Pathways Case Studies: April 2023

Mayo Clinic Laboratories sends a blood sample to the lab with an order to perform a Fetomaternal Bleed, Flow Cytometry test. The mother is Rh negative, and the team needs to know how much RhIg to administer. When the sample is run, the histogram shows a distinct cell peak (B) of 20.23% as shown in Figure 1.

Figure 1
Figure 2

In Figure 1, which cell population is represented by peak B and how much RhIg is needed?

  1. Peak B denotes Adult F cells. Mother likely has HPFH-hereditary persistence of fetal hemoglobin. No additional testing is needed. Administer 1 vial of RhIg.
  2. Peak B denotes Adult F cells. Order Dual Antibody Fetal Cell Count testing.
  3. Peak B denotes fetal cells. Perform a Kleihauer Betke test to quantify fetal cells.
  4. Peak B denotes the fetal cells. Contact the team and inform them that a large fetomaternal bleed has occurred and provide RhIg dose required.

The correct answer is ...

Peak B denotes Adult F cells. Mother likely has HPFH-hereditary persistence of fetal hemoglobin. No additional testing is needed. Administer 1 vial of RhIg.

Figure 1

(A) % Adult cells = 79.71

(B) % Adult F cells = 20.23

(C) % Fetal cells = 0.06

Adult cell: Peak A: RBC population that does not contain Fetal Hemoglobin.

Adult F cells: Peak B: Dim-fluorescing cells that do not stain as brightly as true fetal cells: these are RBCs that contain a small amount of HbF but are not true fetal cells. This “shoulder” will be elevated in hereditary persistence of fetal hemoglobin.

Fetal cells: Peak C. True fetal cell population containing Hemoglobin F and stains brightly positive.

Fetal hemoglobin (hemoglobin F, HbF, alpha2gamma2) is the most abundant hemoglobin during gestation and accounts for 60 to 80 percent of total hemoglobin in the full-term newborn. It is almost completely replaced by adult hemoglobin (hemoglobin A, HbA, alpha2beta2) by 6 to 12 months of age in individuals without hemoglobinopathies. Eventually, in adulthood, it accounts for less than 1% of total hemoglobin. Hereditary persistence of fetal hemoglobin (HPFH) is a benign condition in which significant fetal hemoglobin production continues into adulthood, bypassing the normal shutoff point after which only adult-type hemoglobin should be produced.

In this case the Adult F cells was easily distinguished from true fetal cell gate, which contained 0.06% (C) and further testing was not necessary.

The dosing of RhIg is based on the number of fetal cells in the maternal circulation. The rules, however, provide a safety margin of 1 additional vial based on rounding so even if the calculation of RhIg suggests 0, 1 vial of RhIg is always given if the mother is RhNeg and the child is RhPos. For example:

Calculating the RhIG Dose: % Fetal Bleed x 5000 ml  +1 

                              0.0006 X 5000/30= 0.1(round down) + 1  = 1.0 vial of Rhig

Incorrect answers explanation:

Large amounts of Adult F cells are seen in hereditary persistence of fetal hemoglobin (HPFH) in which the shoulder of HPFH cells (Adult F cells) encroach on the true fetal cell gate. Refer to Figure 2(a) where peak for gate B (Adult F cells 26.21%) extends into gate C (True fetal cells 4.50%). This can interfere with an accurate determination of the fetal cells. The consequences of missing a fetal bleed are significant and can result in an immunizing event leading to hemolytic disease of the newborn in subsequent pregnancies. Therefore, it is imperative to count these cells accurately and dose the appropriate amount of RhIg.

In a mother with known HPFH, this can make it difficult to determine a FMB accurately. Most labs use a single antibody (anti-fetal hemoglobin, HbF) kit to detect fetal hemoglobin. With only one single antibody, there are limitations with the samples that have significant adult F cells containing HbF overlapping with the true fetal cells.

A Dual Antibody Fetal Cell Count Kit includes antibodies for hemoglobin F (HbF) and carbonic anhydrase (CA) providing additional discrimination between the fetal and maternal HbF+ cells in mothers with underlying HPHF. The kit uses anti-carbonic anhydrase (CA) and anti-fetal hemoglobin (HbF). Fetal RBCs will be negative for CA (HbF+CA-) while the mother’s RBCs with high amount of fetal Hb will stain positive for both HbF and CA. (HbF+CA+).

