During the period 2007 to 2009, four PAX5-positive anaplastic large cell lymphomas were identified from the hematopathology practice at Mayo Clinic, Rochester, Minnesota; 198 additional peripheral T-cell lymphomas from the years 1987 to 2009 identified from the Mayo Clinic archives were studied. All cases were classified based on 2008 WHO criteria.1
PAX5 immunohistochemistry was performed on paraffin embedded tissue sections by pretreating in 1mM EDTA buffer at pH 8.0 for 30 min at 97°C (PT Module; Lab Vision, Fremont, CA) and staining for PAX5 (1:200, clone 24, BD Bioscience) on a Dako (Carpinteria, CA) autostainer using the Advance detection system (Dako) with diaminobenzidine as the chromogen. Immunohistochemistry for other markers was performed as previously described35 (link) using antibodies shown in Table 1. Aside from CD30 and PAX5 (discussed below), immunostaining was scored as strong or weak, and designated as negative (-, no staining), focal (-/+, <10% of tumor cells), partial (+/-, 10-30% of tumor cells), or positive (+, >30% of tumor cells).
Polymerase chain reaction (PCR) for T-cell receptor (TCR) γ-chain and immunoglobulin gene rearrangements was performed as described previously.36 (link),37 (link) FISH for PAX5 was performed and scored as described previously using a homebrew breakapart probe.38 Briefly, DNA was isolated from bacterial artificial chromosome probes (ResGen - Invitrogen; Carlsbad, CA) spanning the PAX5 locus as shown in Fig. 3c. Probes were labeled with SpectrumOrange-dUTP or SpectrumGreen-dUTP by nick translation (Abbott Molecular, Des Plaines, IL) and hybridized to tissue sections. Cases with ≥4 fusion signals were considered to have extra copies of the PAX5 gene locus.
Additional peripheral T-cell lymphomas were evaluated by immunohistochemistry and/or FISH as indicated above on tissue microarrays constructed from paraffin blocks as previously described.39 (link) The study was approved by the Mayo Clinic Institutional Review Board and Biospecimens Committee.