SEMIQUANTITATIVE POLYMERASE CHAIN-REACTION FOR T(14 18) IN FOLLICULARLYMPHOMAS - A COLORIMETRIC APPROACH/

BACKGROUND: Autologous bone marrow transplantation is increasingly bei ng used in the management of several types of cancer, and to avoid rei ntroduction of malignant cells, bone marrow purging is often performed . In such cases, sensitive quantitation methods are needed both to ass ess the efficacy of the purging and for surveillance of patients in re mission. Polymerase chain reaction (PCR) has the necessary sensitivity for this application, but it requires that the cancer cells can be re cognized by a defined genetic abnormality. In addition, PCR is in prin ciple a qualitative technique and must be modified for quantitative pu rposes. In follicular non-Hodgkin's lymphomas, the translocation t(14; 18) (q32;q21) is common and is used here for model experiments. EXPERI MENTAL DESIGN: A PCR-based method for the quantitation of translocatio n-positive cells was developed on the basis of coamplification of canc er-specific target molecules with competitor molecules of known concen tration. Gel electrophoresis was substituted by a colorimetric quantit ation system to cope with patient PCR products of the same size as the competitor product and ease automation of the method at a later stage . Cell line Karpas 422, which contains the t(14;18) (q32;21) transloca tion, was used to validate the method. The method was used to assess t he efficacy of a patient bone marrow purging where the initial infiltr ation levels were too low for traditional detection systems. RESULTS: A reproducible and near linear response was obtained between 70 pg and 200 ng K422 DNA, equivalent to 10 K422 cells and 30,000 K422 cells, r espectively. Bone marrow infiltration in one patient was 0.6 to 0.7% b efore malignant cell removal and 3 to 7 x 10(-4) after removal. The co rresponding figures for the other patient were 2% and 3 to 7 x 10(-4), respectively. CONCLUSIONS: The method presented has a sufficient dyna mic range for applications like evaluation of bone marrow purging or m onitoring of minimal residual disease. Adaptation of this method to ot her translocations is discussed.