Prenatal diagnosis of genodermatoses: current scope and future capabilities

The genodermatoses encompass a range of inheritable skin diseases that may be associated with significant mortality and long‐term morbidity. In the past, options for prenatal diagnosis of these diseases were limited to fetal skin biopsy. As a result of recent leaps made in genetics and molecular biology, DNA‐based prenatal diagnosis is now available for an increasing number of genodermatoses, and newer non‐invasive methods are being developed that have the potential for tremendous future impact in dermatology. Dermatologists caring for patients with genodermatoses should be aware of the options for screening and prenatal testing and partake in a multi‐disciplinary approach to patient care.     

Preimplantation genetic diagnosis of severe inherited skin diseases

Abstract: Considerable progress has been made recently in elucidating the molecular pathology underlying several forms of inherited skin diseases. One of the most immediate benefits of these discoveries has been the development of DNA-based prenatal diagnosis in pregnancies at risk for recurrence of a particular disorder. In less than 2 decades, prenatal testing has progressed from mid-trimester fetal skin biopsies or protein analysis in a limited number of conditions to first trimester chorionic villus sampling in a much broader range of genodermatoses. Advances in in vitro fertilization protocols and embryo manipulation techology have further led to the feasibility of even earlier prenatal diagnosis through preimplantation genetic diagnosis. This article details some of the recent advances in genetic skin disease research relevant to prenatal diagnosis and explores the possibilities and practicalities of preimplantation genetic diagnosis in the prevention of these conditions.     

Prenatal exclusion of harlequin ichthyosis; potential pitfalls in the timing of the fetal skin biopsy

BACKGROUND: Harlequin ichthyosis (HI) is a severe and usually fatal congenital skin disorder with autosomal recessive inheritance. Several cases of HI prenatal diagnosis have been performed using fetal skin biopsy, mainly at around 23 weeks estimated gestational age (EGA), and reported in the literature. However, prenatal testing must be done earlier than 21 weeks EGA in several countries including Japan where the present HI families live, because termination is legally allowed only until 22 weeks EGA. OBJECTIVES: We report the successful prenatal exclusion of HI in two fetuses from two independent families and discuss the technical difficulties and potential pitfalls in the prenatal exclusion of HI at early gestation stages. METHODS: Fetal skin biopsy specimens and amniotic fluid samples at 19 and 20 weeks EGA from two fetuses at risk of HI were examined by light and electron microscopy. RESULTS: For the prenatal diagnosis in case 1, the fetal skin biopsy samples were obtained at 20 weeks EGA and showed normal keratinization in the hair canals; no abnormalities were observed in the keratinized cells. In case 2, the interfollicular epidermis and the hair follicles in the samples obtained at 19 weeks EGA had not differentiated enough to show proper keratinization. However, lamellar granules were normally formed in the inner root sheath cells of the late bulbous hair pegs. From these ultrastructural findings, the case 1 fetus was diagnosed as unaffected with HI, and the case 2 fetus was diagnosed as unlikely to be affected. Subsequently, both were born as healthy, unaffected babies. CONCLUSIONS: The timing of biopsies at 19 weeks EGA is not ideal for fetal skin biopsy because the samples are not always sufficiently differentiated for the prenatal diagnosis of HI. However, morphological observations of lamellar granules gives us important additional information useful for HI prenatal diagnosis.     

