Skip to main content
PMC home page
Sarcoma logoLink to Sarcoma
. 1999 Mar;3(1):25–32. doi: 10.1080/13577149977839

Detection of EWS/FLI-1 by Immunostaining. An Adjunctive Tool in Diagnosis of Ewing's Sarcoma and Primitive Neuroectodermal Tumour on Cytological Samples and Paraffin-Embedded Archival Material

Gunnar Nilsson 1,2, Min Wang 1, Johan Wejde 1, Andris Kreicbergs 2, Olle Larsson 1,✉
PMCID: PMC2395406  PMID: 18521261

Abstract

Purpose. Recently we showed that the 68-kDa fusion protein derived from the EWS/FLI1 hybrid gene can be specifically detected by Western blotting using a polyclonal antibody to the C-terminal of FLI1 on biopsy material from Ewing's sarcoma. The aim of this study was to investigate whether this antibody also could be used for immunocytochemistry and immunohistochemistry in diagnosis of Ewing's sarcoma.

Methods. Immunostaining on paraffin-embedded archival material, fine-needle aspirates and tumour touch imprints from Ewing's sarcomas and primitive neuroectodermal tumours (PNET) for detection of the fusion protein was performed. Most cases were also analysed by Western blotting.Tumours of differential diagnostic importance were also included.

Results. Eighty per cent (12/15 cases) of the Ewing tumours exhibited a positive immunoreactivity for the FLI1 antibody. The signal was mainly localised in the nuclei of the tumour cells, which seems reasonable since EWS/FLI1 is a transcription factor. The signal was found to be specific since it did not appear when the blocking peptide was added to the antibody solution.Moreover, two other types of small-round cell tumours (i.e. neuroblastoma and alveolar rhabdomyosarcoma) were negative as well as most normal tissues.

Discussion. Immunostaining of histological and cytological specimens with the FLI1 antibody can be of diagnostic relevance in Ewing tumours carrying t(11;22).The absence of immunoreactivity in non-Ewing cells is most likely due to a low expression of the wild-type FLI1 protein.

