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Papers by Ana M Abreu Velez

In many countries and laboratories, techniques such as direct immunoﬂ uorescence (DIF) are not a... more In many countries and laboratories, techniques such as direct immunoﬂ uorescence (DIF) are not available for the diagnosis of skin diseases. Thus, these laboratories are limited in the full diagnoses of autoimmune skin diseases, vasculitis, and rheumatologic diseases. In our experience with these diseases and the patient’s skin biopsies, we have noted a positive correlation between periodic acid-Schiff (PAS) staining and immunoﬂ uorescence patterns; however, these were just empiric observations. In the current study, we aim to conﬁ rm these observations, given the concept that the majority of autoantibodies are glycoproteins and should thus be recognized by PAS staining. Aims: To compare direct immunoﬂ uorescent and PAS staining, in multiple autoimmune diseases that are known to exhibit speciﬁ c direct immunoﬂ uorescent patterns. Materials and Methods: We studied multiple autoimmune skin diseases: Five cases of bullous pemphigoid, ﬁ ve cases of pemphigus vulgaris, ten cases of cutaneous lupus, ten cases of autoimmune vasculitis, ten cases of lichen planus (LP), and ﬁ ve cases of cutaneous drug reactions (including one case of erythema multiforme). In addition, we utilized 45 normal skin control specimens from plastic surgery reductions. Results: We found a 98% positive correlation between DIF and PAS staining patterns over all the disease samples. Conclusion: We recommend that laboratories without access to DIF always perform PAS staining in addition to hematoxylin and eosin (H&E) staining, for a review of the reactivity pattern

Background: The clinical presentation of bullous pemphigoid (BP) is variable; blistering skin les... more Background: The clinical presentation of bullous pemphigoid (BP) is variable; blistering skin lesions may be present, but an urticarial or erythematous rash may also precede the appearance of the blisters. The patients themselves may also traumatize blister lesions, and spontaneous ulceration of the skin may occur. Aim: We sought to compare the immune changes in intact bullous pemphigoid lesions, versus ulcerated lesions. Here we aim to describe these changes, utilizing a skin biopsy containing both intact and ulcerated bullous pemphigoid lesional areas via hematoxylin and eosin histology (H&E), as well as direct immunofluorescence (DIF) and immunohistochemistry (IHC). Results: The findings in these areas demonstrated distinctly different patterns of the immune response. In the intact blister areas, markers such as HLA-DP, DQ, DR antigen, cyclooxygenase-2 (COX-2), B-cell lymphoma 2(BCL2), CD3, CD68, alpha-1 antitrypsin, mast cell tryptase, von Willembrand factor and Factor XIIIa demonstrated positive staining in some areas of the blister and around adjacent dermal blood vessels. However, in ulcerated areas, most of these markers primarily compartmentalized in a linear manner at the base of the ulcer. We interpreted the ulcerated area pattern as evidence of the immune system attempting to phagocytose or extrude the ulcerated tissue.

Background: Bullous pemphigoid (BP) is an acquired autoimmune disease characterized by subepiderm... more Background: Bullous pemphigoid (BP) is an acquired autoimmune disease characterized by subepidermal vesicles, with clinical macules and bullae. In contradistinction, drug induced bullous pemphigoid (DBP) may be triggered by medications and other agents. Presently, minimal pathologic differences have been documented to differentiate between these two entities. We present a case of a drug-induced bullous pemphigoid-like reaction, and review critical aspects that seem to differentiate these entities. Case report: A 68 year old male was consulted for the presence of erythematosus plaques, papules and tense blisters on the abdomen and thighs, after the intake of multiple medications. Materials and Methods: Skin biopsies were taken for hematoxylin and eosin review and immunohistochemistry, and for direct immunofluorescence studies.

