Positive Direct Immunofluorescence Research Articles

Identification of a Basement Membrane Zone Antigen Reactive with Circulating IgA Antibody in Ocular Cicatricial Pemphigoid

Ocular cicatricial pemphigoid is a rare vesiculobullous disease characterized by linear deposition of IgG and/or IgA along the basement membrane zone of conjunctival biopsies. This study identifies a tissue antigen detected by ocular cicatricial pemphigoid patient sera. Patient selection was based on the presence of only ocular involvement and a positive direct immunofluorescence of conjunctiva. We evaluated patient and control sera using indirect immunofluorescence of basement membrane zone separated skin, Western blot, and purified antibodies from nitrocellulose and epidermal sheets. Direct immunofluorescence performed on the patients' conjunctival biopsy showed linear deposition of IgA along the basement membrane zone in all seven patients, and five of seven also demonstrated deposition of IgG along the basement membrane zone. Indirect immunofluorescence performed on the patients' sera demonstrated linear deposition of IgA along the epidermal side of the basement membrane zone of ethylenediaminetetraacetic acid-separated skin in all seven patients. IgA titers ranged from 1:20 to 1:80. No IgG was detected. Immunoblots detected IgA binding to a 45-kD antigen in all patients as well as sporadic IgA binding to a number of other proteins. Immunoblots stained with sera from patients did not show reactivity to the 230- or 180-kD bullous pemphigoid antigens or the 97-kD linear IgA bullous dermatosis antigen. Eluting IgA from the 45-kD region and other regions revealed that only antibodies eluted from the 45-kilodalton region bound linearly to the basement membrane on separated skin. Purification of IgA using epidermal sheets confirmed that the antibody responsible for staining on indirect immunofluorescence bound to the 45-kD region on Western blot. Sera from normals and patients with bullous pemphigoid, dermatitis herpetiformis, and linear IgA bullous dermatosis failed to demonstrate basement membrane zone IgA on elution of the 45-kD region. We conclude that these ocular cicatricial pemphigoid sera contain a unique IgA antibody that binds to a 45-kD basement membrane zone antigen.

Immunologic Abnormalities Associated With Primary Anetoderma

Primary anetoderma is a rare cutaneous elastolytic disorder, the etiopathogenesis of which has not yet been established. Six patients with primary anetoderma were studied in an attempt to assess the role of the immunologic system in the elastolytic process. The investigation included the medical history, physical examination, routine blood tests, specific tests for collagen diseases, prothrombin time, activated partial thromboplastin time, thyroxine, indirect immunofluorescence test, and skin biopsies for histopathologic study and direct immunofluorescence. Two of the patients presented with autoimmune disorders: the first had Graves' disease, lupus anticoagulant, and autoimmune hemolysis, and the second had systemic scleroderma. There were positive direct immunofluorescence findings in most of the patients. Furthermore, all of them were found to have serologic immunologic abnormalities, of which the most common was a positive antinuclear factor. These findings indicate that there is an immunologic involvement in primary anetoderma.

Immunologic abnormalities associated with primary anetoderma

<h3>• Background and Design.—</h3> Primary anetoderma is a rare cutaneous elastolytic disorder, the etiopathogenesis of which has not yet been established. Six patients with primary anetoderma were studied in an attempt to assess the role of the immunologic system in the elastolytic process. The investigation included the medical history, physical examination, routine blood tests, specific tests for collagen diseases, prothrombin time, activated partial thromboplastin time, thyroxine, indirect immunofluorescence test, and skin biopsies for histopathologic study and direct immunofluorescence. <h3>Results.—</h3> Two of the patients presented with autoimmune disorders: the first had Graves' disease, lupus anticoagulant, and autoimmune hemolysis, and the second had systemic scleroderma. There were positive direct immunofluorescence findings in most of the patients. Furthermore, all of them were found to have serologic immunologic abnormalities, of which the most common was a positive antinuclear factor. <h3>Conclusions.—</h3> These findings indicate that there is an immunologic involvement in primary anetoderma. (<i>Arch Dermatol.</i>1992;128:799-803)

Determination of the optimum site for diagnostic biopsy for direct immunofluorescence in bullous pemphigoid

The distributions in deposition of immunoreactants in bullous pemphigoid before and after initiating treatment were investigated. Punch biopsies of skin were performed on each patient from up to five different sites and studied by direct immunofluorescence (DIF). Both groups showed the highest diagnostic yield for DIF from perilesional biopsies, with positivity of 78% from the pretreatment group and 83% from the post-treatment group. The percentage of positive DIF for remaining sites in the pre-treatment group were as follows: 50% lower back, 62% oral mucosa, 70% flexor aspect of forearm, 70% anterior aspect of thigh. The post-treatment group had positive DIF of other sites biopsied as follows: 59% anterior aspect of thigh, 64% flexor aspect of forearm, 68% back. We conclude that a single perilesional biopsy is usually sufficient to provide positive DIF and more than two biopsies is seldom justified as it is unlikely to increase the yield of positive DIF. If it is not possible to obtain a perilesional biopsy, then the anterior aspect of thigh or flexor aspect of forearm is a suitable alternative site. If a second biopsy is considered after initiating treatment, it should be taken from the oral mucosa as this has been shown in a separate study to have a higher rate of positive DIF than uninvolved skin.

