Extensive variant of cutaneous amyloidosis: report of a case with electron-microscopic and immunohistochemical studies of the basement membrane zone at sites of amyloid production.

A 60-year-old Japanese female developed widespread lichenoid eruptions with pigmentation, which initially appeared in preceding erythematous skin lesions due to dermatomyositis. Thioflavine T and Dylon stainings, electron microscopy and immunohistochemistry revealed that thick amyloid deposits were present in the papillary dermis particularly beneath the epidermis. Autopsy showed no evidence of systemic amyloidosis. Electron microscopy of the lesional skin disclosed the disturbance of lamina densa formation in the epidermal basement membrane zone (BMZ). There was disruption and dissociation of the lamina densa from the basal cell, and a lamina-densa-like substance was found in the amyloid deposits. Immunofluorescence and immunoelectron microscopy showed that type IV and VII collagens, LDA-1 antigen (a noncollagenous component of the BMZ) and laminin were distributed in irregular thick deposits along the BMZ and were also present within the amyloid itself. These findings indicate that morphological and immunohistochemical abnormalities of the lamina densa may be involved in amyloid production at the interface of the epidermis and dermis, at least in this case.     

Bullous amyloidosis: the mechanism of blister formation revealed by electron microscopy

BACKGROUND: Few electron microscopic studies of blister lesions in bullous amyloidosis have been reported, and the mechanism of blister formation remains to be elucidated. This study was designed to examine the nature of bullous amyloidosis ultrastructurally, and clarify the pathogenesis of blister formation. METHODS: We examined a 47-year-old woman with IgD-lambda type myeloma, suffering from bullous lesions on her hands and feet caused by trauma or rubbing. Light and electron microscopic studies were performed. RESULT: Ultrastructurally, amyloid deposits aggregated under the lamina densa. Keratinocyte protrusions penetrated the dermis through the gap in the lamina densa and enfolded amyloid deposits. Amyloid globules were found in enlarged intercellular spaces of keratinocytes. Desmosomes were sparsely distributed in some areas of the epidermis. CONCLUSION: These results indicate that keratinocytes enfold the amyloid globules and take them in the intercellular space of epidermis, and that the breakdown of the lamina densa and widening of the intercellular space between keratinocytes induce skin fragility. Trauma or rubbing of her hands and feet appears to act as the localized precipitating factor of blister formation in bullous amyloidosis.     

Bullous poikilodermatitic amyloidosis of the skin with junctional bulla development in IgG light chain plasmacytoma of the lambda type. Histology, immunohistology and electron microscopy]

A 75-year-old patient presented with bullae, poilokilodermatic skin and nail dystrophy as signs of systemic amyloidosis 1 year before an IgG myeloma of the lambda type was diagnosed. The skin lesions appeared at mechanically irritated locations on the trunk and at the tensor sites of the extremities. Histology showed a subepidermal blister and necrotic keratinocytes in the epidermis. There were amyloid deposits in the papillary dermis perivascular, and in the deep dermis around eccrine glands and in arrectores pilorum muscles. Polyclonal antibodies allowed classification of the deposits as amyloid composed of lambda light-chain immunoglobulins. Electron microscopy revealed globoid deposits of non-branching filaments typical of amyloid. The blister was formed at the level of the lamina lucida, with the lamina densa at the base of the bottom of the bulla. So far, junctional blister formation in bullous amyloidosis of the skin has been described only once. Our case is the second report of this blister type, and to our knowledge the first published report of a junctional blister in myeloma-associated systemic amyloidosis.     

Amyloidogenesis in a case of cutaneous amyloidosis: an electron microscopic study using the refixation-reembedding method

A case of the hyperkeratotic type of macular amyloidosis was electron microscopically examined using the refixation-reembedding method. All the cytoid bodies seen in the dermis were amyloid islets associated with malformed basal lamina-like material. The uppermost amyloid islets were in close apposition to the basal cells, which had neither hemidesmosomes nor intact basal lamina. Anchoring fibril-like filaments were seen between the amyloid islets and the basal lamina-like material. Many fibrillar bodies and clusters were present in the spinous layer, but not in the dermis. Masses of eosinophilic, PAS-positive ovoid bodies seen in the horny layer were tightly packed fibrillar bodies shifted from the spinous layer to the horny layer. These findings suggest that the disturbed basal cells produce an amyloid substance in the dermis, as well as malformed basal lamina. The colloid bodies formed by the degeneration of the basal cells seemed to shift up to the horny layer but not to contribute to the amyloidogenesis in this case.     