Bullous pemphigoid (BP) is a bullous disease of autoimmune origin. These disorders are the result... more Bullous pemphigoid (BP) is a bullous disease of autoimmune origin. These disorders are the result of an inflammatory process that causes skin lesions, with local increases of pro-inflammatory mediators. Alterations of the vessels, nerves or/or neurovascular structures have been previously described in BP. Here we aim to confirm reactivity to vessels and nerves in a case of BP. Case report: A 68 year old Caucasian male presented with a sudden appearance of erythematous papules, plaques and few blisters on the abdomen, back, arms, and thighs; focal excoriations were present, with some itching and burning sensations. Materials and Methods: Skin biopsies were taken for hematoxylin and eosin (H&E) staining and immunohistochemistry (IHC), and for

Subcorneal pustular dermatosis (SPD) represents a chronic, relapsing sterile pustular eruption, ... more Subcorneal pustular dermatosis (SPD) represents a chronic, relapsing sterile pustular eruption, involving the trunk and flexoral proximal extremities. A 54-year-old female presented with recurrent, flaccid pustules measuring several millimeters in diameter, on normal and mildly erythematous skin of the groin and submammary areas. Biopsies for hematoxylin and eosin (H&E) examination, direct immunofluorescence (DIF) and immunohistochemistry (IHC) analysis were performed. The H&E staining demonstrated typical features of SPD, including some damage within dermal pilosebaceous units subjacent to the subcorneal blistering process. DIF revealed strong deposits of immunoreactants IgG, IgM, fibrinogen and complement/C3, present in a shaggy pattern within the subcorneal disease areas; in focal, areas of the basement membrane junction and in focal pericytoplasmic areas of epidermal keratinocytes. IHC revealed strong positivity to HLA-DPDQDR, mast cell tryptase, CD68, and ZAP-70 in the subcorneal inflammatory infiltrate, and surrounding dermal blood vessels. Myeloperoxidase was also positive. Positive staining with the anti-ribosomal protein S6-pS240 at the edges of hair follicles and sebaceous glands subjacent to the subcorneal blisters was also noted. Conclusions: We conclude that this disorder may have several components in its etiopathology, including a possible restricted immune response and a possible genetic component; these possibilities warrant further investigation.

Introduction: Autoimmune bullous skin diseases (ABDs) represent a group of disorders of the skin ... more Introduction: Autoimmune bullous skin diseases (ABDs) represent a group of disorders of the skin and mucosa commonly associated with deposits of immunoglobulins, complement and fibrinogen, and usually directed against distinct adhesion molecules. After studing these diseases for many years, we noted alterations not only between the cells junctions of the epidermis and/or the dermal/epidermal junction, but also in dermal skin appendageal structures and in mesenchymal tissue around the blisters. Based on our findings, we wanted to determine if the observed patterns of autoimmunity correlated with cutaneous vimentin expression. Materials and Methods: Archival biopsies previously diagnosed with ABDs by clinical, hematoxylin and eosin (H&E) and direct and/or immunofluorescence data were stained with antibodies directed against vimentin via immunohistochemistry (IHC). We tested 30 patients affected by endemic pemphigus, 30 controls from the endemic area, and 15 normal controls. We also tested 30 biopsies from patients with bullous pemphigoid (BP), 20 with pemphigus vulgaris (PV), 8 with pemphigus foliaceus, 14 with dermatitis herpetiformis (DH) and 3 with Senear-Usher syndrome. Results: The H&E, DIF and vimentin patterns of positivity in the different ABDs confirmed that vimentin was compartmentalized around areas of dermal inflammation, around skin appendages and in epidermal, dermal and mesenchymal cell junction areas. Conclusion: Vimentin may be a useful tool for highlighting patterns of microenvironmental tissue alteration in multiple ABDs. The vimentin staining pattern observed was analogous to that we have previously described for proteases and protease inhibitors in patients affected by ABDs, expanding the concept that the autoimmune process extends beyond cell junctions. Abbreviations: Autoimmune bullous diseases (ABDs), bullous pemphigoid (BP), pemphigus vulgaris (PV), pemphigus foliaceus (PF), der-matitis herpetiformis (DH), endemic pemphigus foliaceus (EPF), linear IgA disease (LAD), immunohistochemistry (IHC), direct and indirect immunofluorescence (DIF, IIF), hematoxylin and eosin (H&E), basement membrane zone (BMZ), intercellular staining between keratinocytes (ICS), intermediate filament (IF), epithelial-to-mesenchymal transition (EMT).

