Patient plasma samples
A library of plasma samples were collected from all patients attending one general oncology clinic for postoperative follow up of breast cancer; samples were taken without any patient selection and irrespective of clinical stage, menopausal status, histological type or degree of differentiation. The operative management of each patient was according to the tenets of their surgeons at the time of presentation; the breast was conserved wherever possible. The majority of patients were treated with adjuvant radio-therapy. Adjuvant cytotoxic chemotherapy, with or without tamoxifen, was the subject of clinical trial at the time.
The clinical status of each patient at time of sampling had been summarized on a database using Helix Express (version 1.0.1; Helix Technologies, Northbrook, IL, USA). The plasma library had been established over a 15-year period, with an average of eight plasma samples per patient being cryopreserved. It is known that cryopreserved p53 autoantibodies are stable in either sera or plasma, and that several cycles of freezing/thawing do not cause loss of titre (Soussi T, personal communication). Although it is theoretically possible that shed p53 protein might partially block autoantibody in some patients, it has already been shown [17] that autoantibody to p53 is normally in excess, and is thus detectable by ELISA.