This Article Full Text Full Text (PDF) Purchase Article View Shopping Cart Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Gaffney, R. Articles by Viswanatha, D. S. Search for Related Content PubMed PubMed Citation Articles by Gaffney, R. Articles by Viswanatha, D. S.

JMD 2003, Vol. 5, No. 2
Copyright © 2003 American Society for Investigative Pathology & Association for Molecular Pathology

Novel Fluorescent Ligase Detection Reaction and Flow Cytometric Analysis of SYT-SSX Fusions in Synovial Sarcoma

Robyn Gaffney^*, Artemis Chakerian^*, John X. O’Connell, Joan Mathers, Kelly Garner, Nancy Joste^* and David S. Viswanatha^*

From the University of New Mexico Health Sciences Center, * Albuquerque, New Mexico; Surrey Memorial Hospital, Surrey, British Columbia, Canada; Children’s and Women’s Health Centre of British Columbia, Vancouver, British Columbia, Canada; and Tricore Reference Laboratories, Albuquerque, New Mexico

Synovial sarcomas (SS) are characterized by the t(X;18)(p11;q11) translocation and its resultant fusion gene, SYT-SSX. Two homologues of the SSX gene (ie, SSX1 and SSX2) are involved in the vast majority of SS and the SYT-SSX1 type of fusion has been associated with inferior clinical outcome. Thus, detection of the presence and type of SYT-SSX fusion is critical for diagnosis and prognosis in SS. Identification of SYT-SSX fusion type is typically accomplished by reverse-transcription polymerase chain reaction (RT-PCR) followed by a post-PCR analytic method. As mRNA nucleotide sequences of the SSX1 and SSX2 segments involved in the SYT-SSX fusion are nearly identical, post-PCR methods must be highly discriminatory. We describe a novel method to identify and differentiate these two chimeric transcripts using RT-PCR followed by fluorescent thermostable ligase detection reaction (f-LDR), microparticle bead capture and flow cytometric detection. Evaluation of this unique approach in 11 cases of SS without prior knowledge of SYT-SSX status, six cases of control sarcomas (CS) and three hematopoietic cell lines, revealed that the f-LDR technique was rapid, unambiguous, and highly specific. The f-LDR results were compared to XmnI enzyme digestion patterns and sequencing of PCR products, revealing a 100% concordance for all cases of SS with regards to SYT-SSX transcript type. In addition, there was a strong association of transcript type detected by f-LDR and morphological subclassification of SS, as previously reported. We conclude that this f-LDR method with flow-based detection is a robust approach to post-PCR detection of specific nucleotide sequences in SS and may be more broadly applicable in molecular oncology.