Introduction: Pemphigus foliaceus (PF) is endemic in some South American countries, especially in... more Introduction: Pemphigus foliaceus (PF) is endemic in some South American countries, especially in Colombia and Brazil; in Brazil, it is also known as fogo selvagem (FS). We aimed to study the presence of mast cells and the expression of the mast/stem cell growth factor receptor (c-kit/CD117) in PF skin biopsies, as well as the role of IgE in the disease pathogenesis. Methods: Forty-four skin biopsies from patients affected by endemic PF (EPF) (30 patients from El Bagre, Colombia, and 14 from the northeastern region of São Paulo State, Brazil), 48 control biopsies from Colombian and Brazilian endemic areas, and additional control biopsies from none endemic areas in Colombia and the USA non were studied. Immunohistochemistry (IHC) was performed to evaluate skin biopsies with anti-mast cell tryptase (MCT), anti-c-kit and anti-IgE antibodies. We also searched for serum IgE in 30 EPF and 30 non-atopic controls from the El Bagre region via ELISA. In our El Bagre patients and controls, we also searched for IgE in skin samples by direct immunofluorescence. Results: In 100% of the EPF biopsies, MCT, c-kit and IgE were identified with stronger expression relative to control biopsies, especially in the inflammatory infiltrates around upper dermal blood vessels and dermal eccrine glands. IgE staining was positive along the BMZ in some EPF skin samples. The DIF results confirmed a linear deposition of IgE at the BMZ. Increased IgE serum levels were also noted in PF patients relative to controls.. Conclusions: In patients with EPF, the observed increased expression of MCT, c-kit and IgE in lesional skin, associated with higher serum IgE levels may indicate possible IgE participation in the antigenic response.

En muchas enfermedades dermatológicas se presentan ampollas, pero no todas son de etiología autoi... more En muchas enfermedades dermatológicas se presentan ampollas, pero no todas son de etiología autoinmune. Para el estudio de las enfermedades ampollosas se deben tener en cuenta las manifestaciones clínicas, la historia de cómo y cuándo empezaron las ampollas, las caracterís-ticas epidemiológicas e histológicas (por ejemplo, el nivel de la piel en el que se producen las ampollas) y la presencia o no de infiltrados inflamatorios. Para corroborar la etiología autoin-mune de la enfermedad ampollosa es importante contar con los resultados de pruebas como la inmunofluorescencia directa e indirecta, el inmunoblotting, el ensayo inmunoenzimático (ELISA), la inmunoprecipitación y la microscopía electrónica. La información sobre los títulos séricos de autoanticuerpos ayuda a orientar mejor el tratamiento inmunosupresor. SUMMARY Autoimmune blistering diseases of the pemphigus group Blisters may appear in many dermatological diseases, but they are not necessarily of autoim-mune etiology. For the study of blistering diseases, it is necessary to take into account the clinical aspects, the history of when and how blisters appeared, the epidemiological and histological information (for instance, the skin level at which blisters are located), and whether inflammatory infiltrates are present. In order to corroborate the autoimmune etiology of blisters, it is important to have the results of confirmatory tests such as direct and indirect immunofluorescence, immune blotting, enzyme-linked immune-assay (ELISA), immune precipitation , and electronic microscopy. Information on autoantibodies serum titers may help to conduct a more precise immunosuppressive therapy.

A clinical pruritus and inflammation is present following most insect bites. Here we document imm... more A clinical pruritus and inflammation is present following most insect bites. Here we document immunoreactivity to Meissner corpuscles and dermal nerves following arthropod bites.

Erythema multiforme is an acute, often self-limited and occasionally lethal syndrome with distinc... more Erythema multiforme is an acute, often self-limited and occasionally lethal syndrome with distinctive skin lesions and/or mucosal lesions; the disease has multiple triggers, including medications. We describe a 68 year old African American female on multiple medications; the patient suddenly presented with recurrent, flaccid bullae and erythematous circular lesions on her face, neck, genitals, hands and legs. Biopsies for hematoxylin and eosin (H&E) examination, direct immunofluorescence (DIF) and immunohistochemistry (IHC) stains were performed. The histology displayed erythema multiforme with epidermal blisters, defragmented melanocytes and other cell debris inside the blister. DIF revealed deposits of IgG, IgM, IgA, IgD, Complement/C3, C4, and fibrinogen within the epidermis; and around upper dermal vessels. Anti-neutrophil cytoplasmic antibodies (c-ANCAS) were present as part of “large round bodies” inside the blisters, and as part of both cytoid and colloid bodies. Staining for Von Willembrand factor, CD68, myeloperoxidase and CD45 was seen on cell fragments within the blisters, as well as within the inflammatory infiltrate in the dermis. In this case cystoid, colloid and “large round bodies” may represent amalgamations of multiple cell fragments, with immunoglobulins and complement. Overall, a strong immunologic response was observed with c-ANCAs, caused by multiple medications.

