Abstract
Implementation of interphase fluorescence in situ hybridization (FISH) assays in the clinical laboratory requires validation against established methods. Validation tools in common use include exchange of consecutive sections with another institution that has already established the FISH assay, comparison with conventional banded metaphase cytogenetics, confirmation of specificity using probed normal metaphases, consecutive paraffin sections of a validation set tested by a reference laboratory, and specificity assessment against well characterized cell lines. We have investigated the feasibility of using tissue microarrays (TMA) constructed from murine xenografts as a preliminary specificity-screening tool for validation of interphase FISH assays. Cell lines currently in use for FISH controls are used to generate xenografts in SCID mice which are fixed in formalin and paraffin embedded. A TMA is constructed using duplicate donor cores from the xenograft blocks. Xenografts used represent a wide range of translocations used routinely for formalin fixed paraffin embedded sections evaluated by FISH. Probe cocktails (Abbott–Vysis), for several non-random translocations associated with hematologic neoplasms and soft tissue sarcomas have been used in this manner. On-line deparaffinization, cell conditioning, and prehybridization steps are automated using a staining workstation (Ventana Discovery XT); hybridization and stringency washes are performed manually offline. FISH-probed TMAs are tracked using a Metasystems image scanner and analyzed using classifiers specifically developed for each molecular abnormality. FISH results for each xenograft in the TMA correspond exactly to the genotype previously established for the parent cell line from which the xenograft was prepared. Moderate complexity tissue microarrays constructed from murine xenografts are excellent validation tools for initial assessment of interphase FISH probe specificity.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Barrans SL, O’Connor SJ, Evans PA, Davies FE, Owen RG, Haynes AP, Morgan GJ, Jack AS (2002) Rearrangement of the BCL6 locus at 3q27 is an independent poor prognostic factor in nodal diffuse large B-cell lymphoma. Br J Haematol 117:322–332
Barrans SL, Evans PA, O’Connor SJ, Owen RG, Morgan GJ, Jack AS (2003) The detection of t(14;18) in archival lymph nodes: development of a fluorescence in situ hybridization (FISH)-based method and evaluation by comparison with polymerase chain reaction. J Mol Diagn 5:168–175
Bijwaard KE, Fetsch JF, Przygodzki R, Taubenberger JK, Lichy JH (2002) Detection of SYT-SSX fusion transcripts in archival synovial sarcomas by real-time reverse transcriptase-polymerase chain reaction. J Mol Diagn 4:59–64
Cataldo KA, Jalal SM, Law ME, Ansell SM, Inwards DJ, Fine M, Arber DA, Pulford KA, Strickler JG (1999) Detection of t(2;5) in anaplastic large cell lymphoma: comparison of immunohistochemical studies, FISH, and RT-PCR in paraffin-embedded tissue. Am J Surg Pathol 23:1386–1392
Chaganti SR, Chen W, Parsa N, Offit K, Louie DC, Dalla-Favera R, Chaganti RS (1998) Involvement of BCL6 in chromosomal aberrations affecting band 3q 27 in B-cell non-Hodgkin lymphoma. Genes Chromosomes Cancer 23:323–327
Chaganti RS, Nanjangud G, Schmidt H, Teruya-Feldstein J (2000) Recurring chromosomal abnormalities in non-Hodgkin’s lymphoma: biologic and clinical significance. Semin Hematol 37:396–411
Cook J (2004) Paraffin section interphase fluorescence in situ hybridization in the diagnosis and classification of non-hodgkin lymphomas. Diagn Mol Pathol 13:197–206
Cook JR, Sherer M, Craig FE, Shekhter-Levin S, Swerdlow SH (2003) T(14;18)(q32;q21) involving MALT1 and IGH genes in an extranodal diffuse large B-cell lymphoma. Hum Pathol 34:1212–1215