Conjunctival involvement in pemphigus vulgaris: a clinical, histopathological and immunofluorescence study

Eleven patients with pemphigus vulgaris and with eye complaints had histopathological and direct immunofluorescence (DIF) studies on biopsies of their conjunctivae. In eight of the patients the ocular symptoms preceded the other manifestations of pemphigus. None of the 11 patients had any detectable conjunctival blisters or erosions. The conjunctivitis in three of the patients proved on histopathology and DIF to be a manifestation of pemphigus vulgaris. In the remaining patients, conjunctival hyperaemia with or without a mucoid discharge was observed, but there were no specific histopathological features, although one of them had a positive DIF.

Conjunctival involvement in cicatricial and bullous pemphigoid: a clinical and immunopathological study.

Patients with bullous pemphigoid were found to have significant ocular abnormalities. In a group of 18 patients one had conjunctival shrinkage, and 11 of 15 (73%) had positive linear direct immunofluorescence on conjunctival biopsy from a clinically uninvolved site. Our ocular findings in a group of 14 with cicatricial pemphigoid are also reported and compared with those from a control group of 20. Our findings suggest there is overlap between the pemphigoid groups and raise further questions about the pathogenicity of immunoreactants within the basement membrane zone. Bulbar conjunctival biopsy was simple and well tolerated, and the rate of immunofluorescence positivity of conjunctiva was twice that of skin in both pemphigoid groups.

Reactivity of circulating antineuronal antibodies (CANA) on peripheral nervous system structures.

The appearance of circulating antineuronal antibodies (CANA) in patients with malignant tumors has been correlated with the occurrence of paraneoplastic neurological syndromes. However, the effect of CANA on the peripheral nervous system is poorly understood. The reactivity of sera from CANA-positive and -negative patients were investigated on cryostat sections of peripheral nerves and skeletal muscle, and on nerve tease-fiber preparations. Only CANA-positive sera showed staining of Schwann cell nuclei on cryostat sections, whereas nerve tease-fiber preparations and sections of skeletal muscle remained negative. Positive direct immunofluorescence of small cell lung cancer (SCLS) cells was confined to CANA-positive patients only. These findings suggest the existence of a common antigen between SCLC and Schwann cells.

Dermatitis herpetiformis: Oral lesions as an early manifestation

Dermatitis herpetiformis is an uncommon chronic skin disease that rarely affects the oral cavity. An unusual case of dermatitis herpetiformis, starting with oral lesions 6 months prior to the onset of skin lesions, is reported. The diagnosis was based on the typical histopathologic features and on positive direct immunofluorescence. The literature is briefly reviewed. Evaluation of laboratory findings, clinical picture, and response to sulfones is essential to the final diagnosis and is emphasized.

Chronic idiopathic and secondary neutropenia: clinical and serological investigations.

Clinical data on 49 patients with chronic idiopathic neutropenia (CIN) and 42 patients with neutropenia secondary to a well-defined immunological disorder (SN) were collected and related to serological parameters. In 47% of the patients with CIN and 53% of those with SN, a positive direct immunofluorescence test was obtained with granulocytes from the patients. In the sera from the patients in the two groups, antibodies against donor granulocytes were detected by the indirect immunofluorescence test, the leucoagglutination test and/or the granulocytotoxicity test in 15%, 19% and 15%, respectively. The results of the above tests could not be correlated with any clinical or haematological parameter. Immune complexes in the serum were detected by the 125I-Clq-binding test in 29% of patients with CIN and in 58% of those with SN. The presence of serum immune complexes correlated well with the existence of a low neutrophil count, but not with the presence of recurrent infections, with bone-marrow abnormalities, or with positive reactions in other serological tests. The sera of eight out of 14 patients with CIN and seven out of 12 patients with SN had inhibitory activity for myeloid colony formation in vitro (CFU-GM). This CFU-GM inhibitory activity was correlated with the presence of recurrent infections and with hypoplasia of the myeloid compartment of the bone marrow, but not with positive reactions in other tests. We conclude that the 125I-Clq-binding test probably detects circulating immune complexes that induce a shift neutropenia, whereas serum activity inhibitory for CFU-GM possibly relates to clinically more serious forms of neutropenia. The significance of neutrophil-bound Ig and granulocyte-reactive antibodies in the serum is not clear.

