Skip to main content

Advertisement

SpringerLink
Log in

Assessment of BRAFV600E mutation in pulmonary Langerhans cell histiocytosis in tissue biopsies and bronchoalveolar lavages by droplet digital polymerase chain reaction

  • Original Article
  • Published:
Virchows Archiv Aims and scope Submit manuscript

Abstract

The neoplastic nature of pulmonary Langerhans cell histiocytosis (PLCH) is still debated. As the detection of BRAF V600E and MAP2K1 mutations in patients with PCLH is now considered for such assessment, the aim of our study was to evaluate digital droplet polymerase chain reaction (ddPCR) in PCLH diagnosis. We retrospectively analyzed BRAFV600E detection in a cohort of 42 PCLH tissues and 18 bronchoalveolar lavages (BALs) by ddPCR, immunohistochemistry, high-resolution melting PCR (HRM), and next-generation sequencing (NGS). The presence of BRAFV600E mutation was assessed by at least two concordant techniques to further evaluate specificity and sensitivity of each method. The BRAF V600E mutation prevalence was detected in 18 out of 41 cases by ddPCR, 10 out of 36 cases by HRM PCR, and 16 out of 31 cases by NGS. BRAFV600E immunohistochemistry sensitivity was 94%, and specificity was 79%. HRM PCR sensitivity was only 59%, and specificity was 100%. NGS sensitivity and specificity were 100% for interpretable cases (n = 31), but in 11 cases, this technique was non-contributive. The analysis of BAL samples by ddPCR revealed a BRAFV600E mutation both in tissue and in BAL samples in one patient, a wild-type status both in tissue and in BAL samples in two patients, and a wild-type BRAF status in BAL and a BRAFV600E mutation in tissue samples in four patients. The study supports the usefulness of ddPCR for BRAF status assessment in either tissue or BAL samples to increase the accuracy of PLCH diagnosis.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  1. Swerdlow SH, Campo E, Harris NL, Jaffe ES, Pileri SA, Stein H, Thiele J, Vardiman JW (2008) WHO classification of tumours of haematopoietic and lymphoid tissues, fourth edition

  2. Emile JF, Abla O, Fraitag S, Horne A, Haroche J, Donadieu J, Requena-Caballero L, Jordan MB, Abdel-Wahab O, Allen CE, Charlotte F, Diamond EL, Egeler RM, Fischer A, Herrera JG, Henter JI, Janku F, Merad M, Picarsic J, Rodriguez-Galindo C, Rollins BJ, Tazi A, Vassallo R, Weiss LM (2016) Revised classification of histiocytoses and neoplasms of the macrophage-dendritic cell lineages. Blood 127:2672–2681

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Willman CL, Busque L, Griffith BB, Favara BE, McClain KL, Duncan MH, Gilliland DG (1994) Langerhans cell histiocytosis (histiocytosis X)—a clonal proliferative disease. N Engl J Med 331:154–160

    Article  CAS  PubMed  Google Scholar 

  4. Yu RC, Chu C, Buluwela L, Chu AC (1994) Clonal proliferation of Langerhans cells in Langerhans cell histiocytosis. Lancet 343:767–768

    Article  CAS  PubMed  Google Scholar 

  5. Badalian-Very G, Vergilio JA, Degar BA, MacConaill LE, Brandner B, Calicchio ML, Kuo FC, Ligon AH, Stevenson KE, Kehoe SM, Garraway LA, Hahn WC, Meyerson M, Fleming MD, Rollins BJ (2010) Recurrent BRAF mutations in Langerhans cell histiocytosis. Blood 116:1919–1923

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Haroche J, Charlotte F, Arnaud L, von Deimling A, Helias-Rodzewicz Z, Hervier B, Cohen-Aubart F, Launay D, Lesot A, Mokhtari K, Canioni D, Galmiche L, Rose C, Schmalzing M, Croockewit S, Kambouchner M, Copin MC, Fraitag S, Sahm F, Brousse N, Amoura Z, Donadieu J, Emile JF (2012) High prevalence of BRAF V600E mutations in Erdheim-Chester disease but not in other non-Langerhans cell histiocytoses. Blood 120:2700–2703