Prenatal diagnosis and screening

We have limited the scope of this article to those disorders that have already been successfully diagnosed or excluded in utero. We currently have the potential to diagnose a number of others for which the opportunity has not yet arisen. If a biochemical, morphologic, chromosomal, or DNA alteration is known for a specific condition and is likely to be expressed in one of the fetal tissues or secretions, attempt at prenatal diagnosis is reasonable. Our ability to detect the inherited disorders of the skin in utero will continue to improve both in the number of specific disorders successfully diagnosed or excluded and in the increasingly earlier stages of pregnancy at which the disorder can be detected. Advances in instrumentation will, it is hoped, decrease the risk of the invasive methods of prenatal diagnosis, and improvement in noninvasive methods, such as maternal serum screening, may eliminate the need for invasive procedures altogether. Detection of useful DNA polymorphisms linked to genes for specific genodermatoses and development of specific gene probes will improve the accuracy of diagnosis and reduce the need for specific fetal tissues. The entire genome of an individual is present in each cell, even though a specific gene product may not be expressed in that cell. Thus, DNA restriction endonuclease studies can be performed on amniotic fluid cells, chorionic villi, fetal cells in maternal circulation, and fetal tissues with equal facility. The usefulness of prenatal diagnosis will always be limited by the ability to detect pregnancies at risk. If carrier detection is unavailable, the only way to identify couples at risk for offspring with an autosomal recessive condition is by the birth of an affected child. For autosomal dominant and X-linked recessive and dominant conditions, new mutations will continue to occur. As mentioned previously, screening of all pregnancies for all defects is not possible now and is unlikely ever to be feasible, either economically or technically. The reliability of prenatal diagnosis will continue to depend upon accurate diagnosis in the index case and upon the availability of a specific and sensitive test (or tests), with no overlap in values between heterozygotes and homozygotes for autosomal recessive conditions or between normal and affected fetuses with autosomal dominant and X-linked recessive disorders. Correct interpretation of test results is subject to experience, recognition of artifact, and variation in the expression of a given disorder in utero.(ABSTRACT TRUNCATED AT 400 WORDS)     

Translational benefits from research on rare genodermatoses

Significant new discoveries about many of the genodermatoses have been made recently through an improved knowledge of the human genome, advances in molecular screening strategies and also through more comprehensive Internet DNA databases. By 2003, over 350 single gene skin disorders had been characterized at a molecular level. These new data provide more detailed information for patients, allow for more accurate diagnoses, and help improve genetic counselling. Other benefits include the feasibility of carrier screening and DNA-based prenatal testing, as well as a platform for devising new treatments, including somatic gene therapy. Research on rare single gene disorders also provides new insight into more common skin conditions. For example, new ideas about photosensitivity are emerging from discoveries of mutations in a novel component of the actin cytoskeleton (kindlin-1) in the rare inherited poikiloderma disorder, Kindler syndrome. Likewise, new clues to understanding disease pathology in lichen sclerosus have been gleaned from the discovery of pathogenic mutations in the skin protein, extracellular matrix protein 1, in the rare sclerosing inherited skin disorder, lipoid proteinosis. Finally, new insight into what can cause exuberant granulation tissue in chronic wounds has been provided by the discovery of specific mutations in the basement membrane protein, laminin 5, in the rare inherited condition, laryngo-onychocutaneous syndrome. It is clear that a precise research focus on the rare genodermatoses is providing practical benefits for sufferers of these disorders, as well as new lessons and ideas about more common acquired skin conditions.     

Antenatal diagnosis of hereditary epidermolysis bullosa

Among the genetic disorders of the skin, the heterogeneous group of epidermolysis bullosa includes some of the most severe. Prenatal diagnosis is of considerable importance to the families who have had an affected child, or in which one of the parents is affected. The prenatal diagnosis is performed using fetoscopy and fetal skin biopsy. Fetal skin samples are taken at 20 weeks of gestation and are examined by light and electron microscopy to determine whether the fetus is affected. We report here the French experience on prenatal diagnosis of the severe inherited epidermolysis bullosae. Given the severity and frequency of Herlitz syndrome, it is not surprising that this is the most frequently encountered disease in our series of prenatal diagnosis (14 of 21 epidermolysis bullosae), followed by Hallopeau-Siemens (6 cases), and Pasini type (1 case). Our exclusion diagnosis of a Pasini fetus was the first prenatal diagnosis of this type of epidermolysis bullosa performed and reported in the literature. We stress here in this paper that observing the site of separation in the epidermal dermal junction is not sufficient to make a positive prenatal diagnosis. Prenatal diagnosis depends on the observation of the specific ultrastructural marker of the disease such as: hypoplasia and absence of hemidesmosomes and sub-basal dense plate in junctional epidermolysis bullosa-Herlitz, collagenolysis in recessive dystrophic epidermolysis bullosa-Hallopeau-Siemens, and absence and hypoplasia of anchoring fibrils in dominant dystrophic epidermolysis bullosa-Pasini. Until biochemical defects are clarified and suitable tests become available, electron microscopy remains the only current means for reliable, genetically useful, diagnosis of epidermolysis bullosa. In 62 per cent of cases of our series a prenatal diagnosis of exclusion of disease was made and we would stress that in high risk families repeated fetoscopies for prenatal diagnosis are possible in consecutive pregnancies thus allowing the family to have only normal children.     

