Analysis of melanoma cells in peripheral blood by reverse transcription-polymerase chain reaction for tyrosinase and MART-1 after mononuclear cell collection with cell preparation tubes: a comparison with the whole blood guanidinium isothiocyanate RNA isolation method.

Melanoma cell detection in peripheral blood by tyrosinase reverse transcription-polymerase chain reaction (RT-PCR) is usually performed on RNA isolated from whole blood using a guanidinium isothiocyanate (GITC)/phenol extraction method or from Ficoll Hypaque isolated mononuclear cells. The first method contains environmentally harmful reagents, and the second is laborious in the preanalytical steps. Cell preparation tubes (CPTs) are ready-to-use Ficoll Hypaque-based tubes that avoid the time-consuming and critical loading on Ficoll Hypaque.

Melanoma-inhibiting activity (MIA) mRNA is not exclusively transcribed in melanoma cells: low levels of MIA mRNA are present in various cell types and in peripheral blood.

The detection of minimal amounts of melanoma cells by tyrosinase reverse transcription polymerase chain reaction (RT-PCR) is seriously hampered by false negative reports in blood of melanoma patients with disseminated melanoma. Therefore, additional assays which make use of multiple melanoma markers are needed. It has been shown that introduction of multiple markers increases the sensitivity of detection.

Transcription of the MAGE-1 gene and the methylation status of its Ets binding promoter elements: a quantitative analysis in melanoma cell lines using a real-time polymerase chain reaction technique.

The human MAGE gene family comprises at least 12 highly homologous genes. This makes it very difficult to assess expression of a single member quantitatively by means of Northern blotting. In order to investigate expression of the MAGE-1 gene quantitatively we therefore used the recently developed real-time polymerase chain reaction (PCR), a novel fluorescence-based quantitative PCR technique.

Reproducibility of detection of tyrosinase and MART-1 transcripts in the peripheral blood of melanoma patients: a quality control study using real-time quantitative RT-PCR.

In recent years, large discrepancies were described in the success rate of the tyrosinase reverse transcription polymerase chain reaction (RT-PCR) for detecting melanoma cells in the peripheral blood of melanoma patients. We present a quality control study in which we analysed the reproducibility of detection of tyrosinase and MART-1 transcripts in 106 blood samples from 68 melanoma patients (mainly stages III and IV). With this study, we aimed to improve insight in the reproducibility of a RT-PCR for the detection of (minimal) amounts of circulating melanoma cells.

Heterogeneous expression of immunotherapy candidate proteins gp100, MART-1, and tyrosinase in human melanoma cell lines and in human melanocytic lesions.

In recent years, it has become evident that T cells can recognize peptides of melanocytic lineage antigens such as gp100, MART-1, and tyrosinase at the tumor cell surface and can subsequently destroy these cells. It is thus feasible to develop immunotherapeutic approaches based on the melanocytic marker profiles of melanoma cells. One of the predictors of the success rate of such a treatment is the extent of positive (target) tumor cells within the lesions of the patient.