ISPO

Published in Cancer Detection and Prevention 1995; 19(1).

Accurate quantitation of residual acute lymphoblastic leukemia (ALL) by limiting-dilution PCR

WM Roberts, MV Ouspenskaia, DA Johnston, Z Estrov, TF Zipf

The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA.

The polymerase chain reaction (PCR) can provide a sensitivity for detecting one malignant cell amongst 106 normal cells. Quantitation of rare target sequences, such as occurs in residual leukemia studies, is problematic when single PCR reactions are performed, resulting in estimates with errors as large as one order of magnitude. Our objective was to design a PCR method that generated estimates with narrow error margins. This was achieved by using a limiting-dilution assay measuring the fraction of ten positive (all-or-none) reactions of the PCR amplification of the leukemia-specific IgH gene rearrangement in serial dilutions of DNA obtained from diagnostic bone marrow specimens from 15 children with B-precursor ALL. A best-fit equation was developed for the resulting sigmoid curve that allows estimates that are accurate within 1/4 an order of magnitude when the concentration of leukemia cells is >=10(-5). The method is reliable and exhibits small interspecimen variation. We have applied this technique to a prospective study of residual disease in childhood ALL; the technique, however, can apply to any PCR assay that requires accurate quantitation of the target number with an assessment of relative error probabilities.

Paper presented at the International Symposium on the Impact of Biotechnology on Predictive Oncology and Therapy; Boston, Massachusetts; December 11 - 13, 1994; in the section on Diagnostic Markers.

http://www.cancerprev.org/Journal/Issues/19/1/303/1109