Alterations of basement membrane zone in bullous and non-bullous variants of extragenital lichen sclerosus

The aim of the study was to compare alterations of various regions of the basement membrane zone (BMZ) in lichen sclerosus (LS) using laser scanning confocal microscopy. The study included three cases of bullous LS, one case of bullous LS that developed in the course of graft-versus-host disease (GVHD), and six cases of non-bullous LS. Three cases of morphea served as a control. Biopsies from patients' skin and control biopsies from normal human skin were cut into 30-microm thick slides and labeled with antibodies against beta4-intergin (lamina lucida marker), collagen IV, and the N-terminal end of collagen VII (lamina densa markers) and the C-terminal end of collagen VII (sublamina densa marker) using routine immunofluorescence (IF). Three-dimensional (3D) reconstruction of various regions of the BMZ showed a decrease in the number and size of the dermal papillae in LS and morphea as compared with normal skin. In LS numerous invaginations and holes were present in the BMZ at the level of the lamina lucida and lamina densa. Computer animation of 3D projections revealed that the thickness of the lamina densa observed under the light microscopy is an optical artifact dependent on periodical tortion of the lamina densa along its axis. Torsions and invaginations of the BMZ are equally responsible for the phenomenon of artificial reduplication of the lamina densa observed at the ultrastructural level. IF labeling with antibody against the N-terminal end of collagen VII disclosed the presence of a large hole (up to 25 microm) in the lamina densa and the presence of granular material in deep dermis suggestive of partial degradation of lamina densa at the level of anchoring fibers. An IF mapping study showed blister formation below the lamina densa in three patients with bullous LS, whereas in a case of LS associated with GVHD, a blister formed through the basal layer of the epidermis. In morphea, there was flattening of BMZ at the level of lamina lucida, lamina densa, and sublamina densa but the continuity of BMZ was preserved. Three-dimensional reconstruction of dermal-epidermal junction in LS revealed alterations of the BMZ, most pronounced at the level of the lamina densa and sublamina densa.     

Alterations in the basement membrane zone in pili annulati hair follicles as demonstrated by electron microscopy and immunohistochemistry

BACKGROUND: Pili annulati is a rare autosomal dominant inherited hair shaft abnormality in which clinical examination reveals alternating light and dark bands leading to a shiny appearance of the hair. The clinically light bands are the abnormal areas due to cavities within the cortex. The pathogenesis remains unknown. OBJECTIVES: To investigate the expression of the basement membrane zone (BMZ) components in pili annulati hair follicles of the scalp. METHODS: Transmission electron microscopy (TEM) was carried out on scalp sections of six individuals with pili annulati and six controls. Longitudinal sections of scalp tissues from four individuals with pili annulati and six normal controls were studied by immunohistochemistry with a panel of monoclonal antibodies to the following BMZ components: alpha(6)beta(4) integrin, laminin 5, LH39 antigen, laminin 1, collagen IV and collagen VII. RESULTS: Using TEM, pili annulati scalp specimens exhibited a reduplicated lamina densa in the region of the root bulb in comparison with the single thin electron-dense band in controls. Using immunohistochemistry, there was a wavy BMZ in pili annulati follicles with antibodies to components of the lamina lucida, lamina densa and anchoring fibrils, whereas the BMZ in control hair follicles was as a smooth linear band. The expression of the hemidesmosome-associated alpha(6)beta(4) integrin was linear in both pili annulati and control hair follicles. CONCLUSIONS: Our results suggest that the genetic defect may be a mutation in proteins involved in signalling and regulation of formation and degradation of the lamina densa and sublamina densa region resulting in abnormal assembly or remodelling of the BMZ.     