Progress in epidermolysis bullosa: from eponyms to molecular genetic classification

Epidermolysis bullosa, a clinically and genetically diverse group of heritable mechanobullous disorders characterized by skin fragility in the cutaneous basement membrane zone, has become a prototype for the recent progress in molecular genetics of genodermatoses. The different forms of epidermolysis bullosa have been linked to mutations in no less than 10 distinct genes encoding the major structural basement membrane zone proteins. This information has formed a basis for refined molecular classification with prognostic implications, improved genetic counseling, and prenatal and preimplantation genetic diagnosis.     

The molecular genetics of the genodermatoses: progress to date and future directions

The Human Genome Mapping Project and allied rapid advances in genetic technology over the past decade have facilitated accurate association of allelic variations in several genes with specific skin phenotypes. Currently the genetic bases of the majority of the more common genodermatoses have been elucidated. In scientific terms this work has been extraordinarily successful and has yielded many new biological insights. These advances, although exciting, have yet to be translated into direct benefit for patients with these diseases. Genetic counselling has been greatly aided by gene identification, by the better understanding of genotype-phenotype correlation and by the disclosure of unexpected genetic mechanisms in some families. Knowledge of the molecular basis of these disorders has also been vital in enabling DNA-based prenatal diagnosis in several conditions and DNA-based preimplantation diagnosis has been used in a selected few. While this successful period of gene mapping is now nearing completion, progress towards the next goal, that of developing therapeutic strategies based on the knowledge of these underlying genetic mechanisms, has proven frustratingly slow. Despite the ready access to the skin compared with solid internal organs, the challenges of cutaneous gene therapy are legion and many technical issues need to be surmounted to enable gene replacement or modification of gene expression to have a useful role in these disorders. In this article we make a comprehensive review of progress to date in gene identification, genotype-phenotype correlation, prenatal diagnosis and cutaneous gene therapy, and we examine future directions for research in this field.     

Prenatal diagnosis for severe inherited skin disorders: 25 years' experience

BACKGROUND: Over the last 25 years there have been major advances in methods for prenatal testing of inherited skin disorders. Since 1979, our group at the St John's Institute of Dermatology has performed 269 prenatal diagnoses, using a variety of approaches, including fetal skin biopsy (FSB), chorionic villus sampling (CVS) and preimplantation genetic diagnosis (PGD). OBJECTIVES: This study was designed to review the clinical indications, testing procedures and laboratory analyses for all prenatal tests conducted at St John's over this period. METHODS: FSBs were examined for morphological and, when relevant or feasible, immunohistochemical abnormalities. The DNA-based tests involved screening by nucleotide sequencing, restriction enzyme digests or, in a few cases, by linkage analysis. Results Of the 269 tests, 191 were FSB, 76 were CVS and two were PGD. The major indications for FSB were epidermolysis bullosa (EB) (138 cases, including 88 junctional and 48 dystrophic), ichthyoses (37 cases, including 22 tests for harlequin ichthyosis) and oculocutaneous albinism (12 cases). Of the CVS procedures, 75 were for EB (40 junctional, 35 dystrophic) and one was for the EEC (ectrodactyly, ectodermal dysplasia, clefting) syndrome. Both of the PGD procedures were for the skin fragility-ectodermal dysplasia syndrome. All tests provided accurate diagnoses and the fetal loss rate was approximately 1% for both FSB and CVS. CONCLUSIONS: The development of prenatal testing has proved to be of great benefit for individuals or couples at risk of having children with severe inherited skin disorders and, in the absence of a cure, prenatal testing along with appropriate counselling has become an important translational benefit of basic research and an integral part of clinical management.     

Nucleic acid‐based non‐invasive prenatal diagnosis of genetic skin diseases: are we ready?

The discovery of circulating fetal nucleic acids is a great step on the way of developing non‐invasive prenatal diagnosis (NIPD) for genetic disorders. Here, we briefly discuss the current applications of circulating fetal nucleic acids in genetic testing for different kinds of hereditary diseases with an emphasis on using circulating cell‐free fetal DNA in diagnosis of monogenic disorders. As the genetic skin disorders impair the quality of life at different levels, we next discuss some ethical issues in NIPD for genetic skin diseases of various severities and in particular, the responsibility of doctors and parents, respectively, in the prenatal genetic testing.     