Direct immunofluorescence for the study of cutaneous drug eruptions

Forty-one patients with drug eruptions were retrospectively evaluated to correlate clinical, direct immunofluorescence, and dermatopathologic findings. Forty-six biopsy specimens were obtained for immunofluorescence in addition to specimens for dermatopathologic study. Eighteen patients had blood vessel fluorescence. Fourteen patients had basement membrane zone fluorescence. Eleven patients had dermatopathologic evidence of lichenoid changes; 8 of these had basement membrane zone fluorescence. Triamterene-hydrochlorothiazide caused the most eruptions (six patients). Positive direct immunofluorescence findings may aid the diagnosis of drug eruptions, especially in the presence of a nonspecific clinical picture or histopathologic findings.

Polymorphous Light Eruption

Of 138 patients with polymorphous light eruption originally investigated in 1972, one hundred fourteen were contacted seven years later (1979), and 47 subjected to a clinical and laboratory reevaluation. Sixty-four of the 114 patients (57%) described a diminution of sun-related skin complaints, including 12 patients who reported a total freedom from sun sensitivity during the previous two years. In contrast, two patients who in 1972 had claimed temporary freedom from sun sensitivity had again noted their reactions to sunlight by 1979. In five of 43 cases examined at uninvolved skin sites, weak (1+) positive direct immunofluorescence staining for basement membrane zone immunoglobulins was reported by both laboratories employed in this study. Low or moderate antinuclear antibody serum titers were found in nine patients. No patient, however, met diagnostic criteria for systemic lupus erythematosus.

Diagnosis of autoimmune skin disease in the dog: Correlation between histopathologic, direct immunofluorescent and clinical findings

Histopathologic, immunopathologic and clinical evaluation of two hundred thirty dogs resulted in a diagnosis of canine autoimmune skin disease (AISD) in eighty four of the suspected cases. Pemphigus foliaceus was the most common diagnosis, followed by bullous pemphigoid and pemphigus vulgaris. The highest percentage of cases were mixed breeds, followed by shelties, collies, German shepherds and poodles. There was no apparent age or sex predilection. Eighty seven percent of the eighty four cases of AISD were diagnosed on the basis of supportive clinical criteria in addition to compatible histopathologic and/or direct immunofluorescent findings. Thirteen percent of the cases with a final clinical diagnosis of AISD lacked supportive immunohistopathologic criteria on biopsy and the diagnosis was based upon clinical appearance, exclusion of other dermatoses, and response to immunosuppressive therapy. This latter group included four cases of SLE diagnosed using a combination of clinical criteria and a positive test for antinuclear antibody. Thirty three percent of the AISD cases were diagnosed on the basis of compatible findings on both histopathology and direct immunofluorescence. A histopathologic diagnosis of AISD was made in sixty nine percent of the cases. Positive direct immunofluorescence was found in fifty two percent of the cases. Thirty five percent of the AISD cases were diagnosed on histopathologic criteria without supportive immunofluorescent findings, while nineteen percent had positive direct immunofluorescence with nonspecific histopathologic changes. The number of definitive positive results was greater for direct immunofluorescence (38%) than for histopathology (25%). There was an equal incidence (21%) of false positive results among non-AISD cases with both tests. Indirect immunofluorescence appears to have little value in the diagnosis of canine AISD. The recommended diagnostic approach to canine AISD involves the use of both histopathologic and immunopathologic evaluations.

The duration and quality of positive direct immunofluorescence in skin biopsies using michel's fixative on a case of equine pemphigus foliaceus

A special buffered ammonium sulfate fixative (Michel's fixative) designed for use as a liquid media capable of preserving in vivo tissue fixed immunoglobulins and complement was evaluated. The preservative ability of this fixative was studied by using skin biopsies from a confirmed case of spontaneously occurring equine pemphigus foliaceus. Once obtained, 16 skin biopsies were placed in Michel's fixative. Eight samples were stored at ambient temperature (21°C) and 8 samples were stored in a conventional refrigerator at 4°C. Over an 8 month time period, direct immunofluorescence was performed on these skin biopsies looking for the abnormal presence of IgG, IgM (when done) and C 3 deposition within the intercellular spaces of the epidermis. The results of this study indicated that Michel's fixative was capable of preserving in vivo tissue fixed immunoglobulin and complement for 8 months. This was demonstrated by positive direct immunofluoresence spaces of the epidermis in all of the skin biopsies examined. In addition, it was shown that refrigerated and non-refrigerated skin biopsies in Michel's fixative both yielded a diagnostic quality of direct immunofluorescence. This will allow the mailing of skin biopsies without requiring refrigeration and specimens mailed in the winter should demonstrate reliable results on fluorescent antibody testing when performed.

