Abstract
Retinoic acid receptor beta (RAR-beta) seems to be a useful intermediate marker in trials of retinoids, and the aim of this work was to describe a fast, sensitive, and routine applicable method to measure RAR-beta expression in tumor biopsies.
We developed a new technique combining reverse transcription-PCR with a colorimetric ELISA detection of amplification products. The principle of this nonradioactive method is based on digoxigenin labeling of PCR products during amplification.
Amplified DNA is hybridized with a biotinylated capture probe. The generated hybrid is immobilized on a streptavidin-coated microtiter plate, and detection is performed with the use of an antidigoxigenin peroxidase conjugate. We applied this method to quantify the expression of RAR-beta and an internal control (beta2 microglobulin) in laryngeal tumors. We found a detection threshold at 50 pg of PCR products, which represents a 100-fold improvement when compared to the detection limit of ethidium bromide detection.
The method was reproducible (intra- and interassay reproducibilities at 7 and 5%, respectively). We used this technique for determining RAR-beta expression in 20 patients with laryngeal carcinoma and in 20 patients without cancer.
The data show that the value of the RAR-beta:beta2 microglobulin ratio is decreased in tumoral versus nontumoral specimens (P = 0.0012), which is consistent with previously published results.
See also:
- All-Trans-Retinoic Acid (ATRA - analogues and/or derivatives);
- Solution of retinoids in vitamin E in the Di Bella Method biological multitherapy;
- The Di Bella Method (A Fixed Part - All-Trans Retinoic Acid, Analogues and/or Derivatives);
- The Di Bella Method (A Fixed Part - Alpha tocopheryl acetate/Vitamin E);
- Neuroblastoma: Complete objective response to biological treatment;
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