    Article  CAS  PubMed  Google Scholar 

  7. Sahm F, Capper D, Preusser M, Meyer J, Stenzinger A, Lasitschka F, Berghoff AS, Habel A, Schneider M, Kulozik A, Anagnostopoulos I, Mullauer L, Mechtersheimer G, von Deimling A (2012) BRAFV600E mutant protein is expressed in cells of variable maturation in Langerhans cell histiocytosis. Blood 120:e28–e34

    Article  CAS  PubMed  Google Scholar 

  8. Satoh T, Smith A, Sarde A, Lu HC, Mian S, Trouillet C, Mufti G, Emile JF, Fraternali F, Donadieu J, Geissmann F (2012) B-RAF mutant alleles associated with Langerhans cell histiocytosis, a granulomatous pediatric disease. PLoS One 7:e33891

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Wei R, Wang Z, Li X, Shu Y, Fu B (2013) Frequent BRAFV600E mutation has no effect on tumor invasiveness in patients with Langerhans cell histiocytosis. Biomed Rep 1:365–368

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Yousem SA, Dacic S, Nikiforov YE, Nikiforova M (2013) Pulmonary Langerhans cell histiocytosis: profiling of multifocal tumors using next-generation sequencing identifies concordant occurrence of BRAF V600E mutations. Chest 143:1679–1684

    Article  CAS  PubMed  Google Scholar 

  11. Berres ML, Lim KP, Peters T, Price J, Takizawa H, Salmon H, Idoyaga J, Ruzo A, Lupo PJ, Hicks MJ, Shih A, Simko SJ, Abhyankar H, Chakraborty R, Leboeuf M, Beltrao M, Lira SA, Heym KM, Bigley V, Collin M, Manz MG, McClain K, Merad M, Allen CE (2014) BRAF-V600E expression in precursor versus differentiated dendritic cells defines clinically distinct LCH risk groups. J Exp Med 211:669–683

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Bubolz AM, Weissinger SE, Stenzinger A, Arndt A, Steinestel K, Bruderlein S, Cario H, Lubatschofski A, Welke C, Anagnostopoulos I, Barth TF, Beer AJ, Moller P, Gottstein M, Viardot A, Lennerz JK (2014) Potential clinical implications of BRAF mutations in histiocytic proliferations. Oncotarget 5:4060–4070

    Article  PubMed  PubMed Central  Google Scholar 

  13. Chilosi M, Facchetti F, Calio A, Zamo A, Brunelli M, Martignoni G, Rossi A, Montagna L, Piccoli P, Dubini A, Tironi A, Tomassetti S, Poletti V, Doglioni C (2014) Oncogene-induced senescence distinguishes indolent from aggressive forms of pulmonary and non-pulmonary Langerhans cell histiocytosis. Leuk Lymphoma 55:2620–2626

    Article  CAS  PubMed  Google Scholar 

  14. Roden AC, Hu X, Kip S, Parrilla Castellar ER, Rumilla KM, Vrana JA, Vassallo R, Ryu JH, Yi ES (2014) BRAF V600E expression in Langerhans cell histiocytosis: clinical and immunohistochemical study on 25 pulmonary and 54 extrapulmonary cases. Am J Surg Pathol 38:548–551

    Article  PubMed  Google Scholar 

  15. Brown NA, Furtado LV, Betz BL, Kiel MJ, Weigelin HC, Lim MS, Elenitoba-Johnson KS (2014) High prevalence of somatic MAP2K1 mutations in BRAF V600E-negative Langerhans cell histiocytosis. Blood 124:1655–1658

    Article  CAS  PubMed  Google Scholar 

  16. Chakraborty R, Hampton OA, Shen X, Simko SJ, Shih A, Abhyankar H, Lim KP, Covington KR, Trevino L, Dewal N, Muzny DM, Doddapaneni H, Hu J, Wang L, Lupo PJ, Hicks MJ, Bonilla DL, Dwyer KC, Berres ML, Poulikakos PI, Merad M, McClain KL, Wheeler DA, Allen CE, Parsons DW (2014) Mutually exclusive recurrent somatic mutations in MAP2K1 and BRAF support a central role for ERK activation in LCH pathogenesis. Blood 124:3007–3015

