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Vet Pathol 42:596-607 (2005)
© 2005 American College of Veterinary Pathologists

Characterization of Feline Immunoglobulin Heavy Chain Variable Region Genes for the Molecular Diagnosis of B-cell Neoplasia

J. A. Werner, J. C. Woo, W. Vernau, P. S. Graham, R. A. Grahn, L. A. Lyons and P. F. Moore

Departments of Veterinary Pathology, Microbiology, and Immunology (JAW, WV, PFM) and Population Health and Reproduction (RAG, LAL), School of Veterinary Medicine, University of California, Davis, CA; and MedImmune Vaccines Inc., Mountain View, CA (JCW, PSG)

To develop a molecular-based assay so that the diagnosis of feline B-cell neoplasia can be facilitated, we have characterized 24 feline immunoglobulin heavy chain variable region (IGH V) complementary DNA (cDNA) transcripts. Structural homology with rearranged human IGH V genes was found, and the sequence information was used to design a feline-specific polymerase chain reaction (PCR)-based assay to amplify the complementarity determining region 3 as a marker for B-cell clonality. Conserved primers derived from the second and third framework regions of V gene segments were used in conjunction with 2 sequence-specific primers and 1 degenerate primer derived from the J gene segments. Each PCR reaction was run in duplicate, and both native and denatured PCR products were evaluated using polyacrylamide gel electrophoresis. Formalin-fixed, paraffin-embedded (FFPE) tissue sections from cats with confirmed B-cell neoplasia (diffuse large B-cell lymphoma, plasmacytoma, and myeloma) were examined, and 15/22 (68.2%) cats produced results indicative of the presence of a monoclonal population of B cells. The evaluation of denatured PCR products (heteroduplex analysis) facilitated a more accurate interpretation in 3/15 (20%) cats. Pseudoclonality was a major reason for the failure to detect monoclonality. Poor DNA quality is a significant concern and was responsible for the removal of 2 cats from the study. Using this assay, FFPE normal feline lymphoid tissues and unfixed peripheral blood mononuclear cells were determined to be composed of polyclonal populations of B cells. This assay represents a useful adjunctive diagnostic tool for the diagnosis and investigation of feline B-cell lymphoproliferative disorders.

Key words: Cats; clonality; complementarity determining region 3; heteroduplex analysis; immunoglobulin heavy chain gene; lymphoma; myeloma; plasmacytoma; polymerase chain reaction.

Request reprints from Dr. P. F. Moore, Department of Pathology, Immunology, and Microbiology, School of Veterinary Medicine, University of California, Davis, CA 95616 (USA). E-mail: pfmoore{at}ucdavis.edu.






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