![]() Of note, sample sources for SBH analysis are limited to fresh or frozen cells, owing to the requirement for high-quality, long-strand genomic DNA. The multi-step, technically demanding requirements of SBH also increase the possibility of analytic delays, because errors in procedure are seldom identified before the blot is finally exposed. Visual interpretation of exposed blots is usually straightforward however, occasional difficulties may arise owing to degraded DNA quality, incomplete enzymatic digestion of DNA, or when novel pathologic bands co-migrate with germline fragments. SBH has been most widely applied to detect clonotypic rearrangements of the antigen receptor genes in abnormal B- or T-cell lymphoid proliferations, but assays to evaluate specific oncogenes (e.g., KMT2A, BCL2, CCND1) have also been described. Exposure of the resultant band pattern on radiographic film reveals the presence of any atypical rearrangement(s) at the genetic locus in question, relative to the expected or “germline” (unaltered) band configuration. A particular region of immobilized DNA can then be interrogated by stringent hybridization with a specific double-stranded genomic probe, which has been labeled with a radioisotope (e.g., P 32) or by a non-isotopic reagent (e.g., for chemiluminescent detection). SBH involves digestion of high-molecular-weight DNA with site-specific restriction endonucleases, followed by size separation of the DNA by gel electrophoresis and transfer of the nucleic acid fragments onto a nylon or nitrocellulose membrane. Southern blot hybridization (SBH) technique occupies a diminishing role in the molecular diagnostic laboratory but continues to have relevance for the analysis of relatively large (kilobase)-scale alterations in genomic DNA. Viswanatha MD, in Hematopathology (Third Edition), 2018 Southern Blot Hybridization ![]()
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