Infancy- and childhood-onset dyschromatoses

The dyschromatoses are a group of pigmentary disorders characterized clinically by mixed and often guttate hypopigmented and hyperpigmented lesions. There are many conditions that present with dyschromatosis, including genodermatoses, inflammatory skin diseases, infections, drug and chemical use, and nutritional disorders. Some conditions have extracutaneous features. Poikiloderma (a combination of hypo- and hyperpigmentation with telangiectasia and atrophy) must be excluded. In this article, we describe the dyschromatoses typically presenting in infancy and childhood, most of which are genodermatoses. The approach we have taken in classifying them is based on organ involvement. We hope this article will serve as a guide for dermatologists to the recognition of these uncommon conditions.     

Prenatal diagnosis as a test for genodermatoses: its past,present and future

The prenatal diagnosis (PND) of severe hereditary skin diseases started in the early 1980s using fetal skin biopsy techniques based on ultrastructural and immunohistochemical abnormalities of the fetal skin. Recent success in identifying responsible genes and demonstrating mutations in such genes has set the stage for DNA-based PND in the 1990s. Common examples of skin conditions which can be prenatally diagnosed include epidermolysis bullosa, oculocutaneous albinism and Harlequin ichthyosis in which the severity of the clinical phenotype appears to justify PND in families at risk. More recently, preimplantation diagnoses of inherited diseases have become possible using in vitro fertilization techniques. The diagnosis consists of a blastomere biopsy of the six to ten-cell embryo and a DNA analysis of single blastomeres. Disease-free embryos are selected for transfer to the uterus, thereby avoiding the need for termination of a fetus found to be affected by conventional PND. Furthermore, carrying out a PND using a single fetal cell from the maternal blood, such as nucleated erythrocytes, has become technically feasible. Although there are many questions that remain unanswered, the outlook for further development of noninvasive PND in the future appears optimistic.     

Praktische Aspekte der molekularen Diagnostik bei Genodermatosen

Genodermatoses are rare genetic disorders with a broad spectrum of cutaneous and extracutaneous manifestations that have a genetic background. A thorough clinical examination, laboratory workup and morphological analyses of the skin remain crucial for the diagnosis in the era of next generation sequencing (NGS). The diagnostic algorithm depends on the clinical and molecular heterogeneity and should be adapted for each group of genodermatoses. In cases with uncharacteristic phenotypes which cannot be classified, NGS-based testing accelerates the time to diagnosis and leads to the identification of new disorders and new disease-associated genes. The new knowledge on genotype–phenotype correlations should enable revision of the classification of genodermatoses on a molecular basis.     

Facial involvement in genodermatoses

Facial involvement represents a characteristic feature of a wide range of genodermatoses. Specific facial findings often help point to the correct diagnosis, which improves counseling and management. In particular, this can facilitate the identification and treatment of associated extracutaneous disease. The highly visible nature of facial lesions in genodermatoses and facial birthmarks can result in stigmatization and frequently leads to particular concern in patients and their family members. It is therefore critical for dermatologists to be aware of the broad spectrum of facial manifestations in genetic skin disease, especially when these findings have important implications with regard to monitoring and treatment. In this contribution, facial involvement in genodermatoses is divided into five morphologic categories based on the most prominent feature: Papules, scaling, photosensitivity/findings associated with aging (eg, telangiectasias, atrophy, lentigines), blisters/erosions, and birthmarks. Hopefully, this will provide a practical and clinically useful approach to a large and diverse assortment of genetic skin conditions.     

Prenatal diagnosis of genetic skin disease by fetoscopy and electron microscopy: report on 5 years' experience

Since our first report 5 years ago (Rodeck, Eady & Gosden, 1980) the combination of fetoscopy and electron microscopy to obtain and examine fetal skin biopsy specimens has become an established method for assessing mid-trimester fetuses at risk for different genodermatoses characterized by specific ultrastructural abnormalities. Our present report concerns 39 fetuses ( Table ).  

  TABLE. Skin biopsy examination of 39 fetuses     

16.