See Figure 2

Anti-HbF is used on the Y axis and Anti-CA on the X. Thus, relevant true fetal bleeds Fetal cells would be in the LUQ (HbF+, HbCA-). In this case they represent 0.25%.

In the lower right a second antibody to carbonic anhydrase (Anti CA) is used to distinguish Adult cells from fetal cells (HBF-, CA+) (78.74%).

Adult F cells with increased amounts of fetal Hb seen in HPFH are in the RUQ (Right upper quadrant) (HbF+, CA+) (20.38%).

Adding a secondary antibody such as carbonic anhydrase (CA) improved the delineation of fetal cells from 4.50% in Fig 2a to 0.25% fetal cells shown in the LUQ in Fig 2b. This demonstrates the utility of CA in discriminating fetal and maternal HbF+ cells in mothers with an underlying hereditary persistence of fetal hemoglobin.

Peak C, not Peak B denotes fetal cells and although the Kleihauer-Betke assay is a quantification test, it is not able to reliably separate maternal and fetal HbF+ cells and would not be useful in a maternal sample with an underlying hemoglobinopathy.


  1. Sharma DC, Singhal S, Woike P, Rai S, Yadav M, Gaur R. Hereditary persistence of fetal hemoglobin. Asian J Transfus Sci. 2020 Jul-Dec;14(2):185-186. doi:10.4103/ajts.AJTS_71_16. Epub 2020 Dec 19. PMID: 33767547; PMCID: PMC7983139.
  2. Clark G, Svensson AM, Lieberman L. Perinatal issues in transfusion practice. Technical Manual. 20th Ed. Bethesda (MD); 2020:659-672.
  3. Krywko DM, Yarrarapu SNS, Shunkwiler SM. Kleihauer Betke Test. 2022 Aug 8. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2022 Jan–. PMID: 28613626.

Saadiya Nazli, M.B.B.S.

Fellow, Cellular Therapy
Mayo Clinic

Margaret (Maggie) DiGuardo, M.D.

Consultant, Transfusion Medicine
Mayo Clinic
Assistant Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science

A 21-year-old man with a past medical history of recurrent epistaxis, recent weight loss, joint pain, and lower extremity paresthesia presented with diffuse alveolar hemorrhage and upper airway ulcerations. High-resolution chest CT revealed multiple non-cavitary lung nodules with ground glass opacities. CRP was elevated at 252 mg/L and antibody serology testing was positive for antineutrophil cytoplasmic antibodies with a granular cytoplasmic staining pattern (c-ANCA), as well as PR3 and GBM autoantibodies. Antinuclear antibody (ANA) testing by Hep-2 substrate was negative. Urinalysis revealed hematuria with RBCs >100.

Figure 1

Based on these findings, what is the most likely diagnosis?

  • Systemic lupus erythematosus (SLE)
  • Granulomatosis with polyangiitis (GPA) with renal involvement
  • Anti-glomerular basement membrane disease (anti-GBM disease)
  • Microscopic polyangiitis (MPA)

The correct answer is ...

Granulomatosis with polyangiitis (GPA) with renal involvement.

Antineutrophil cytoplasmic antibody (ANCA) associated vasculitides (AAV) are a group of small vessel vasculitis disorders in which the development of autoantibodies to the neutrophil proteins myeloperoxidase (MPO) and proteinase 3 (PR3) play a central role in pathogenesis. AAV include three clinical diseases, namely granulomatosis with polyangiitis (GPA), microscopic polyangiitis (MPA), and eosinophilic GPA (EGPA), which are characterized by neutrophil-driven blood vessel inflammation, endothelial injury and tissue damage.1

Autoantibodies against PR3 and MPO produce unique staining patterns when indirect immunofluorescence (IIF) is performed on ethanol-fixed neutrophils. Although this technique is still routinely available, recent guidelines recommend antigen-specific immunoassays as the preferred screening test for patients with suspected AAV.2

Autoantibodies to PR3 (PR3-ANCA) occur in 75%3 of patients with GPA and produce a characteristic pattern of granular cytoplasmic fluorescence on ethanol-fixed neutrophils called the cANCA pattern (Figure 1A). Clinically, GPA is most often associated with upper and lower respiratory tract involvement.

