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Validation of immunoglobulin gene rearrangement detection by PCR using commercially available BIOMED-2 primers

Leukemia volume 20, pages 176–179 (2006)Cite this article

The BIOMED-2 primers are now commercially available for clinical use but a recent search of the English language literature revealed essentially no expansion of the initial validation studies such that laboratories introducing the primers must rely on data from the relatively few samples tested exclusively by the laboratories of the BIOMED-2 group prior to commercial production. We have performed independent validation of BIOMED-2 primers for IG gene rearrangement analysis in anticipation of primer use for routine clinical laboratory testing.

Clinical sensitivity and specificity data is summarized in Table 1. Sensitivity for non-Hodgkin neoplasms ranged from 82 to 100% depending on tumor and specimen type. The diagnostic categories with <100% sensitivity were plasma cell neoplasms (95%), mantle cell lymphoma (86%), and diffuse large B-cell lymphoma (82%), none of which commonly pose a diagnostic dilemma requiring molecular evaluation. The cause of less than 100% sensitivity in these categories is not entirely clear. Less than perfect assay precision could be a factor in some cases, particularly if there should happen to be the combination of positivity in a single size range and poor amplification. When formally tested, precision was adequate but not 100%, as there was some inter-run variation in peak heights. However, this did not interfere with the result status of any of these cases and all negative cases in the study were retested with identical negative results. Thus, we do not feel routine duplicate testing is warranted, although it may be helpful in cases interpreted as equivocal on initial testing. Of course it is essential that adequate and representative tissue be tested to ensure optimal sensitivity and despite our attempts to control for this variable, it could still be a contributing factor, particularly in the plasma cell neoplasms where patchy tissue involvement is common. Clinical sensitivity was not significantly different between neoplasms considered prefollicular (expected to have no somatic hyper mutation) and those considered follicular or postfollicular (expected to have somatic hyper mutation). This indicates that the approach of analyzing multiple genes with multiple primers largely overcomes the problem of primer incompatibility. Further support for this concept comes from the surprising result of moderate clinical sensitivity (35%) using the primer sets to evaluate Hodgkin lymphoma. Although the neoplastic cells in both nodular lymphocyte predominant and classical Hodgkin lymphoma contain rearranged IG genes,6, 7 they contain extensive somatic hypermutation and represent a small percentage of the total cells in most samples such that clinical laboratory clonality studies are usually negative.

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  1. Division of Hematopathology, Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN, USA

    R F McClure, P Kaur, E Pagel, P D Ouillette, C E Holtegaard, C L Treptow & P J Kurtin

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  1. R F McClure
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  2. P Kaur
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McClure, R., Kaur, P., Pagel, E. et al. Validation of immunoglobulin gene rearrangement detection by PCR using commercially available BIOMED-2 primers. Leukemia 20, 176–179 (2006). https://doi.org/10.1038/sj.leu.2404049

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