Prenatal diagnosis of lethal junctional epidermolysis bullosa by fetal skin biopsy   总被引：1，自引：0，他引：1  

H Shimizu  O M Schofield  R A Eady 《Nippon Hifuka Gakkai zasshi. The Japanese journal of dermatology》1991,101(5):539-545

A 22-year-old woman, whose first infant had died of lethal junctional epidermolysis bullosa (JEB), requested prenatal diagnosis for her third pregnancy. At 20 weeks gestation, fetal biopsy was performed under direct vision by fetoscopy. A semithin section of epon-embedded skin showed dermo-epidermal separation at the light microscopic level. Electron microscopy revealed the site of separation to be within the lamina lucida of the epidermal basement membrane (EBM). Indirect immunofluorescence on a 5 microns cryostat specimen of skin showed a complete absence of GB3 monoclonal antibody immunostaining at the EBM compared with a control 18 week old normal fetal skin sample. The diagnosis was therefore made that the fetus was affected with lethal JEB and a prostaglandin termination performed. The diagnosis was confirmed by further studies on the aborted fetus. 54 cases of prenatal diagnosis of various types of epidermolysis bullosa performed at Institute of Dermatology over the last 10 years are briefly reviewed. Several social and practical problems to launch prenatal diagnosis in Japan are also discussed.     

17.

A multistep approach to the diagnosis of rare genodermatoses     

《Clinics in Dermatology》2020,38(4):399-407

Genodermatoses are heritable skin diseases that can cause significant morbidity and mortality. Most of them show characteristic cutaneous findings. Genodermatoses can be associated with extracutaneous system abnormalities. Diagnosing hereditary skin disorders is still a challenging task due to their rarity and diversity, due to diseases evolving over many years, and the initial manifestations not always being diagnostic; therefore, ongoing evaluation and surveillance is often required to make the accurate diagnosis. The algorithm for the diagnosis depends on a combination of thorough clinical and family history clinical examination, laboratory findings, consultation of multiple medical specialists, and molecular analysis. Diagnostic testing targeted at differentiation of similar genodermatoses may be required. Recognition is crucial for the initiation of the treatment for skin manifestations and detection of other extracutaneous abnormalities, including malignancy. Diagnostic accuracy and molecular diagnosis may help in providing a template for ongoing management, testing, and education and prognostication for families of children with genodermatoses.     

18.

Nails: diagnostic clue to genodermatoses     

Inamadar AC  Palit A 《Indian journal of dermatology, venereology and leprology》2012,78(3):271-278

Nails are cutaneous appendages mostly involved in mechanical functions. However, nails may reflect presence of various systemic disorders evidenced by alteration of their shape, size, color or texture. Genodermatoses are multisystem disorders with cutaneous involvement. Many of the genodermatoses present with nail changes and some of these may be the clinical pointers to the diagnosis. Diagnostic clues to various genodermatoses derived from nail findings have been discussed.     

19.

Epidermolytic hyperkeratosis: ultrastructure and biochemistry of skin and amniotic fluid cells from two affected fetuses and a newborn infant   总被引：5，自引：0，他引：5  

K A Holbrook  B A Dale  V P Sybert  R W Sagebiel 《The Journal of investigative dermatology》1983,80(4):222-227

Skin biopsy samples and amniotic fluid cells obtained in utero from two fetuses at risk for epidermolytic hyperkeratosis were examined by light and electron microscopy. Both fetuses were affected; the second was carried to term. Epidermal extracts were prepared from blisters of the newborn for analysis of keratin and filaggrin proteins. Abnormal clumps of keratin filaments were present in all layers of the prekeratinized fetal epidermis except the periderm and stratum germinativum. A significant population of amniotic fluid cells also contained the filament aggregations. Prenatal diagnosis of the disease should be possible using cells obtained at amniocentesis, thus avoiding fetal skin biopsy. Biochemical studies showed abnormalities in keratin and filaggrin proteins. The structural alterations in the tissue might be a consequence of altered interaction between these two abnormal epidermal proteins.     

20.

Rapid prenatal diagnosis of epidermolysis bullosa letalis using GB3 monoclonal antibody   总被引：1，自引：0，他引：1  

A.H.M. HEAGERTY  R.A.J. EADY  A.R. KENNEDY  K.H. NICOLALADES  C.H. RODECK  B-L. HSI  J-P. ORTONNE 《The British journal of dermatology》1987,117(3):271-275

The prenatal diagnosis of epidermolysis bullosa letalis was made by demonstrating a marked reduction of normal immunofluorescence staining with the monoclonal antibody GB3 in a fetal skin biopsy obtained at 18 weeks' gestation. The diagnosis was confirmed by conventional electron microscopy using established techniques. The affected pregnancy continued to term and a baby was delivered who rapidly developed blistering affecting the buttocks, lower limbs and mouth. This technique is simpler and quicker than electron microscopy, yet appears to retain the same degree of accuracy.     