Antibodies to MPO (MPO-ANCA) occur in approximately 60% of patients with MPA and 80%3 of patients with renal limited vasculitis and produce a pattern of perinuclear cytoplasmic fluorescence on ethanol-fixed neutrophils called the pANCA pattern (Figure 1B). MPA commonly manifests with severe renal involvement; however, pulmonary symptoms overlapping with GPA have been described.

Certain cases GPA or MPA may be associated with pANCA or cANCA patterns, making the differential diagnosis difficult.4 Additionally, 5%-10% of patients may have GPA, MPA, or renal limited vasculitis in the absence of and ANCA. Interestingly, Recent genetic and clinical evidence has suggested that these clinical syndromes may be more appropriately classified as PR3-positive AAV or MPO-positive AAV.5,6

In this case, the patient presented with upper respiratory symptoms including recurrent epistaxis and diffuse alveolar hemorrhage which are generally consistent with GPA. Further supporting this diagnosis, serology testing revealed PR3-ANCA with a cANCA staining pattern. The patient’s urinalysis, however, showed hematuria and additional serology testing revealed the presence of anti-GBM antibodies, which could be consistent with anti-GBM disease. To differentiate between the two, biopsies of the sinus mucosa and kidney were performed. Sinus mucosa biopsy could not definitively identify vasculitis; however, the renal biopsy revealed necrotizing glomerulonephritis with focal cellular crescents. The patient’s clinical presentation, serology results, biopsies, and additional laboratory studies finalized the diagnosis of GPA with renal involvement.


  1. Jennette JC, Falk RJ, Bacon PA, Basu N, Cid MC, Ferrario F, et al. 2012 revised International Chapel Hill Consensus Conference nomenclature of vasculitides. Arthritis Rheum. 2013;65:1-11
  2. Bossuyt X, Cohen Tervaert JW, Arimura Y, Blockmans D, Flores-Suárez LF, Guillevin L, et al. Position paper: Revised 2017 international consensus on testing of ANCAs in granulomatosis with polyangiitis and microscopic polyangiitis. Nat Rev Rheumatol. 2017;13:683-92
  3. Geetha D, Jefferson JA. ANCA-Associated Vasculitis: Core Curriculum 2020. Am J Kidney Dis. 2020;75:124-37
  4. Monach PA, Warner RL, Lew R, Tómasson G, Specks U, Stone JH, et al. Serum biomarkers of disease activity in longitudinal assessment of patients with ANCA-associated vasculitis. ACR Open Rheumatol. 2022;4:168-76
  5. Mahr A, Moosig F, Neumann T, Szczeklik W, Taillé C, Vaglio A, et al. Eosinophilic granulomatosis with polyangiitis (Churg-Strauss): evolutions in classification, etiopathogenesis, assessment and management. Curr Opin Rheumatol. 2014;26:16-23
  6. Lyons PA, Peters JE, Alberici F, Liley J, Coulson RMR, Astle W, et al. Genome-wide association study of eosinophilic granulomatosis with polyangiitis reveals genomic loci stratified by ANCA status. Nat Commun. 2019;10:5120

Patrick Vanderboom, Ph.D., M.S.

Fellow, Clinical Chemistry
Mayo Clinic

Melissa Snyder, Ph.D.

Consultant, Clinical Biochemistry
Mayo Clinic
Associate Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science

A 56-year-old woman presented for the evaluation of a pelvic mass. PET scan showed intense activity in the mass along the left side of the rectum. She had a prior diagnosis of right tubo-ovarian high-grade serous carcinoma status post hysterectomy and bilateral salpingo-oophorectomy 10 years ago and multiple local recurrences in the past decade. A pelvic mass biopsy was performed, and the hematoxylin and eosin-stained (H&E) slides were reviewed. Several immunohistochemical (IHC) studies were also performed and the pictures are shown below. 

Figure 1: H&E 20x
Figure 2: H&E 40x
Figure 3: H&E 2 20x
Figure 4: Calretinin 20x
Figure 5: CK7
Figure 6: p53 wild type (20X)
Figure 7: WT1

Based on histomorphology and immunohistochemical studies, what is the likely diagnosis? 