The localization of target antigens and autoantibodies in linear IgA disease is variable: correlation of immunogold electron microscopy and immunoblotting

Linear IgA disease (LAD) of adults and children is a dapsone-responsive, autoimmune subepidermal blistering disease characterized by linear IgA deposits at the basement membrane zone (BMZ) of the skin and mucosa. Circulating IgA antibodies to BMZ components are often present. In this study we investigated the ultrastructural localization of the antigens and autoantibodies in six patients with LAD (five adults and one child). Using a direct postembedding immunogold electron microscopy (EM) technique, three different patterns of IgA antibody deposition were seen in the skin of four patients with LAD. The IgA deposits localized within the uppermost part of the lamina lucida and to the basal surface of the hemidesmosome in two patients, to the lamina lucida in one, and to the lamina densa in the fourth patient. Using an indirect immunogold EM technique and serum or purified blister fluid from two additional LAD patients, we showed that the serum autoantibodies of one patient bound to the hemidesmosome of the BMZ, while the autoantibodies in the blister fluid of the other patient bound to the lamina densa and sublamina densa including the anchoring fibrils in a labelling pattern similar to that of the monoclonal antibody (LH7.2) to collagen VII. All the autoantibodies binding to the hemidesmosome or lamina lucida recognized a protein in epidermal extracts of molecular weight 180 kDa or its breakdown product of 97 kDa, 200 kDa or 230 kDa. The antibodies binding to the lamina densa recognized proteins of 180 and 285 kDa. The antibodies that bound to the lamina densa and anchoring fibrils recognized collagen VII. In this immunogold EM study we have shown four patterns of IgA labelling in six patients with LAD, associated with five different antigens as recognized by immunoblotting. These results, together with our previous immunofluorescence and immunoblotting findings add support to the contention that LAD is a heterogeneous disease as regards both the target antigens and epitopes.     

The basement membrane zone in lichen sclerosus: an immunohistochemical study

The alteration in expression of basement membrane zone (BMZ) components in lichen sclerosus was investigated by immunohistochemical staining of skin biopsies from seven patients with histologically confirmed disease compared with controls. Monoclonal antibodies and polyclonal sera directed against proteins of the hemidesmosomes, anchoring fibrils, lamina lucida, lamina densa and BMZ collagens were used. Characteristic histological appearances at the dermo-epidermal junction were reflected in widespread alterations in antigen expression in the epidermal basement membrane and the papillary dermis. Expression of the proteins which constitute the structural scaffold (collagen IV and VII) were increased in lichen sclerosus. Expression of hemidesmosomal proteins which mediate adhesion and cell to matrix interaction (α6/β4 and hullous pemphigoid antigen) and expression of anchoring filament components were markedly reduced, suggesting that the epidermal cells are exposed to selective damage.     

Ontogenesis of the basement membrane zone after grafting cultured human epithelium: a morphologic and immunoelectron microscopic study.

Sheets of cultured epithelial cells have been successfully used as autografts for the permanent coverage of patients with full-thickness burns and as allografts to stimulate the healing of chronic skin ulcers. The basement membrane zone (BMZ), composed of lamina lucida, lamina densa, and anchoring fibrils, plays a pivotal role in the firm adherence of the epidermis to the dermis. The present study describes the ultrastructural development during various stages of wound healing after resurfacing different wound areas by cultured epithelial grafts. For this purpose, biopsies were obtained from five patients 5 d to 4.5 years after resurfacing full-thickness burns with cultured autografts, and from five patients at various stages after treatment of excised tattoos and chronic skin ulcers with cultured allografts. One biopsy was taken from a spontaneously healed burn wound 30 years post-injury. Ultrathin sections were prepared for transmission and immunoelectron microscopy, using monoclonal antibodies against type IV and VII collagen. Findings were compared to controls of age- and site-matched normal skin. Eleven days after grafting, the first BMZ features had regenerated, including lamina lucida, a discontinuous lamina densa, hemidesmosomes, and sparse anchoring fibrils. The process of de novo synthesis of BMZ components had begun, and within 4 to 5 weeks complete reformation of BMZ was observed, including normal distribution of anchoring fibrils. Immunolabeling of type VII collagen was first observed upon the lower part of lamina densa at day 11 and steadily increased, reaching normal values 5 weeks after grafting. In contrast, gold deposition of type IV collagen upon lamina densa was strongly increased at day 19 compared to normal. This high expression reduced a little at 5 weeks, but remained high up to 30 years after injury. Long-term burn scars exhibited pseudopodia-like extensions of all basal cells, abundant anchoring fibrils, and an increased amount of arching anchoring fibrils. These features might compensate for the lack of proper rete ridges.     

