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Chemiluminescent restriction fragment length polymorphism analysis (DNA fingerprinting) to identify keratinocytes from two donors in co‐culture
Author(s) -
Young David M.,
Summers William C.,
Cuono Charles B.
Publication year - 1993
Publication title -
experimental dermatology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.108
H-Index - 96
eISSN - 1600-0625
pISSN - 0906-6705
DOI - 10.1111/j.1600-0625.1993.tb00195.x
Subject(s) - restriction fragment length polymorphism , dna , biology , transplantation , microbiology and biotechnology , cell culture , restriction fragment , chimera (genetics) , chemiluminescence , dna profiling , heterologous , genetics , chemistry , medicine , polymerase chain reaction , gene , chromatography , surgery
The transplantation of allogeneic cells such as pancreatic islet cells, bone marrow, and keratinocytes for skin replacement is in growing use clinically. Present techniques which are used to distinguish transplanted allogeneic cells from phenolypically identical host cells in chimeric tissue have serious limitations. This study assesses the feasibility of using Chemiluminescent restriction fragment length polymorphism (RFLP) analysis to distinguish between keratinocyte cell lines from 2 different donors grown in co‐culture. We evaluated the sensitivity of this technique in tissue cultured cells in order to define its usefulness in clinical situations. RFLP analysis readily detects heterologous DNA comprising only 10% of a 5 μg sample. This was a detection threshold of 500 ng of high molecular wefght DNA. With standard extraction methods, this represents the DNA obtained from 4000 cells. This new chemiluminescent technique is as sensitive as older techniques which employ radioactive probes, but avoids the hazards of handling radioactive material. Using this RFLP analysis, the presence of transplanted allogeneic cells can be readily delected in small biopsy specimens. This technique has wide potential application in allogeneic cellular transplantation investigations.