  • High-grade serous carcinoma
  • Female adnexal tumor of probable Wolffian origin (FATWO)
  • Ovarian sex-cord stromal tumor
  • STK11 adnexal tumor

The correct answer is ...

STK11 adnexal tumor.

The tumor shows closely packed tubules with solid and sieve-like areas composed of ovoid tumor cells with scant eosinophilic cytoplasm, and nuclear hyperchromasia with conspicuous nucleoli. Occasional pseudo-inclusions are also identified. There is a characteristic background of the myxoid stroma. The neoplastic cells are positive for CK7 (focal), WT1, and calretinin. P53 shows a wild-type staining pattern. CK5, EMA, and SF1 showed rare positive cells. The IHC stains for Pax-8, CDX2, CK20, Napsin A, and S100 are negative. Mayo Clinic solid tumor panel (next-generation sequencing) was also performed that demonstrated an STK11 c.580G>A (p.D194N) (VAF: 91%) mutation. The combined histology along with immunohistochemistry and molecular studies is most consistent with STK11 adnexal tumor.

STK11 adnexal tumor is a novel, usually para tubal, adnexal tumor occurring in females with a median age of 39 years. Very few cases are reported in the literature. Approximately 50% of these tumors are associated with the Peutz-Jeghers syndrome that harbors STK11 mutation, however, the histogenesis is still unclear. Tumors show diverse histological patterns with inter-anastomosing cords and trabeculae. Prominent myxoid stroma imparts a distinct appearance. These tumors are overtly malignant with atypical cytologic features often with prominent nucleoli and a variable mitotic index. Half of these cases are associated with metastatic disease at the time of diagnosis. The tumor variably expresses CK and sex cord-stromal markers (inhibin, calretinin, and WT1). The tumor can be easily confused with other ovarian tumors including high-grade serous carcinoma and female adnexal tumor of probable Wolffian origin (FATWO). 

High-grade serous carcinoma is a malignant epithelial tumor showing serous (tubal type) differentiation with papillary, solid and/or glandular growth and moderate to severe nuclear atypia. It is associated with TP53 mutation (>97%). The tumor showed wild type p53 staining making a diagnosis of high-grade serous carcinoma less likely.

FATWO arises in the broad ligament or ovary and is considered Wolffian in origin. Morphologically, the tumor shows an admixture of hollow to solid tubules, sieve-like cysts, or diffuse growth with interspersed hyalinized bands. Eosinophilic luminal secretions may be present but cytological atypia and mitoses are usually minimal. Rarely, metastasis has been reported. When considering a diagnosis of malignant FATWO, consider the possibility of STK11 adnexal tumor if architectural patterns are not those typical of FATWO.


  1. Bennett JA, et al. A distinctive adnexal (usually paratubal) neoplasm often associated with Peutz-Jeghers syndrome and characterized by STK11 alterations (STK11 adnexal tumor): a report of 22 cases. Am J Surg Pathol. 45(8):1061-1074, August 2021.
  2. WHO Classification of Tumours Editorial Board. Female genital tumours [Internet]. Lyon (France): International Agency for Research on Cancer; 2020. (WHO classification of tumours series, 5th ed.; vol. 4). 

Qandeel Sadiq, M.B.B.S.

Fellow, Surgical Pathology
Mayo Clinic

Maryam Shahi, M.D.

Senior Associate Consultant, Anatomic Pathology
Mayo Clinic
Assistant Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science

A 70-year-old man with no significant medical history presented with progressive headache and visual disturbances. Head imaging demonstrated bilateral, symmetric masses involving the retrobulbar orbital soft tissue. Clinical evaluation showed no evidence of sarcoidosis, lymphoproliferative disease, monoclonal proteins, or elevated serum IgG4. The patient was treated with prednisone and showed partial improvement of the symptoms. A biopsy of the left orbital lesion was performed.