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Alayed K, Medeiros LJ, Patel KP, Zuo Z, Li S, Verma S, Galbincea J, Cason RC, Luthra R, Yin CC (2016) BRAF and MAP2K1 mutations in Langerhans cell histiocytosis: a study of 50 cases. Hum Pathol 52:61–67

    Article  CAS  PubMed  Google Scholar 

  18. Li Y, Cai HR, Ma M, Wang LJ, Yang J, Ding JJ, Miao LY (2016) A case of pulmonary Langerhans cell Histiocytosis with BRAF V600E-negative and MAP2K1-posivtive mutations presenting as diffuse nodules in chest high-resolution computed tomography after smoking cessation. Chin Med J 129:1384–1386

    Article  PubMed  PubMed Central  Google Scholar 

  19. Zeng K, Ohshima K, Liu Y, Zhang W, Wang L, Fan L, Li M, Li X, Wang Z, Guo S, Yan Q, Guo Y (2016) BRAFV600E and MAP2K1 mutations in Langerhans cell histiocytosis occur predominantly in children. Hematol Oncol

  20. Zeng K, Wang Z, Ohshima K, Liu Y, Zhang W, Wang L, Fan L, Li M, Li X, Wang Y, Yu Z, Yan Q, Guo S, Wei J, Guo Y (2016) BRAF V600E mutation correlates with suppressive tumor immune microenvironment and reduced disease-free survival in Langerhans cell histiocytosis. Oncoimmunology 5:e1185582

    Article  PubMed  PubMed Central  Google Scholar 

  21. Nelson DS, Quispel W, Badalian-Very G, van Halteren AG, van den Bos C, Bovee JV, Tian SY, Van Hummelen P, Ducar M, MacConaill LE, Egeler RM, Rollins BJ (2014) Somatic activating ARAF mutations in Langerhans cell histiocytosis. Blood 123:3152–3155

    Article  CAS  PubMed  Google Scholar 

  22. Nelson DS, van Halteren A, Quispel WT, van den Bos C, Bovee JV, Patel B, Badalian-Very G, van Hummelen P, Ducar M, Lin L, MacConaill LE, Egeler RM, Rollins BJ (2015) MAP2K1 and MAP3K1 mutations in Langerhans cell histiocytosis. Genes Chromosom Cancer 54:361–368

    Article  CAS  PubMed  Google Scholar 

  23. Diamond EL, Durham BH, Haroche J, Yao Z, Ma J, Parikh SA, Wang Z, Choi J, Kim E, Cohen-Aubart F, Lee SC, Gao Y, Micol JB, Campbell P, Walsh MP, Sylvester B, Dolgalev I, Aminova O, Heguy A, Zappile P, Nakitandwe J, Ganzel C, Dalton JD, Ellison DW, Estrada-Veras J, Lacouture M, Gahl WA, Stephens PJ, Miller VA, Ross JS, Ali SM, Briggs SR, Fasan O, Block J, Heritier S, Donadieu J, Solit DB, Hyman DM, Baselga J, Janku F, Taylor BS, Park CY, Amoura Z, Dogan A, Emile JF, Rosen N, Gruber TA, Abdel-Wahab O (2016) Diverse and targetable kinase alterations drive histiocytic neoplasms. Cancer Discov 6:154–165

    Article  CAS  PubMed  Google Scholar 

  24. Kamionek M, Ahmadi Moghaddam P, Sakhdari A, Kovach AE, Welch M, Meng X, Dresser K, Tomaszewicz K, Cosar EF, Mark EJ, Fraire AE, Hutchinson L (2016) Mutually exclusive extracellular signal-regulated kinase pathway mutations are present in different stages of multi-focal pulmonary Langerhans cell histiocytosis supporting clonal nature of the disease. Histopathology 69:499–509

    Article  PubMed  Google Scholar 

  25. Mourah S, How-Kit A, Meignin V, Gossot D, Lorillon G, Bugnet E, Mauger F, Lebbe C, Chevret S, Tost J, Tazi A (2016) Recurrent NRAS mutations in pulmonary Langerhans cell histiocytosis. Eur Respir J 47:1785–1796

