Fluorescent Assay Determines Free DNA in Cancer Patient's Serum

An assay has been developed that measures circulating cell-free DNA in the serum of patients with cancer using a fluorochrome and without extraction and amplification.

The assay is simply performed by adding diluted fluorochrome to the samples and measuring the fluorescence obtained. The assay is accurate, sensitive, and reproducible.

The assay was developed at the Soroka University Medical Center, (Be'er Sheva, Israel) and compared with a commercial kit for carcinoembryonic antigen (CEA). The innovative fluorescent assay was first evaluated on a mouse model and then tested on colorectal cancer (CRC) patients' serum. There were 38 unselected patients with confirmed primary CRC recruited among whom 55% were women and 45% were men. The age range was 43-86 years, and the mean age was 68 years. The primary colon carcinoma sites were as follows: right colon, sigmoid colon, left colon, rectum, and simultaneous carcinoma in the right and left colon. Serum from healthy volunteers was also tested. The concentration of CEA was determined by using the ARCHITECT CEA assay (Abbott Laboratories, Abbott Park, IL, USA).

The cell-free DNA (CFD) assay used diluted SYBR Gold Nucleic Acid Gel Stain (Invitrogen; Carlsbad, CA, USA), which was added to the serum samples and the resulting fluorescence measured with a 96-well fluorometer. Concentrations of unknown samples were calculated from a DNA standards curve by extrapolation in a linear regression model. The 38 patients with CRC had higher preoperative CFD levels of 798 ± 409 ng/mL, when compared with healthy subjects with CFD levels of 308 ± 256 ng/mL. CFD levels were elevated in patients who remained with the disease or died as compared with patients free of disease at one year.

Colorectal cancer (CRC) is the third most common type of cancer worldwide, with an estimated one million new cases and a half million deaths each year. Early diagnosis is fundamental in reducing morbidity and mortality, with patients diagnosed at early stages demonstrating increased long-term survival. The authors concluded that by using a simple fluorometric assay, they were able to find elevated CFD levels in cancer patients, confirming the findings of previous studies that were performed with a complex and time-consuming method. The study was published in February 2011, in the American Journal of Clinical Pathology.

Related Links:
Soroka University Medical Center
Invitrogen
Abbott Laboratories