Figure 1: A skull base to mid-thigh PET-CT scan revealed marked, bilateral hypermetabolism of the retrobulbar orbital soft tissue (A, B). There was also extensive osseous hypermetabolism associated with sclerotic change and cortical thickening, predominantly involving the bilateral humeral and femoral heads (C). Increased prominence of the kidneys with perinephric fat stranding and hypermetabolic soft tissue along the perirenal fascia was also noted (C).
Figure 2: Histologic sections demonstrate fibroadipose tissue extensively involved by an atypical histiocytic infiltrate consisting of cells with abundant, foamy cytoplasm admixed with lymphoplasmacytic inflammation and rare eosinophils. Touton giant cells were not identified (A, B). Grocott-Methenamine silver (GMS) and AFB stains resulted negative for fungal and acid-fast bacilli microorganisms, respectively (not shown). By immunohistochemistry, the histiocytic infiltrate is positive for CD163 (C), Factor XIIIa (D), CD68, and cyclin D1 (not shown), while negative for S100 (E) and langerin (not shown). Histiocytes show cytoplasmic BRAFV600E positivity, (F), consistent with this BRAF mutation.
Figure 3: Digital droplet PCR (ddPCR) confirmed the presence of a BRAF p.V600E mutation (asterisks).

The most likely diagnosis is:

  • Adult-onset xanthogranuloma
  • Erdheim-Chester disease
  • IgG4-related disease
  • Langerhans cell histiocytosis

The correct answer is ...

Erdheim-Chester disease.

Erdheim-Chester disease (ECD) is a rare and aggressive histiocytic proliferative disorder that affects most frequently men between the fourth and seventh decades of life. ECD is multisystemic and can affect essentially any organ with relative sparing of liver, spleen, and lymph nodes. Bilateral, symmetric involvement of long bones is characteristic, while perinephric and periaortic infiltration is present in approximately 50% of cases. Pituitary, central nervous system (CNS), lung, heart, and skin involvement can also be seen. ECD may be asymptomatic or may present with symptoms that vary significantly depending on the extent and distribution of the lesions. These include fatigue, fever, long bone pain, diabetes insipidus, neurologic, and respiratory symptoms.

Ophthalmic involvement by ECD was first described by Alper et al in 1983 and occurs in up to 30% of cases. Patients typically present with bilateral exophthalmos, pain, and ophthalmoplegia due to orbital involvement. Other ocular manifestations include palpebral xanthelasmas, anterior uveitis and/or vitritis, edema and atrophy of the optic disk, recurrent retinal detachments, drusen-like deposits, and atrophy of the retinal pigmented epithelium. Imaging studies usually demonstrate hypodense, infiltrating retro-orbital lesions encasing the optic nerve and extraocular muscles. Extraconal and preseptal extension, secondary involvement of the lacrimal gland, and osteosclerosis of skull and facial bones are uncommonly seen.

Histologically, ECD typically consists of an infiltrative histiocytic proliferation composed of cells with abundant, foamy (xanthomatous) cytoplasm admixed with Touton multinucleated giant cells, variable lymphoplasmacytic inflammation, and neutrophils. Background fibrosis is frequently observed and may represent the dominant component, which may lead to misinterpreting it as a reactive process. By immunohistochemistry, ECD histiocytes are positive for CD68 and CD163 and may show Factor XIIIa expression. Langerin, CD1a, S100, and OCT2 are negative. Most ECD cases harbor somatic alterations that result in activation of the mitogen-activated protein kinase (MAPK) cell signalling pathway. Among these, BRAF p.V600E mutation is present in approximately 50%–60% of cases, followed by other BRAF, ARAF, NRAS, KRASMAP2K1, or PIK3CA mutations. Therefore, the BRAFV600E (VE1) antibody is a useful surrogate marker to assess for this BRAF mutation, which can be subsequently confirmed by targeted sequencing.

In the orbit, the differential diagnosis of ECD includes a number of neoplastic and nonneoplastic lesions that are also rich in xanthomatous histiocytes. This group of lesions referred as orbital xanthogranulomatous disease (AXD) include: 1) adult-onset xanthogranuloma (AOX), 2) adult-onset asthma with periocular xanthogranuloma (AAPOX), and 3) necrobiotic xanthogranuloma (NBX). All three lesions show sheets of mononuclear, foamy (xanthomatous) histiocytes admixed with variable amounts of Touton multinucleated giant cells and lymphoplasmacytic inflammation. By immunohistochemistry, these histiocytes are positive for CD68, CD163, and variably for Factor XIIIa, and lack CD1a, langerin, and S100 expression.