    Article  CAS  PubMed  Google Scholar 

  26. Heritier S, Emile JF, Barkaoui MA, Thomas C, Fraitag S, Boudjemaa S, Renaud F, Moreau A, Peuchmaur M, Chassagne-Clement C, Dijoud F, Rigau V, Moshous D, Lambilliotte A, Mazingue F, Kebaili K, Miron J, Jeziorski E, Plat G, Aladjidi N, Ferster A, Pacquement H, Galambrun C, Brugieres L, Leverger G, Mansuy L, Paillard C, Deville A, Armari-Alla C, Lutun A, Gillibert-Yvert M, Stephan JL, Cohen-Aubart F, Haroche J, Pellier I, Millot F, Lescoeur B, Gandemer V, Bodemer C, Lacave R, Helias-Rodzewicz Z, Taly V, Geissmann F, Donadieu J (2016) BRAF mutation correlates with high-risk Langerhans cell histiocytosis and increased resistance to first-line therapy. J Clin Oncol 34:3023–3030

    Article  PubMed  PubMed Central  Google Scholar 

  27. Girschikofsky M, Arico M, Castillo D, Chu A, Doberauer C, Fichter J, Haroche J, Kaltsas GA, Makras P, Marzano AV, de Menthon M, Micke O, Passoni E, Seegenschmiedt HM, Tazi A, McClain KL (2013) Management of adult patients with Langerhans cell histiocytosis: recommendations from an expert panel on behalf of Euro-Histio-Net. Orphanet J Rare Dis 8:72

    Article  PubMed  PubMed Central  Google Scholar 

  28. Dimmler A, Geddert H, Werner M, Faller G (2017) Molecular analysis of BRAF V600E mutation in multiple nodules of pulmonary Langerhans cell histiocytosis 470:429–35

  29. Chollet S, Soler P, Dournovo P, Richard MS, Ferrans VJ, Basset F (1984) Diagnosis of pulmonary histiocytosis X by immunodetection of Langerhans cells in bronchoalveolar lavage fluid. Am J Surg Pathol 115:225–232

    CAS  Google Scholar 

  30. Auerswald U, Barth J, Magnussen H (1991) Value of CD-1-positive cells in bronchoalveolar lavage fluid for the diagnosis of pulmonary histiocytosis X. Lung 169

  31. Lommatzsch M, Bratke K, Stoll P, Mulleneisen N, Prall F, Bier A (2016) Bronchoalveolar lavage for the diagnosis of pulmonary Langerhans cell histiocytosis. Vircho Respir Med 119:168–174

    Article  PubMed  Google Scholar 

  32. Hyman DM, Puzanov I, Subbiah V, Faris JE, Chau I, Blay JY, Wolf J, Raje NS, Diamond EL, Hollebecque A, Gervais R, Elez-Fernandez ME, Italiano A, Hofheinz RD, Hidalgo M, Chan E, Schuler M, Lasserre SF, Makrutzki M, Sirzen F, Veronese ML, Tabernero J, Baselga J (2015) Vemurafenib in multiple nonmelanoma cancers with BRAF V600 mutations. N Engl J Med 373:726–736

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Haroche J, Cohen-Aubart F, Emile JF, Arnaud L, Maksud P, Charlotte F, Cluzel P, Drier A, Hervier B, Benameur N, Besnard S, Donadieu J, Amoura Z (2013) Dramatic efficacy of vemurafenib in both multisystemic and refractory Erdheim-Chester disease and Langerhans cell histiocytosis harboring the BRAF V600E mutation. Blood 121:1495–1500

    Article  CAS  PubMed  Google Scholar 

  34. Boursault L, Haddad V, Vergier B, Cappellen D, Verdon S, Bellocq JP, Jouary T, Merlio JP (2013) Tumor homogeneity between primary and metastatic sites for BRAF status in metastatic melanoma determined by immunohistochemical and molecular testing. PLoS One 8:e70826

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Ilie M, Long E, Hofman V, Dadone B, Marquette CH, Mouroux J, Vignaud JM, Begueret H, Merlio JP, Capper D, von Deimling A, Emile JF, Hofman P (2013) Diagnostic value of immunohistochemistry for the detection of the BRAFV600E mutation in primary lung adenocarcinoma Caucasian patients. Ann Oncol 24:742–748