The effects of epidermal keratinocytes and dermal fibroblasts on the formation of cutaneous basement membrane in three-dimensional culture systems

The cutaneous basement membrane (BM) plays an important role in normal and pathological conditions. However, few studies have addressed the formation of the cutaneous BM using three-dimensional culture systems. In this study, to elucidate the effects of human epidermal keratinocytes and dermal fibroblasts on the formation of the cutaneous BM, keratinocytes were cultured on several dermal substrates in the presence or absence of fibroblasts at the air–liquid interface. After 2 weeks of culture, immunohistochemical stainings for the components of the BM and electron microscopic studies of the BM zone (BMZ) were performed. In cultures of keratinocytes alone on dead reticular dermis or collagen gel without fibroblasts, 4 integrin chain, laminin, type IV and VII collagens were all expressed. However, ultrastructurally, BMZ was not formed. In cultures of keratinocytes on fibroblast-populated collagen matrix, laminin, and type IV and VII collagens were expressed more strongly than in the absence of fibroblasts. In addition, elements of the BMZ such as hemidesmosomes, lamina lucida, lamina densa and anchoring fibrils were formed, although it was still incomplete. In the culture of keratinocytes alone on de-epidermized dermis (DED) (surface up), 4 integrin chain, laminin, and type IV and VII collagens were strongly expressed. Also, the BMZ appeared similar to that in normal skin. In cocultures of keratinocytes and fibroblasts on DED or cultures of keratinocytes on DED combined with fibroblast-populated collagen matrix, type IV collagen was expressed more strongly than in cultures of keratinocytes alone. Ultrastructurally, similar findings to those of cultures of keratinocytes alone on DED were seen. Interestingly, when keratinocytes and fibroblasts were cocultured on DED, some fibroblasts were seen in the upper dermis as a result of migration into the dermis through partial loss of the lamina densa. These results show that keratinocytes produce most of the components of the BM such as laminin, and type IV and VII collagens. In addition, fibroblasts stimulate the expression of the components of the BM and the formation of a BMZ, suggesting that fibroblasts may produce laminin, and type IV and VII collagens or influence the effects of keratinocytes on the formation of the BM through a keratinocyte–fibroblast interaction.This investigation was supported by a grant (04-2001-027) from the Seoul National University Hospital Research Fund and partly by the Pacific Corporation.     

Presence of basal lamina-like substance with anchoring fibrils within the amyloid deposits of primary localized cutaneous amyloidosis

The dermal-epidermal (DE) junction areas of skin specimens obtained from 16 patients with either lichen amyloidosis or macular amyloidosis were studied. In the dermal papillae where amyloid was deposited, elastic fibers frequently were absent, but periodic acid-Schiff reaction after diastase digestion was homogenously positive. Ultrastructural studies revealed that a basal lamina-like substance with anchoring fibrils was present between and within amyloid deposits. By indirect immunofluorescence technique using an anti-basement membrane zone antiserum obtained from a patient with bullous pemphigoid, specific linear fluorescence occurred at the DE junction, and in a reticular pattern in dermal papillae. It seemed that apoptotic keratinocytes of the epidermis brought down basal lamina and fine fibrous components attached to it when these cells dropped down to the papillary dermis and became the source of amyloid. These findings support the hypothesis that epidermal keratinocyte degeneration plays an important role in the histogenesis of cutaneous amyloidoses.     

Heterogeneity of Brunsting-Perry type pemphigoid: a case showing blister formation at the lamina lucida, immune deposition beneath the lamina densa and autoantibodies against the 290-kD polypeptide along the lamina densa

An otherwise healthy 31-year-old man presented with multiple, vesicular, subepidermal blistering on the head, face, chest and oral cavity, leaving shallow scar formation, typical of Brunsting-Perry type pemphigoid. Direct immunofluorescence showed linear deposition of immunoglobulin (Ig)G and C3 along the basement membrane zone (BMZ), and indirect showed anti-BMZ autoantibodies (IgG, >40×) reacting with the dermal side under the salt-split study. Immunofluorescence staining for type IV collagen and laminins, as well as routine electron microscopy, demonstrated that the cleavage level of the blister was intra-lamina lucida. The immunoperoxidase method applied to lesional skin demonstrated IgG deposits along the lamina densa. The post-embedding immunogold method demonstrated that the autoantibodies against BMZ reacted with the lamina densa and the dermis just beneath it. Immunoblot studies demonstrated that the autoantibodies reacted with the 290-kD polypeptide (suggesting type VII collagen) when dermal extract was used as the substrate. The patient was treated with combination therapy consisting of 30 mg prednisolone, 900 mg nicotinamide and 750 mg tetracycline, and the number of newly forming blisters decreased. We concluded that Brunsting-Perry type pemphigoid, a rare autoimmune blistering disease, includes cases showing characteristics of epidermolysis bullosa acquisita as well as bullous pemphigoid. This case showed discrepancy between the blistering level (intra-lamina lucida) and location of antigen (lamina densa and sub-lamina densa areas).     