AOX presents clinically as a localized process without systemic involvement.

AAPOX, usually bilateral and anteriorly located in the orbit, show in addition to the foamy histiocytes presence of lymphoid follicles. As its name implies, a defining feature of this entity is an association with adult-onset asthma, which may develop concomitantly or months to years following the onset of the ocular symptoms.

NBX shows areas of geographic necrosis surrounded by palisading epithelioid histiocytes and interstitial mucin deposits, which can be highlighted by Alcian blue and mucicarmine stains. NBX presents with indurated nodules and plaques, most commonly involving eyelids and periocular skin, which can ulcerate over time, leading to local tissue destruction. NBX is typically associated with paraproteinemia, multiple myeloma, B-cell lymphoma, and other lymphoproliferative disorders, so regular surveillance of these patients should be granted.

Interestingly, increased numbers of IgG4 positive cells might be seen in association with these lesions raising consideration that there is an association among IgG4 related disease and these entities. When IgG4-related disease is suspected, an increase in absolute numbers of IgG4+ plasma cells and/or increase in the IgG4+/IgG+ plasma cell ratio should be evaluated by immunohistochemistry in the affected tissue, along with clinical and serological correlation.

Lastly, ECD must be distinguished from other histiocytic neoplasms including Langerhans cell histiocytosis (LCH) and Rosai-Dorfman disease (RDD). The presence of CD1a and langerin-positive histiocytes supports the diagnosis of LCH, while strong, diffuse S100 immunoreactivity is consistent with RDD. A subset of cases of ECD can occur concurrently with LCH (so-called overlap syndrome or mixed histiocytosis) or show histologic and immunohistochemical overlap with Rosai-Dorfman disease, with many of these cases also showing mixed clinical and radiological features.

The development of new therapies has drastically improved prognosis of patients with ECD. Nonetheless, prognosis is still influenced by the site and extent of infiltration, with patients showing CNS involvement carrying a worse outcome. Currently, combined targeted therapy with BRAF/MEK-inhibitors is the standard regimen of treatment, as it overcomes the limitations of using monotherapy with BRAF-inhibitors and leads to a better response, significant delay in development of treatment resistance, and longer progression-free survival. The combined use of BRAF/MEK inhibitors may also be beneficial in patients harboring other non-BRAF p.V600E mutations. Although prednisone and azathioprine are at times successful in controlling the disease, they can also delay the diagnosis if ECD is not suspected.

In summary, the diagnosis of ECD must be based on a detailed clinical, radiological, and histopathological analysis. Histologic evaluation is essential not only to confirm the diagnosis, but also to assess for the BRAF gene mutational status, as this carries significant therapeutic implications.


  1. Salomao DR, Van Halteren A. Erdheim-Chester disease. In: WHO Classification of Tumours Editorial Board. Eye tumours [Internet; beta version ahead of print]. Lyon (France): International Agency for Research on Cancer; 2023. (WHO classification of tumours series, 5th ed.; vol. 13). Available from:
  2. Alper MG, Zimmerman LE, Piana FG. Orbital manifestations of Erdheim-Chester disease. Trans Am Ophthalmol Soc. 1983;81:64-85.
  3. Ozkaya N, Rosenblum MK, Durham BH, et al. The histopathology of Erdheim-Chester disease: a comprehensive review of a molecularly characterized cohort. Mod Pathol. 2018 Apr;31(4):581-597.
  4. Kanakis M, Petrou P, Lourida G, Georgalas I. Erdheim-Chester disease: a comprehensive review from the ophthalmologic perspective. Surv Ophthalmol. 2022 Mar-Apr;67(2):388-410.
  5. Sivak-Callcott JA, Rootman J, Rasmussen SL, et al. Adult xanthogranulomatous disease of the orbit and ocular adnexa: new immunohistochemical findings and clinical review. Br J Ophthalmol. 2006 May;90(5):602-8.
  6. McKelvie P, McNab AA, Hardy T, Rathi V. Comparative study of clinical, pathological, radiological, and genetic features of patients with adult ocular adnexal xanthogranulomatous disease, Erdheim-Chester disease, and IgG4-related disease of the orbit/ocular adnexa. Ophthalmic Plast Reconstr Surg. 2017 Mar/Apr;33(2):112-119.
  7. Guo J, Wang J. Adult orbital xanthogranulomatous disease: review of the literature. Arch Pathol Lab Med. 2009 Dec;133(12):1994-7.
  8. Park JK, Huang LC, Kossler AL. Erdheim-Chester disease and vemurafenib: a review of ophthalmic presentations and clinical outcomes. Orbit. 2022 Jun 15:1-12.
Photo of Maria Adelita (Adelita) Vizcaino Villalobos, M.D.