Prenatal diagnosis of lethal junctional epidermolysis bullosa by fetal skin biopsy

A 22-year-old woman, whose first infant had died of lethal junctional epidermolysis bullosa (JEB), requested prenatal diagnosis for her third pregnancy. At 20 weeks gestation, fetal biopsy was performed under direct vision by fetoscopy. A semithin section of epon-embedded skin showed dermo-epidermal separation at the light microscopic level. Electron microscopy revealed the site of separation to be within the lamina lucida of the epidermal basement membrane (EBM). Indirect immunofluorescence on a 5 microns cryostat specimen of skin showed a complete absence of GB3 monoclonal antibody immunostaining at the EBM compared with a control 18 week old normal fetal skin sample. The diagnosis was therefore made that the fetus was affected with lethal JEB and a prostaglandin termination performed. The diagnosis was confirmed by further studies on the aborted fetus. 54 cases of prenatal diagnosis of various types of epidermolysis bullosa performed at Institute of Dermatology over the last 10 years are briefly reviewed. Several social and practical problems to launch prenatal diagnosis in Japan are also discussed.     

A multistep approach to the diagnosis of rare genodermatoses

Genodermatoses are heritable skin diseases that can cause significant morbidity and mortality. Most of them show characteristic cutaneous findings. Genodermatoses can be associated with extracutaneous system abnormalities. Diagnosing hereditary skin disorders is still a challenging task due to their rarity and diversity, due to diseases evolving over many years, and the initial manifestations not always being diagnostic; therefore, ongoing evaluation and surveillance is often required to make the accurate diagnosis. The algorithm for the diagnosis depends on a combination of thorough clinical and family history clinical examination, laboratory findings, consultation of multiple medical specialists, and molecular analysis. Diagnostic testing targeted at differentiation of similar genodermatoses may be required. Recognition is crucial for the initiation of the treatment for skin manifestations and detection of other extracutaneous abnormalities, including malignancy. Diagnostic accuracy and molecular diagnosis may help in providing a template for ongoing management, testing, and education and prognostication for families of children with genodermatoses.     

Nails: diagnostic clue to genodermatoses

Nails are cutaneous appendages mostly involved in mechanical functions. However, nails may reflect presence of various systemic disorders evidenced by alteration of their shape, size, color or texture. Genodermatoses are multisystem disorders with cutaneous involvement. Many of the genodermatoses present with nail changes and some of these may be the clinical pointers to the diagnosis. Diagnostic clues to various genodermatoses derived from nail findings have been discussed.     

Epidermolytic hyperkeratosis: ultrastructure and biochemistry of skin and amniotic fluid cells from two affected fetuses and a newborn infant

Skin biopsy samples and amniotic fluid cells obtained in utero from two fetuses at risk for epidermolytic hyperkeratosis were examined by light and electron microscopy. Both fetuses were affected; the second was carried to term. Epidermal extracts were prepared from blisters of the newborn for analysis of keratin and filaggrin proteins. Abnormal clumps of keratin filaments were present in all layers of the prekeratinized fetal epidermis except the periderm and stratum germinativum. A significant population of amniotic fluid cells also contained the filament aggregations. Prenatal diagnosis of the disease should be possible using cells obtained at amniocentesis, thus avoiding fetal skin biopsy. Biochemical studies showed abnormalities in keratin and filaggrin proteins. The structural alterations in the tissue might be a consequence of altered interaction between these two abnormal epidermal proteins.     

Rapid prenatal diagnosis of epidermolysis bullosa letalis using GB3 monoclonal antibody

The prenatal diagnosis of epidermolysis bullosa letalis was made by demonstrating a marked reduction of normal immunofluorescence staining with the monoclonal antibody GB3 in a fetal skin biopsy obtained at 18 weeks' gestation. The diagnosis was confirmed by conventional electron microscopy using established techniques. The affected pregnancy continued to term and a baby was delivered who rapidly developed blistering affecting the buttocks, lower limbs and mouth. This technique is simpler and quicker than electron microscopy, yet appears to retain the same degree of accuracy.