previous chemotherapy or radiation therapy. Minor surgical procedures, fine needle aspirations or core biopsies within 7 days prior to sampling No concurrent disease(s) Serious, non-healing wound, ulcer, or bone Signed informed client consent Biomarker Quantitation Plasma concentrations of MDK were quantified by sandwich ELISA assay (Peprotech, Rocky Hill, NJ, USA) that utilises rabbit antibodies raised to human midkine for both capture and detection stages of the assay. Reverse transcriptase The assay was performed in Costar half-well immunoassay plates (Corning) coated with 50 μl of capture antibody at a concentration of 1 ug/ml in 50 mM carbonate buffer and incubated at 4°C for 18 h. Following four washes in PBS/Tween20 (Sigma), the plate was blocked with 150 μl/well of 0.1% BSA (Sigma) in PBS, for one hour at room temperature. Plasma samples diluted 1:2 in PBS containing 0.05% Tween20 and 0.1% BSA were applied to the plate following blocking, alongside a standard curve, from 2000 pg/ml down in doubling dilutions, constructed from a stock recombinant midkine.