Eczema herpeticum is an entity usually seen in pediatric and young adult patients, and presents p... more Eczema herpeticum is an entity usually seen in pediatric and young adult patients, and presents perils of systemic compromise and a ten percent mortality rate [1,2]. We describe a 53 year old Caucasian female with a childhood history of atopy who consulted her dermatologist for a sudden presentation of generalized, annular, scaly, itchy, erythematous plaques with some raised ridges and blistering; these lesions were observed concurrent with malaise (Fig. 1a, black arrow). Our study was performed accordance with the Code of Ethics of the World Medical Association (Declaration of Helsinki); informed consent was obtained. Lesional skin biopsies were taken for hematoxylin and eosin (H&E) review, direct immunofluorescence (DIF) and immunohistochemistry (IHC) staining. Review of the H&E sections demonstrated an infectious process, suggestive of a herpes virus infection. Focal areas of the epidermis displayed ballooning of keratinocytes, with margination of cell chromatin and multinucleated cells seen inside an epidermal blister (Fig. 1b, black arrows) (400x). The entire dermis was edematous, including the appendageal structures. A multilevel epidermal blistering process was present. Within the blister lumen, numerous neutrophils, eosinophils and Langerhans histiocytes were also noted. Inflammatory cell debris and acantholytic keratinocytes were also present within the blister lumen. In the subjacent dermis, a superficial, perivascular, mild mixed inflammatory infiltrate was present, featuring numerous lymphocytes, histiocytes, neutrophils and eosinophils. No frank vasculitis was noted. Our DIF was performed as previously described [3], and demonstrated positivity to the lateral aspects of arrector pili muscles with FITC conjugated anti-human fibrinogen antibodies (Fig. 1c, green staining; white arrow). Other findings included IgG (+/-, dermal eccrine glands); IgA (+/-, superficial dermal perivascular); Complement/C1q (+/-, superficial dermal perieccrine); Complement/C3 (+, BMZ granular); Complement/ C5b-9(+/-, superficial dermal perivascular); albumin (+, superficial dermal eccrine); and fibrinogen (+, superficial dermal interstitial).

Allergies and autoimmune diseases may both be considered hyper-immune responses, where the body’s... more Allergies and autoimmune diseases may both be considered hyper-immune responses, where the body’s immune system becomes supercharged and attacks or responds to inappropriate antigens. We describe a skin rash with a mixed allergic and autoimmune host response. A 65 year old female consulted her dermatologist for a pruritic rash. The patient had taken many medications without improvement of the rash, and lived in an area affected by environmental spills; other patients had presented with similar rashes concurrently. A clinical evaluation was performed, and skin biopsies were obtained for hematoxylin and eosin (H&E) examination, as well as for immunohistochemical (IHC) and direct immunofluorescence (DIF) studies. The H&E review revealed a mild, superficial, perivascular dermal infiltrate of lymphocytes, histiocytes, mast cells and eosinophils. Dermal sclerodermoid alterations were also noted. A mild peripheral blood eosinophilia was found; cutaneous IHC staining revealed staining for anti-HLA-DP, DQ, DR antigen and Complement/C5b-9/MAC, in the areas of the perivascular infiltrate and the sclerodermoid changes. The DIF confirmed these findings. Our case is characterized by a mixed allergic/autoimmune reaction, which did not fit exclusively into any single Gell Coombs immune response category.

The subepidermal vesiculobullous disorders include a wide variety of pathogenically unrelated ent... more The subepidermal vesiculobullous disorders include a wide variety of pathogenically unrelated entities, which share the formation of clefts or bullae. A 59 year old female presented with a sudden eruption of pruritic skin vesicles and blisters on several areas of her body. The patient was taking multiple medications. We decided to test for the expression of p0071 and ARVCF because they are linked with tight, adherens, and occludens cell junctions in the epidermis and the dermis, and study if these molecules participate in the bullae formation. Skin biopsies were taken from lesional skin and were tested by hematoxylin and eosin(H&E) staining, as well as via immunohistochemistry (IHC) and direct immunofluorescence(DIF) using multiple antibodies. ARVCF and p0071 were overexpressed in the epidermis, and in dermal cell junctions around the blisters along with ribosomal protein S6-pS240, Factor XIIIa, CD15, CD45, multiple immunoglobulins, complement, fibrinogen and HLA-DP, DQ, DR antigen. All these molecules were also overexpressed around dermal vessels, eccrine glands and in neurovascular cell junctions below the blister. A normal control did not display the overexpression. Drug reactions may cause blisters; regional cell junctions may be altered, as demonstrated by overexpression of ARVCF and p0071. The overexpression likely contributes to passage of immunologic cells and formation of edema, directly contributing to blister formation.