Cook JR, Shekhter-Levin S, Swerdlow SH (2004) Utility of routine classical cytogenetic studies in the evaluation of suspected lymphomas: results of 279 consecutive lymph node/extranodal tissue biopsies. Am J Clin Pathol 121:826–835
Dierlamm J, Wlodarska I, Michaux L, Stefanova M, Hinz K, Van Den Berghe H, Hagemeijer A, Hossfeld DK (2000) Genetic abnormalities in marginal zone B-cell lymphoma. Hematol Oncol 18:1–13
Dohner H, Stilgenbauer S, Fischer K, Bentz M, Lichter P (1997) Cytogenetic and molecular cytogenetic analysis of B cell chronic lymphocytic leukemia: specific chromosome aberrations identify prognostic subgroups of patients and point to loci of candidate genes. Leukemia 11(Suppl 2):S19–S24
Frater JL, Tsiftsakis EK, Hsi ED, Pettay J, Tubbs RR (2001) Use of novel t(11;14) and t(14;18) dual-fusion fluorescence in situ hybridization probes in the differential diagnosis of lymphomas of small lymphocytes. Diagn Mol Pathol 10:214–222
Godon A, Moreau A, Talmant P, Baranger-Papot L, Genevieve F, Milpied N, Zandecki M, Avet-Loiseau H (2003) Is t(14;18)(q32;q21) a constant finding in follicular lymphoma? An interphase FISH study on 63 patients. Leukemia 17:255–259
Haralambieva E, Kleiverda K, Mason DY, Schuuring E, Kluin PM (2002) Detection of three common translocation breakpoints in non-Hodgkin’s lymphomas by fluorescence in situ hybridization on routine paraffin-embedded tissue sections. J Pathol 198:163–170
Haralambieva E, Banham AH, Bastard C, Delsol G, Gaulard P, Ott G, Pileri S, Fletcher JA, Mason DY (2003) Detection by the fluorescence in situ hybridization technique of MYC translocations in paraffin-embedded lymphoma biopsy samples. Br J Haematol 121:49–56
Kumar S, Pack S, Kumar D, Walker R, Quezado M, Zhuang Z, Meltzer P, Tsokos M (1999) Detection of EWS-FLI-1 fusion in Ewing’s sarcoma/peripheral primitive neuroectodermal tumor by fluorescence in situ hybridization using formalin-fixed paraffin-embedded tissue. Hum Pathol 30:324–330
Lee W, Han K, Harris CP, Shim S, Kim S, Meisner LF (1993) Use of FISH to detect chromosomal translocations and deletions. Analysis of chromosome rearrangement in synovial sarcoma cells from paraffin-embedded specimens. Am J Pathol 143:15–19
Li JY, Gaillard F, Moreau A, Harousseau JL, Laboisse C, Milpied N, Bataille R, Avet-Loiseau H (1999) Detection of translocation t(11;14)(q13;q32) in mantle cell lymphoma by fluorescence in situ hybridization. Am J Pathol 154:1449–1452
Mathew P, Valentine MB, Bowman LC, Rowe ST, Nash MB, Valentine VA, Cohn SL, Castleberry RP, Brodeur GM, Look AT (2001) Detection of MYCN gene amplification in neuroblastoma by fluorescence in situ hybridization: a pediatric oncology group study. Neoplasia 3:105–109
Matutes E, Oscier D, Garcia-Marco J, Ellis J, Copplestone A, Gillingham R, Hamblin T, Lens D, Swansbury GJ, Catovsky D (1996) Trisomy 12 defines a group of CLL with atypical morphology: correlation between cytogenetic, clinical and laboratory features in 544 patients. Br J Haematol 92:382–388
Misra DN, Dickman PS, Yunis EJ (1995) Fluorescence in situ hybridization (FISH) detection of MYCN oncogene amplification in neuroblastoma using paraffin-embedded tissues. Diagn Mol Pathol 4:128–135
Paternoster SF, Brockman SR, Mcclure RF, Remstein ED, Kurtin PJ, Dewald GW (2002) A new method to extract nuclei from paraffin-embedded tissue to study lymphomas using interphase fluorescence in situ hybridization. Am J Pathol 160:1967–1972
Rao PH, Houldsworth J, Dyomina K, Parsa NZ, Cigudosa JC, Louie DC, Popplewell L, Offit K, Jhanwar SC, Chaganti RS (1998) Chromosomal and gene amplification in diffuse large B-cell lymphoma. Blood 92:234–240