    Article  CAS  PubMed  Google Scholar 

  36. Ilie MI, Long-Mira E, Hofman V, Mouroux J, Vignaud JM, Gauchotte G, Begueret H, Merlio JP, Emile JF, Hebuterne X, Hofman P (2014) BRAFV600E mutation analysis by immunohistochemistry in patients with thoracic metastases from colorectal cancer. Pathology 46:311–315

    Article  CAS  PubMed  Google Scholar 

  37. Janku F, Vibat CR, Kosco K, Holley VR, Cabrilo G, Meric-Bernstam F, Stepanek VM, Lin PP, Leppin L, Hassaine L, Poole JC, Kurzrock R, Erlander MG (2014) BRAF V600E mutations in urine and plasma cell-free DNA from patients with Erdheim-Chester disease. Oncotarget 5:3607–3610

    PubMed  PubMed Central  Google Scholar 

  38. Hyman DM, Diamond EL, Vibat CR, Hassaine L, Poole JC, Patel M, Holley VR, Cabrilo G, Lu TT, Arcila ME, Chung YR, Rampal R, Lacouture ME, Rosen N, Meric-Bernstam F, Baselga J, Kurzrock R, Erlander MG, Janku F, Abdel-Wahab O (2015) Prospective blinded study of BRAFV600E mutation detection in cell-free DNA of patients with systemic histiocytic disorders. Cancer Discov 5:64–71

    Article  CAS  PubMed  Google Scholar 

  39. Roden AC, Yi ES (2016) Pulmonary Langerhans cell Histiocytosis: an update from the pathologists’ perspective. Arch Pathol Lab Med 140:230–240

    Article  PubMed  Google Scholar 

  40. Capper D, Preusser M, Habel A, Sahm F, Ackermann U, Schindler G, Pusch S, Mechtersheimer G, Zentgraf H, von Deimling A (2011) Assessment of BRAF V600E mutation status by immunohistochemistry with a mutation-specific monoclonal antibody. Acta Neuropathol 122:11–19

    Article  CAS  PubMed  Google Scholar 

  41. Wang K, Li M, Hakonarson H (2010) ANNOVAR: functional annotation of genetic variants from high-throughput sequencing data. Nucleic Acids Res 38:e164

    Article  PubMed  PubMed Central  Google Scholar 

  42. Yousem SA (2001) Pulmonary Langerhans’ cell histiocytosis: molecular analysis of clonality. Am J Surg Pathol 25

  43. Mourah S, Denis MG, Narducci FE, Solassol J, Merlin JL, Sabourin JC, Scoazec JY, Ouafik L, Emile JF, Heller R, Souvignet C, Bergougnoux L, Merlio JP (2015) Detection of BRAF V600 mutations in melanoma: evaluation of concordance between the Cobas(R) 4800 BRAF V600 mutation test and the methods used in French National Cancer Institute (INCa) platforms in a real-life setting. PLoS One 10:e0120232

    Article  PubMed  PubMed Central  Google Scholar 

  44. Barlesi F, Mazieres J, Merlio JP, Debieuvre D, Mosser J, Lena H, Ouafik L, Besse B, Rouquette I, Westeel V, Escande F, Monnet I, Lemoine A, Veillon R, Blons H, Audigier-Valette C, Bringuier PP, Lamy R, Beau-Faller M, Pujol JL, Sabourin JC, Penault-Llorca F, Denis MG, Lantuejoul S, Morin F, Tran Q, Missy P, Langlais A, Milleron B, Cadranel J, Soria JC, Zalcman G (2016) Routine molecular profiling of patients with advanced non-small-cell lung cancer: results of a 1-year nationwide programme of the French Cooperative Thoracic Intergroup (IFCT). Lancet 387:1415–1426

    Article  CAS  PubMed  Google Scholar 

  45. Egeler RM, Neglia JP, Puccetti DM, Brennan CA, Nesbit ME (1993) Association of Langerhans cell histiocytosis with malignant neoplasms. Hematol Oncol Clin North Am 71:865–873

    CAS  Google Scholar 

  46. Egeler RM, Neglia JP, Arico M, Favara BE, Heitger A, Nesbit ME, Nicholson HS (1998) The relation of Langerhans cell histiocytosis to acute leukemia, lymphomas, and other solid tumors. The LCH-Malignancy Study Group of the Histiocyte Society. Hematol Oncol Clin North Am 12:369–378

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We would like to acknowledge Sabine Berouhet, Fabian Martinez, and all the laboratory of Tumor Biology of Bordeaux University Hospital team for their contribution to this work.