Bullous amyloidosis

A patient with a 12-year history of a relapsing bullous dermatosis is presented. Unusual clinical features included urticarial erythema, conspicuous mottled hyper- and depigmentation, lichenification and ichthyosiform hyperkeratosis. Serum immunoglobulin E levels were elevated. Histological examination showing deposits of amyloid in the uppermost dermis confirmed the diagnosis of bullous amyloidosis. Ultrastructurally, blister formation occurred at the level of the lamina lucida. The amyloid did not react with a panel of antibodies directed against amyloid fibril proteins. No underlying systemic disease was found. The bullous eruption responded to prednisolone therapy.     

Immunohistochemical studies on fibrillin in amyloidosis, lichen ruber planus and porphyria.

The pathogenesis of macular amyloidosis and lichen amyloidosis remains unsolved and the primary amyloid fibril protein(s) has not yet been identified. Ultrastructural association of skin amyloid with elastin associated microfibrils has been noted earlier. The presence of fibrillin in conjunction with such microfibrils was recently demonstrated immunohistochemically. The presence of fibrillin immunoreactivity in the amyloid deposits in skin biopsies from 3 patients with macular amyloidosis and 3 patients with lichen amyloidosis was studied, using monoclonal anti-fibrillin antibodies. For comparison, skin specimens were studied from five patients with lichen ruber planus, four patients with erythropoietic protoporphyria and from a patient with myeloma-associated cutaneous amyloidosis. Renal specimens from two cases of the amyloid A type of renal amyloidosis also were investigated. There was no immunostaining either of the keratin bodies in specimens of lichen ruber planus, the cutaneous PAS-positive vascular deposits in patients with erythropoietic protoporphyria, or the amyloid deposits in specimens of systemic amyloidosis and it was faint or absent in amyloid deposits in the specimens from patients with lichen amyloidosis. In contrast, distinct fibrillin immunoreactivity could be demonstrated in amyloid deposits in specimens from patients with macular amyloidosis. It was sometimes absent in deposits located in the upper part of the papillary dermis, close to the dermal epidermal junction zone, while consistently strong in deposits located lower down in the dermis. The results suggest that fibrillin or part of the fibrillin molecule may be present in some of the amyloid deposits in specimens of macular amyloidosis.     

Alterations of basement membrane zone and cutaneous microvasculature in morphea and extragenital lichen sclerosus

The aim of this study was to compare alterations of the basement membrane zone (BMZ) and to visualize changes within the skin vascular network in morphea and extragenital lichen sclerosus with the use of laser scanning confocal microscopy. This work was performed in eight plaques of morphea (three active and five inactive) and eight of lichen sclerosus (three of short duration and five long-lasting). Biopsy specimens from six healthy individuals served as controls. The biopsies were cut into 40-microm-thick sections, labeled with antibodies against beta4-intergin (a lamina lucida marker), collagen IV, and the N-terminal end of collagen VII (lamina densa markers) and C-terminal end of collagen VII (a sublamina densa marker) and studied using laser scanning confocal microscopy. Three-dimensional reconstruction of various regions of the BMZ showed a decreased number and size of the dermal papillae both in morphea and lichen sclerosus compared with normal skin. In morphea, the continuity of the BMZ at the level of lamina lucida, lamina densa, and sublamina densa was preserved whereas in LS numerous invaginations and holes were present in the BMZ at the level of the lamina lucida and lamina densa. Thus the alterations of the BMZ in morphea differ from those in lichen sclerosus. Three-dimensional reconstruction of the skin vascular network showed increased angiogenesis only in the early inflammatory stage of morphea, whereas in inactive morphea and lichen sclerosus various numbers of enlarged vessels were visible. The changes in the vascular network in morphea appear to be related to the activity of the disease.     