Remstein ED, Kurtin PJ, Buno I, Bailey RJ, Proffitt J, Wyatt WA, Hanson CA, Dewald GW (2000) Diagnostic utility of fluorescence in situ hybridization in mantle-cell lymphoma. Br J Haematol 110:856–862
Sanchez-Izquierdo D, Buchonnet G, Siebert R, Gascoyne RD, Climent J, Karran L, Marin M, Blesa D, Horsman D, Rosenwald A, Staudt LM, Albertson DG, Du MQ, Ye H, Marynen P, Garcia-Conde J, Pinkel D, Dyer MJ, Martinez-Climent JA (2003) MALT1 is deregulated by both chromosomal translocation and amplification in B-cell non-Hodgkin lymphoma. Blood,101:4539–4546
Sartelet H, Grossi L, Pasquier D, Combaret V, Bouvier R, Ranchere D, Plantaz D, Munzer M, Philip T, Birembaut P, Zahm JM, Bergeron C, Gaillard D, Pasquier B (2002) Detection of N-myc amplification by FISH in immature areas of fixed neuroblastomas: more efficient than Southern blot/PCR. J Pathol 198:83–91
Schurter MJ, Lebrun DP, Harrison KJ (2002) Improved technique for fluorescence in situ hybridisation analysis of isolated nuclei from archival, B5 or formalin fixed, paraffin wax embedded tissue. Mol Pathol 55:121–124
Shaminie J, Peh SC, Tan MJ (2003) Improvement in the detection rate of t(14;18) translocation on paraffin-embedded tissue: a combination approach using PCR and FISH. Pathology 35:414–421
Streubel B, Lamprecht A, Dierlamm J, Cerroni L, Stolte M, Ott G, Raderer M, Chott A (2003) T(14;18)(q32;q21) involving IGH and MALT1 is a frequent chromosomal aberration in MALT lymphoma. Blood 101:2335–2339
Sun T, Nordberg ML, Cotelingam JD, Veillon DM, Ryder J (2003) Fluorescence in situ hybridization: method of choice for a definitive diagnosis of mantle cell lymphoma. Am J Hematol 74:78–84
Takada S, Yoshino T, Taniwaki M, Nakamura N, Nakamine H, Oshima K, Sadahira Y, Inagaki H, Oshima K, Tadaatsu A (2003) Involvement of the chromosomal translocation t(11;18) in some mucosa-associated lymphoid tissue lymphomas and diffuse large B-cell lymphomas of the ocular adnexa: evidence from multiplex reverse transcriptase-polymerase chain reaction and fluorescence in situ hybridization on using formalin-fixed, paraffin-embedded specimens. Mod Pathol 16:445–452
Tan LH, Do E, Chong SM, Koay ES (2003) Detection of ALK gene rearrangements in formalin-fixed, paraffin-embedded tissue using a fluorescence in situ hybridization (FISH) probe: a search for optimum conditions of tissue archiving and preparation for FISH. Mol Diagn 7:27–33
Tibiletti MG, Bernasconi B, Dionigi A, Riva C (1999) The applications of FISH in tumor pathology. Adv Clin Path 3:111–118
Tubbs REA (2001) Discrepancies in clinical laboratory testing of eligibility for trastuzumab therapy: apparent immunohistochemical false-positives do not get the message. J Clin Oncol 19:2714–2721
Tubbs R, Swain E, Pettay J, Hicks D An approach to the validation of novel molecular markers of breast cancer via TMA-based FISH scanning. J Mol Histol (in press)
Vega F, Medeiros LJ (2003) Chromosomal translocations involved in non-Hodgkin lymphomas. Arch Pathol Lab Med 127:1148–1160
Wolfe KQ, Herrington CS (1997) Interphase cytogenetics and pathology: a tool for diagnosis and research. J Pathol 181:359–361
Wotherspoon AC, Finn TM, Isaacson PG (1995) Trisomy 3 in low-grade B-cell lymphomas of mucosa-associated lymphoid tissue. Blood 85: 2000–2004
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Tubbs, R.R., Pettay, J., Barry, T.S. et al. The specificity of interphase FISH translocation probes in formalin fixed paraffin embedded tissue sections is readily assessed using automated staining and scoring of tissue microarrays constructed from murine xenografts. J Mol Hist 38, 159–165 (2007). https://doi.org/10.1007/s10735-006-9066-1
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10735-006-9066-1