Maria Adelita Vizcaino Villalobos, M.D.

Fellow, Anatomic & Neuropathology
Mayo Clinic

Jorge Trejo-Lopez, M.D.

Senior Associate Consultant, Anatomic Pathology
Mayo Clinic
Assistant Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science

Diva Salomao, M.D.

Consultant, Anatomic Pathology
Mayo Clinic
Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science

A 72-year-old man with history of smoking, cholangiocarcinoma, and urothelial carcinoma underwent a left upper lung lobectomy for invasive adenocarcinoma. An additional tumor, 0.3 x 0.3 x 0.2 cm, was identified in the lobe. H&E photomicrographs of the second tumor are provided below (Figures 1-2). Immunohistochemical studies revealed that the tumor cells stained positive for p40 (staining the basal cell layer, Figure 3), TTF-1 (staining the luminal cell layer, Figure 4) and CK7 (staining both layers).

Figure 1: Low power H&E
Figure 2: High power H&E
Figure 3: p40
Figure 4: TTF-1

What is the correct diagnosis?

  • Peribronchial metaplasia
  • Bronchiolar adenoma
  • Invasive non-mucinous adenocarcinoma of the lung
  • Bronchial papilloma

The correct answer is ...

Bronchiolar adenoma.

Bronchiolar adenomas (BA) are rare, benign lung tumors composed of bronchiolar-type epithelium with continuous basal cell layer. They are located peripherally in the lung and involve the peribronchiolar lung parenchyma. The term bronchiolar adenoma was proposed in 2018 for the entire spectrum of bilayered bronchiolar-type lesions, encompassing the previously described classical and nonclassical ciliated muconodular papillary tumors (CMPT). Like in this case, BA are typically incidental findings and affect middle-aged to elderly patients.

BA can appear as solid, ground-glass or mixed solid/ground-glass on computed tomography (CT). Grossly, they are well-circumscribed, tan-white to grey, with firm, cystic or mucoid cut surface. Histologically, BA are divided into proximal and distal types, based on similarity to normal proximal and distal bronchiolar epithelium. Proximal BA correspond to classic CMPT, and distal BA correspond to nonclassical CMPT. Proximal type BA are composed of mucinous and ciliated luminal cells. They can have a spectrum of architectural patterns, ranging from predominantly papillary to flat. Abundant intratumoral or intraalveolar mucin is characteristic. Distal type BA are predominantly composed of non-ciliated luminal cells, resembling type II pneumocytes, and club cells. They are mostly flat. The basal cell layer is highlighted by basal cell markers, p40, p63 or CK5/6. The luminal cells in distal type BA are more likely to be positive for TTF-1 than in proximal type BA.

The etiology of BA is unknown. Studies have shown BRAF V600E being the most common driver mutation with KRAS being the second most common. The detection of driver mutations, however, is not necessary for diagnosis.

The diagnosis of BA can be challenging during intraoperative consultation and on small biopsies. The distinction from adenocarcinoma is important and can be difficult with limited samples. The differential diagnosis of BA depends on the predominant cell type of the lesion (proximal vs. distal). The most useful features in differentiating BA from mucinous and non-mucinous adenocarcinoma of the lung are the continuous basal cell layer (that can be highlighted by basal cell markers p40, p63 or CK5/6), ciliated cells, and the lack of nuclear atypia. Location and presence of well-defined borders can help differentiate BA from other benign lesions such as bronchial papillomas and peribronchial metaplasia.