Full Text

The Full Text of this article is available as a PDF (728.5 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  2. Cavazzana A. O., Ninfo V., Roberts J., Triche T. J. Peripheral neuroepithelioma: a light microscopic, immunocytochemical, and ultrastructural study. Mod Pathol. 1992 Jan;5(1):71–78. [PubMed] [Google Scholar]
  3. Delattre O., Zucman J., Melot T., Garau X. S., Zucker J. M., Lenoir G. M., Ambros P. F., Sheer D., Turc-Carel C., Triche T. J. The Ewing family of tumors--a subgroup of small-round-cell tumors defined by specific chimeric transcripts. N Engl J Med. 1994 Aug 4;331(5):294–299. doi: 10.1056/NEJM199408043310503. [DOI] [PubMed] [Google Scholar]
  4. Desmaze C., Brizard F., Turc-Carel C., Melot T., Delattre O., Thomas G., Aurias A. Multiple chromosomal mechanisms generate an EWS/FLI1 or an EWS/ERG fusion gene in Ewing tumors. Cancer Genet Cytogenet. 1997 Aug;97(1):12–19. doi: 10.1016/s0165-4608(96)00326-3. [DOI] [PubMed] [Google Scholar]
  5. Dockhorn-Dworniczak B., Schäfer K. L., Dantcheva R., Blasius S., Winkelmann W., Strehl S., Burdach S., van Valen F., Jürgens H., Böcker W. Diagnostic value of the molecular genetic detection of the t(11;22) translocation in Ewing's tumours. Virchows Arch. 1994;425(2):107–112. doi: 10.1007/BF00230345. [DOI] [PubMed] [Google Scholar]
  6. Downing J. R., Head D. R., Parham D. M., Douglass E. C., Hulshof M. G., Link M. P., Motroni T. A., Grier H. E., Curcio-Brint A. M., Shapiro D. N. Detection of the (11;22)(q24;q12) translocation of Ewing's sarcoma and peripheral neuroectodermal tumor by reverse transcription polymerase chain reaction. Am J Pathol. 1993 Nov;143(5):1294–1300. [PMC free article] [PubMed] [Google Scholar]
  7. Hromas R., May W., Denny C., Raskind W., Moore J., Maki R. A., Beck E., Klemsz M. J. Human FLI-1 localizes to chromosome 11Q24 and has an aberrant transcript in neuroepithelioma. Biochim Biophys Acta. 1993 Feb 20;1172(1-2):155–158. doi: 10.1016/0167-4781(93)90283-j. [DOI] [PubMed] [Google Scholar]
  8. Jeon I. S., Davis J. N., Braun B. S., Sublett J. E., Roussel M. F., Denny C. T., Shapiro D. N. A variant Ewing's sarcoma translocation (7;22) fuses the EWS gene to the ETS gene ETV1. Oncogene. 1995 Mar 16;10(6):1229–1234. [PubMed] [Google Scholar]
  9. Kovar H. Progress in the molecular biology of ewing tumors. Sarcoma. 1998;2(1):3–17. doi: 10.1080/13577149878109. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  11. Melot T., Gruel N., Doubeikovski A., Sevenet N., Teillaud J. L., Delattre O. Production and characterization of mouse monoclonal antibodies to wild-type and oncogenic FLI-1 proteins. Hybridoma. 1997 Oct;16(5):457–464. doi: 10.1089/hyb.1997.16.457. [DOI] [PubMed] [Google Scholar]
  12. Mierau G. W., Berry P. J., Malott R. L., Weeks D. A. Appraisal of the comparative utility of immunohistochemistry and electron microscopy in the diagnosis of childhood round cell tumors. Ultrastruct Pathol. 1996 Nov-Dec;20(6):507–517. doi: 10.3109/01913129609016355. [DOI] [PubMed] [Google Scholar]
  13. Ohno T., Rao V. N., Reddy E. S. EWS/Fli-1 chimeric protein is a transcriptional activator. Cancer Res. 1993 Dec 15;53(24):5859–5863. [PubMed] [Google Scholar]
  14. Perlman E. J., Dickman P. S., Askin F. B., Grier H. E., Miser J. S., Link M. P. Ewing's sarcoma--routine diagnostic utilization of MIC2 analysis: a Pediatric Oncology Group/Children's Cancer Group Intergroup Study. Hum Pathol. 1994 Mar;25(3):304–307. doi: 10.1016/0046-8177(94)90203-8. [DOI] [PubMed] [Google Scholar]
  15. Peter M., Couturier J., Pacquement H., Michon J., Thomas G., Magdelenat H., Delattre O. A new member of the ETS family fused to EWS in Ewing tumors. Oncogene. 1997 Mar 13;14(10):1159–1164. doi: 10.1038/sj.onc.1200933. [DOI] [PubMed] [Google Scholar]
  16. Rettig W. J., Garin-Chesa P., Huvos A. G. Ewing's sarcoma: new approaches to histogenesis and molecular plasticity. Lab Invest. 1992 Feb;66(2):133–137. [PubMed] [Google Scholar]
  17. Sorensen P. H., Lessnick S. L., Lopez-Terrada D., Liu X. F., Triche T. J., Denny C. T. A second Ewing's sarcoma translocation, t(21;22), fuses the EWS gene to another ETS-family transcription factor, ERG. Nat Genet. 1994 Feb;6(2):146–151. doi: 10.1038/ng0294-146. [DOI] [PubMed] [Google Scholar]
  18. Thiele C. J. Pediatric peripheral neuroectodermal tumors, oncogenes, and differentiation. Cancer Invest. 1990;8(6):629–639. doi: 10.3109/07357909009018932. [DOI] [PubMed] [Google Scholar]
  19. Triche T. J. Neuroblastoma and other childhood neural tumors: a review. Pediatr Pathol. 1990;10(1-2):175–193. doi: 10.3109/15513819009067106. [DOI] [PubMed] [Google Scholar]
  20. Tsokos M., Linnoila R. I., Chandra R. S., Triche T. J. Neuron-specific enolase in the diagnosis of neuroblastoma and other small, round-cell tumors in children. Hum Pathol. 1984 Jun;15(6):575–584. doi: 10.1016/s0046-8177(84)80012-x. [DOI] [PubMed] [Google Scholar]
  21. Turc-Carel C., Aurias A., Mugneret F., Lizard S., Sidaner I., Volk C., Thiery J. P., Olschwang S., Philip I., Berger M. P. Chromosomes in Ewing's sarcoma. I. An evaluation of 85 cases of remarkable consistency of t(11;22)(q24;q12). Cancer Genet Cytogenet. 1988 Jun;32(2):229–238. doi: 10.1016/0165-4608(88)90285-3. [DOI] [PubMed] [Google Scholar]
  22. Wang M., Nilsson G., Carlberg M., Dricu A., Wejde J., Kreicbergs A., Larsson O. Specific and sensitive detection of the EWS/FLI1 fusion protein in Ewing's sarcoma by Western blotting. Virchows Arch. 1998 Feb;432(2):131–134. doi: 10.1007/s004280050146. [DOI] [PubMed] [Google Scholar]
  23. Wasylyk B., Hahn S. L., Giovane A. The Ets family of transcription factors. Eur J Biochem. 1993 Jan 15;211(1-2):7–18. doi: 10.1007/978-3-642-78757-7_2. [DOI] [PubMed] [Google Scholar]
  24. Weidner N., Tjoe J. Immunohistochemical profile of monoclonal antibody O13: antibody that recognizes glycoprotein p30/32MIC2 and is useful in diagnosing Ewing's sarcoma and peripheral neuroepithelioma. Am J Surg Pathol. 1994 May;18(5):486–494. [PubMed] [Google Scholar]
  25. Xin J. H., Cowie A., Lachance P., Hassell J. A. Molecular cloning and characterization of PEA3, a new member of the Ets oncogene family that is differentially expressed in mouse embryonic cells. Genes Dev. 1992 Mar;6(3):481–496. doi: 10.1101/gad.6.3.481. [DOI] [PubMed] [Google Scholar]
  26. Zucman J., Delattre O., Desmaze C., Plougastel B., Joubert I., Melot T., Peter M., De Jong P., Rouleau G., Aurias A. Cloning and characterization of the Ewing's sarcoma and peripheral neuroepithelioma t(11;22) translocation breakpoints. Genes Chromosomes Cancer. 1992 Nov;5(4):271–277. doi: 10.1002/gcc.2870050402. [DOI] [PubMed] [Google Scholar]

Articles from Sarcoma are provided here courtesy of Wiley

RESOURCES