Fillers, including Gore-Tex, have been long utilized for reconstructive procedures as well as for... more Fillers, including Gore-Tex, have been long utilized for reconstructive procedures as well as for esthetic purposes. We report a 46 year female, who presented to the plastic surgeon to remove a previous nasolabial implant. The tissue was removed and examined using hematoxylin and eosin staining. Three histologic patterns were seen: 1) encapsulation around a large part of the material without giant cells, nor inflammation; 2) peripheral colonization by fibrous tissue, blending with normal soft tissue, and 3) thickening of adjacent skin. We thus document a mixed histologic pattern, featuring a partial peri-implant fibrous capsule, fibrous tissue merging with surrounding normal soft tissue and focal skin thickening.

Background. We identified a new variant of endemic pemphigus foliaceus in El Bagre, Colombia, Sou... more Background. We identified a new variant of endemic pemphigus foliaceus in El Bagre, Colombia, South America, which we term El Bagre-EPF, and observed reactivity to arrector pili muscle (APM), thus we tested for autoimmunity to APM. Methods. We took skin biopsies from 30 patients with El Bagre-EPF and 30 healthy controls (HCs) matched by age, sex and occupation, who were all from the endemic area, and tested these using direct immunofluorescence (DIF), confocal microscopy, immunohistochemistry and immunoblotting (IB). Results. Of the 30 patients with El Bagre-EPF, 27 had autoantibodies to APM that colocalized with commercial antibodies to myocardium-enriched zonula occludens-1-associated protein (MYZAP), desmoplakin (DP)1 and DP2, plakophilin 4, and Arma-dillo repeat gene deleted in velo-cardio-facial syndrome (ARVCF) (P < 0.001, Fisher exact test). The positive staining also colocalized with Junctional Adhesion Molecule 1 (JAM-A), a control antibody for gap cell junctions. No HC samples were positive. In 27 of the 30 patients, serum that was APM-positive also displayed IB colocalization of their autoantibody molecular weights with the Progen antibodies (P < 0.001, Fisher exact test). Conclusions. Patients affected by El Bagre-EPF have autoantibodies to APM, co-localizing with the antibodies MYZAP, ARVCF, p0071, DP1 and DP2, suggesting that these molecules are El Bagre-EPF antigens. Further, all of these antigens represent components of cell junctions, indicating that the immune response is directed, at least partially, against cell junctions. The immune response in patients affected by El Bagre-EPF is polyclonal, and it includes B and T lymphocytes,

Background We previously described a new variant of endemic pemphigus foliaceus in El Bagre, Colo... more Background We previously described a new variant of endemic pemphigus foliaceus in El Bagre, Colombia (El Bagre-EPF).

Citation: Abreu-Velez AM, Gao W, Howard MS. Patients affected by endemic pemphigus foliaceus in C... more Citation: Abreu-Velez AM, Gao W, Howard MS. Patients affected by endemic pemphigus foliaceus in Colombia, South America exhibit autoantibodies to optic nerve sheath envelope cell junctions. Dermatol Pract Concept. 2018;8(1):1-6. Background: The majority of the patients affected by a new variant of endemic pemphigus foliaceus in El Bagre, Colombia (El Bagre EPF or pemphigus Abreu-Manu), have experienced vision problems; we have previously reported several ocular abnormalities. Methods: Here, we aimed to investigate reactivity to optic nerves in these patients. We utilized bovine, rat and mouse optic nerves, and performed immunofluorescence and confocal microscopy to test for optical nerve autoreactivity. We tested 45 patients affected by this disease and 45 controls from the endemic area matched by age, sex and work activity. Results: Overall, 37 of the 45 patient sera reacted to the optic nerve envelope that is composed of leptomeninges; the reactivity was polyclonal and present mostly at the cell junctions (P < 0.001). The immune response was directed against optic nerve sheath cell junctions and the vessels inside it, as well as other molecules inside the nerve. No control cases were positive. Of interest, all the patient autoantibodies co-localized with commercial antibodies to desmoplakins I–II, myocardium-enriched zonula occludens-1-associated protein (MYZAP), armadillo repeat gene deleted in velo-cardio-facial syndrome (ARVCF), and plakophilin-4 (p0071) from Progen Biotechnik (P < 0.001). Conclusion: We conclude that the majority of the patients affected by pemphigus Abreu-Manu have autoantibodies to optic nerve sheath envelope cell junctions. These antibodies also co-localize with ar-madillo repeat gene deleted in velo-cardio-facial syndrome, p0071 and desmoplakins I–II. The clinical significance of our findings remains unknown.