  1. Nicholson AG, Travis WD, et al. Bronchiolar adenoma/ciliated muconodular papillary tumor. In: WHO Classification of Tumours Editorial Board. Thoracic Tumours. Lyon (France): International Agency for Research on Cancer; 2021.
  2. Chang JC, Montecalvo J, et al. Bronchiolar adenoma: Expansion of the concept of ciliated muconodular papillary tumors with proposal for revised terminology based on morphologic, immunophenotypic, and genomic analysis of 25 cases. Am J Surg Pathol. 2018;42:1010-1026.
  3. Shirsat H, Zhou F et al. Bronchiloar adenoma / pulmonary ciliated muconodular papillary tumor: Potential pitfall on frozen sections. Am J Clin Pathol. 2021;155:832-844.
  4. Krishnamurthy K, Kochiyil J et al. Bronchiolar adenomas (BA): A detailed radio-pathologic analysis of six cases and review of literature. Ann Diagn Pathol. 2021;55:151837

Alma Oskarsdottir, M.D.

Resident, Anatomic & Clinical Pathology
Mayo Clinic

Joanne (Eunhee) Yi, M.D.

Consultant, Anatomic Pathology
Mayo Clinic
Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science

A 70-year-old woman with a medical history significant for gastrointestinal adenocarcinoma presented to her primary care provider. She underwent a routine chest X-ray. A small nodule in the lower lobe of the right lung was identified, which was not present on her previous chest X-ray five years earlier. CT-imaging showed a peripherally located 7 mm irregular, solid mass. The mass was biopsied and histologic sections showed a papillary lesion lined by mucinous columnar cells with mild acute inflammation.

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5

What is the most likely diagnosis?

  • Bronchiolar adenoma/ciliated muconodular papillary tumor
  • Metastatic colorectal adenocarcinoma with mucinous features
  • Mucinous adenocarcinoma
  • Peribronchiolar metaplasia

The correct answer is ...

Bronchiolar adenoma/ciliated muconodular papillary tumor.

Bronchiolar adenoma/ciliated muconodular papillary tumor (BA/CMPT) is a benign pulmonary neoplasm. Clinically, BA/CMPT typically occurs in individuals 60 years of age and older and equally affects both sexes. This lesion is most often discovered incidentally on imaging and presents as either a solid nodule or ground-glass opacity in the peripheral lung. Most BA/CMPTs measure between 0.5-1.5 cm. Irregular borders and occasional central cavitation seen on imaging may raise concern for malignancy.

BA/CMPT can be further subdivided into either classic CMPT (demonstrated in this case) and non-classic CMPT. Classic CMPT was first described in 2002 and is characterized by a cellular bilayer consisting of papillary or glandular ciliated columnar epithelium with a continuous layer of basal cells underlying the epithelium. Epithelial cells appear bland and often contain mucin, though this is not required for diagnosis. Non-classic CMPT more closely resembles distal airways with a bilayer consisting of cuboidal, club-like epithelium without cilia or mucin and a continuous basal cell layer.

Molecularly, both classic and non-classic BA/CMPT are positive for BRAF V600E mutations in approximately 40% of cases. Studies have also shown alterations in EGFR, KRAS, HRAS, ALK, and AKT1 genes, though these mutations have not demonstrated as high a prevalence as BRAF V600E.

Key differential diagnoses in these cases include invasive mucinous adenocarcinoma, adenocarcinoma in situ, mucoepidermoid carcinoma, peribronchiolar metaplasia, and metastases. The most important distinguishing feature in BA/CMPT is the presence of a continuous basal cell layer, which stains positive for p40/p63. In adenocarcinoma and mucoepidermoid carcinoma, a continuous basal cell layer will be absent. Additionally, ciliated cells will not be seen in these malignancies but are apparent in classical BA/CMPT. Unlike BA/CMPT, peribronchiolar metaplasia will not form a discrete nodule or opacity, arises in the setting of inflammation or injury, and is often multifocal. Positive TTF-1 staining is useful in establishing pulmonary origin and targeted immunostains can be used to rule out possible metastatic disease (CDX2 was used in this case to rule out an intestinal primary).


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Brittney Thiele, M.D.

Resident, Anatomic & Clinical Pathology
Mayo Clinic

Melanie Bois, M.D.

Consultant, Anatomic Pathology
Mayo Clinic
Assistant Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science

MCL Education

This post was developed by our Education and Technical Publications Team.