Nodular amyloidosis

We present a 38-year-old man with nodular amyloidosis and diabetes mellitus. Skin biopsy showed massive amyloid deposition in the dermis. Immunohistochemical study revealed that amyloid deposits contained amyloid of Kappa light-chain origin and P component. Under electron microscope, vascular changes were noted in addition to amyloid deposits. The origin of nodular amyloidosis and its relationship to diabetes mellitus is discussed in this report.     

Restriction of cicatricial pemphigoid antigens to the lamina densa: confirmation by indirect immunoelectron microscopy.

Circulating anti-basement membrane zone (BMZ) antibodies in a patient with cicatricial pemphigoid (CP) were examined using an indirect immunofluorescence test, indirect immunoperoxidase electron microscopy, and Western blot analysis. An indirect immunofluorescence test on salt-split skin revealed that the anti-BMZ antibodies reacted solely to the dermal side at the separating epidermal-dermal interface, and indirect immunoelectron microscopy on intact skin indicated localization of the corresponding antigens (CP antigens) over the lamina densa and within the lower half of the lamina lucida; there were no CP antigens beneath a melanocyte. Indirect immunoelectron microscopy on salt-split skin demonstrated that the CP antigens were partly dissociated from, but restricted to, the lamina densa. Western blot analysis showed no differences in molecular weight between the CP antigens and bullous pemphigoid (BP) antigens. CP antigens, as detected by this patient's serum, appear to be constituted of molecules quite similar to BP antigens, but with different epitopes. CP antigens may be shed from basal cells and locate in the area of anchoring filaments, where they play a role in connecting basal cells to the underlying lamina densa.     

A unique epithelial basement membrane antigen defined by a monoclonal antibody (KF-1)

The basement membrane zone (BMZ) of human skin is a complex structure which contains several well-defined components including bullous pemphigoid antigen, laminin, type IV collagen, and proteoglycan. Characterization of additional basement membrane (BM) constituents has been limited by their relative inaccessibility, insolubility, and low tissue concentration. We have produced a murine monoclonal antibody that has enabled us to define a unique constituent of the BMZ of human stratified squamous epithelia. The monoclonal antibody (KF-1) was raised by standard techniques using suction blister-derived trypsinized human epidermal cells as the antigen. Indirect immunofluorescence and immunoperoxidase staining of human and rhesus monkey tissues with KF-1 produced linear BMZ staining of stratified squamous epithelia. Glandular and vascular BMs were not stained. Immunoelectron microscopic studies of normal human skin and esophagus showed specific binding of KF-1 to the lamina densa of the BMZ, a localization identical to that of type IV collagen. However, unlike type IV collagen, which is not species specific and is found in all BMs, the antigen defined by KF-1 is collagenase-resistant and is specific for primate stratified squamous epithelia. These findings confirm the existence of regional variation in BM composition, and demonstrate for the first time that the lamina densa of stratified squamous epithelial BMs contains a constituent other than type IV collagen.     

Unusual skin manifestation of cutaneous amyloidosis

A 76-year-old man with a 20-year history of extensive cutaneous amyloidosis is reported. He had asymptomatic symmetric brownish reticulated pigmented patches with well-demarcated borders on his thighs, lower legs, dorsal feet and both arms. The trunk and popliteal fossae were not affected. A skin biopsy specimen showed abundant amyloid deposits in the papillary dermis and reticular dermis. Despite the extensive cutaneous involvement and large amount of amyloid in the deep dermis, no evidence of systemic amyloidosis could be found. Various manifestations of cutaneous amyloidosis are reviewed. We report this case to remind dermatologists of the protean presentations of cutaneous amyloidosis.     

Immunofluorescence and histochemical studies of localized cutaneous amyloidosis

Lichen amyloidosus (LA) and macular amyloidosis (MA) are two forms of localized cutaneous amyloidosis in which the amyloid occurs as larger and smaller deposits respectively in the papillary dermis. The histogenesis of the amyloid of these conditions is unknown. By using an indirect immunofluarescence technique we showed that LA and MA do not react with antibodies against different previously characterized amyloid fibril proteins. These results indicate that the amyloid of LA and MA is different from other known types of amyloid. Protein AP, which was demonstrated in amyloid of MA and LA, is known to be present in all forms of amyloid and is of unknown significance. Antiserum against keratin did not react with the larger homogeneous amyloid bodies, but showed a weak reaction with some small deposits. Histochemical staining failed to show keratin in any of the tissues containing LA or MA.