Journal of the American Academy of Dermatology, 2003

Background: Endemic forms of pemphigus are a unique group of autoimmune diseases that represent o... more Background: Endemic forms of pemphigus are a unique group of autoimmune diseases that represent opportunities to study interactions of the environment and genetics with the immune system. The restriction to relatively well-defined regions of South and Central America and perhaps Africa characterizes these diseases.

Background: In many countries and laboratories, techniques such as direct immunoﬂ uorescence (DIF... more Background: In many countries and laboratories, techniques such as direct immunoﬂ uorescence (DIF) are not available for the diagnosis of skin diseases. Thus, these laboratories are limited in the full diagnoses of autoimmune skin diseases, vasculitis, and rheumatologic diseases. In our experience with these diseases and the patient’s skin biopsies, we have noted a positive correlation between periodic acid-Schiff (PAS) staining and immunoﬂ uorescence patterns; however, these were just empiric observations. In the current study, we aim to conﬁ rm these observations, given the concept that the majority of autoantibodies are glycoproteins and should thus be recognized by PAS staining. Aims: To compare direct immunoﬂ uorescent and PAS staining, in multiple autoimmune diseases that are known to exhibit speciﬁ c direct immunoﬂ uorescent patterns. Materials and Methods: We studied multiple autoimmune skin diseases: Five cases of bullous pemphigoid, ﬁ ve cases of pemphigus vulgaris, ten cases of cutaneous lupus, ten cases of autoimmune vasculitis, ten cases of lichen planus (LP), and ﬁ ve cases of cutaneous drug reactions (including one case of erythema multiforme). In addition, we utilized 45 normal skin control specimens from plastic surgery reductions. Results: We found a 98% positive correlation between DIF and PAS staining patterns over all the disease samples. Conclusion: We recommend that laboratories without access to DIF always perform PAS staining in addition to hematoxylin and eosin (H&E) staining, for a review of the reactivity pattern.

Background: Etanercept is often used for treating patients with plaque psoriasis and psoriatic ar... more Background: Etanercept is often used for treating patients with plaque psoriasis and psoriatic arthritis. Case report:

Journal of the American Academy of Dermatology, 2003

Bullous pemphigoid (BP) is a rare skin condition with tense, fluid-filled blisters, and urticaria... more Bullous pemphigoid (BP) is a rare skin condition with tense, fluid-filled blisters, and urticarial or other lesions on areas of the lower abdomen, upper thighs or armpits (flexoral areas). BP has highest incidence in people older than 60. In BP, the immune system attacks the basement membrane zone (BMZ) of the skin at the junction of the epidermis and dermis; however, current data indicates that the BP autoimmune pathology may also attack dermal blood vessels, some stromal dermal areas, nerves, sweat glands and the BMZs of skin adnexal structures. The BP pathologic immune response that initially was thought to be predominately focused on IgG and Complement/C3 deposition at the cutaneous BMZ; however, recent data indicates that B and T lymphocytes, other immune system cells and inflammatory markers may be involved. In some cases, clinical lesions resembling classic BP can be triggered by taking medications; however, the immunopathogenesis of this disorder seems to be quite different. In BP, direct and indirect immunofluorescence of the skin usually shows reactivity in a linear pattern along the BMZ with IgG, Complement/C3 and sometimes IgE; other immunoglobulins and complement may also be present. Multiple markers have been also described in the BP blisters; however, the data is not always consistent. A number of immunoblotting (IB) analyses have indicated that two major antigenic proteins of epidermal extracts are targets in BP; specifically, the 230 kilodalton (kDa) BP antigen I (BP230 or BPAG1), and the 180-kDa BP antigen II (BP180, BPAG2 or Type XVII collagen). These antigenic proteins are detected by sera from patients with BP in various IB patterns; however, other