Author information

Author notes

  1. Clémence Pierry and Charline Caumont contributed equally to this work.

Authors and Affiliations

  1. Department of Pathology, CHU Bordeaux, Pessac, F-33604, Bordeaux, France

    Clémence Pierry, Marion Marty & Hugues Bégueret

  2. Laboratory of Tumor Biology and Tumor Biobank, CHU Bordeaux, Pessac, F-33604, Bordeaux, France

    Clémence Pierry, Charline Caumont, Audrey Gros, Thomas Bandres, Séverine Verdon & Jean-Philippe Merlio

  3. U1053 Inserm, University Bordeaux, F-33076, Bordeaux, France

    Charline Caumont, Audrey Gros & Jean-Philippe Merlio

  4. Department of Pneumology, CHU Bordeaux, Pessac, F-33604, Bordeaux, France

    Elodie Blanchard & Camille Brochet

  5. Univercity Bordeaux, Centre de Recherche Cardio-Thoracique de Bordeaux, INSERM U1045, CIC 1401, F-33000, 146 rue Léo Saignat, 33076, Bordeaux, France

    Gael Dournes

Authors
  1. Clémence Pierry
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Charline Caumont
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Elodie Blanchard
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Camille Brochet
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Gael Dournes
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Audrey Gros
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Thomas Bandres
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Séverine Verdon
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Marion Marty
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Hugues Bégueret
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Jean-Philippe Merlio
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jean-Philippe Merlio.

Ethics declarations

According to the French Public Health and Bioethical Law, the present study was considered by our research board as a non-interventional study without the need for ethics committee approval (Article L1121-1 and Article R1121-3). Moreover, informed consent was given to all patients when alive for the use of the clinical data and research on their biological material. All data were collected anonymously.

Conflict of interest

There is no conflict of interest to disclose except the fact that Roche Diagnostics (Meylan, France) has provided free of charge the anti-BRAFV600E antibody.

Additional information

Virchows Archiv conforms to the ICMJE recommendation for qualification of authorship. The ICMJE recommends that authorship be based on the following four criteria:

• Substantial contributions to the conception or design of the work, or the acquisition, analysis, or interpretation of data for the work; AND

• Drafting the work or revising it critically for important intellectual content; AND

• Final approval of the version to be published; AND

• Agreement to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

Electronic supplementary material

Supplemental table 1

Genes and exons covered by the Ion Ampliseq panel (XLSX 9 kb)

Supplemental figure 1

Identification of MAP2K1Q56P mutation in pulmonary Langerhans cell histiocytosis tissue (case 2). NGS, BAM alignment with Alamut software (partial sequence, sense strand) of MAP2K1 exon 2. (PPTX 304 kb)

Supplemental figure 2

Identification of BRAFV600E mutation in pulmonary Langerhans cell histiocytosis broncho-alveolar lavage by ddPCR (case 27). a. Fluorescence results are plotted as two-dimensional dot plots. The region of these plots can be sequentially separated based on the fluorescence intensity of each droplet. Gray dots correspond to empty droplets. Green dots correspond to droplets containing WT copies of BRAF codon 600. Blue dots correspond to droplets containing at least BRAFV600E mutation. Orange dots correspond to droplets containing at least one BRAFV600E WT copy and one mutant copy. These droplets are considered for analysis. c. Wild-type(WT) template concentration in copies per μl (cpm) (green square), mutant template cpm (blue square), and fractional abundance of mutant to WT template (red diamond). (PPTX 95 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Pierry, C., Caumont, C., Blanchard, E. et al. Assessment of BRAFV600E mutation in pulmonary Langerhans cell histiocytosis in tissue biopsies and bronchoalveolar lavages by droplet digital polymerase chain reaction. Virchows Arch 472, 247–258 (2018). https://doi.org/10.1007/s00428-017-2185-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00428-017-2185-0

Keywords

Search

Navigation