Specificity of Autoantibodies in Autoimmune Thrombocytopenia

In 42 patients with autoimmune thrombocytopenia (AITP) and a positive direct platelet suspension immunofluorescence test (PSIFT), the antigenic specificity of the autoantibodies was studied. Because the autoantibodies were often not detectable in the serum and additional HLA antibodies may disturb the reaction pattern with the platelet panel, we used eluates prepared from the patients' platelets for this study. Thirty-five patients had antibodies equally reactive with normal platelets, irrespective of their antigenic make-up, but not with the platelets from two Glanzmann's disease patients. Absorption and elution experiments in two patients showed that his was probably not due to the presence of a combination of anti-Zwa and anti-Zwb antibodies. Thus, the majority of autoantibodies against platelets seems to be directed against antigenic determinants not present on Glanzmann's disease platelets, but perhaps located on the platelet-membrane glycoproteins IIb and/or IIIa. In ten patients, antibodies of no, or still unknown, specificity were detected. Three of these had additional antibodies not reactive with the platelets of the two Glanzmann patients.

Specificity of autoantibodies in autoimmune thrombocytopenia

In 42 patients with autoimmune thrombocytopenia (AITP) and a positive direct platelet suspension immunofluorescence test (PSIFT), the antigenic specificity of the autoantibodies was studied. Because the autoantibodies were often not detectable in the serum and additional HLA antibodies may disturb the reaction pattern with the platelet panel, we used eluates prepared from the patients' platelets for this study. Thirty-five patients had antibodies equally reactive with normal platelets, irrespective of their antigenic make-up, but not with the platelets from two Glanzmann's disease patients. Absorption and elution experiments in two patients showed that his was probably not due to the presence of a combination of anti-Zwa and anti-Zwb antibodies. Thus, the majority of autoantibodies against platelets seems to be directed against antigenic determinants not present on Glanzmann's disease platelets, but perhaps located on the platelet-membrane glycoproteins IIb and/or IIIa. In ten patients, antibodies of no, or still unknown, specificity were detected. Three of these had additional antibodies not reactive with the platelets of the two Glanzmann patients.

Juvenile pemphigus vulgaris

Although pemphigus vulgaris is uncommon in adolescence, its occurrence in two patients in this age group is reported. In both cases the lesions were confined to the oral mucosa and the diagnosis was established by the typical histologic picture and positive direct and indirect immunofluorescence. The literature on childhood pemphigus vulgaris is reviewed, and the importance of prompt diagnosis by early biopsy and supportive immunofluorescent tests is emphasized.

Idiopathic thrombocytopenic purpura in children; detection of platelet autoantibodies by immunofluorescence.

There are 3 varieties of idiopathic thrombocytopenic purpura (ITP) in children: the acute, the chronic and the recurrent forms. We have tested the platelets from 80 children with these forms of ITP by an immunofluorescence test. Platelet-bound immunoglobulins were detected in 19/22 acute, 41/48 chronic and 5/10 recurrent cases of ITP, i.e. in 65 of a total of 80 cases. A significance differences between acute and chronic ITP was found in the observation that only IgM was bound to the platelets of 7/16 patients with acute ITP. In all cases of chronic ITP with a positive direct immunofluorescence test, IgG was present on the platelet membrane. It is assumed because some patients with chronic ITP do not have elevated platelet-bound immunoglobulins, and show refractoriness to immunosuppressive therapy, that in these patients the disorder is not of a humoral autoimmune nature.

Experimentelle Candidiasis: Pathogenese und serologische Untersuchungen in einem Kaninchenmodell

Three methods of inducing candidiasis in a rabbit model were studied in an effort to better understand the host responses and pathogenesis in this infection. Serological studies of both antigen and antibody were included. Endocarditis and cystitis were induced by inserting catheters containing viableCandida albicans into the right carotid artery and bladder, respectively, and pyelonephritis was induced by intravenous inoculation of viableCandida albicans. All rabbits with cystitis had evidence of infection only in the urinary bladder. Blood cultures in only one (with endocarditis) of 31 rabbits were positive forCandida albicans. All rabbits withCandida albicans endocarditis and pyelonephritis developed agglutinating antibody and immunodiffusion bands within seven to fourteen days from induction of infection, while none of the rabbits withCandida albicans cystitis developed agglutinating antibody titer changes or immunodiffusion bands. All rabbit urine and serum samples were examined forCandida albicans antigen by a latex agglutination and gel diffusion test. No free antigen was detected in any of the specimens using the methods described. A direct immunofluorescence technique was utilized to detect antibody coated yeast cells in the urine of infected animals as evidence of invasive disease. All rabbits with endocarditis, and four of eight rabbits with pyelonephritis had positive direct immunofluorescence studies of their urine, while all rabbits with cystitis had negative urines for antibody coated yeast cells.

Immunofluorescence findings in human renal amyloidosis.

Positive direct immunofluorescence findings have been occasionally reported for human renal amyloidosis. However, no substantial information describing the frequency of immunomicroscopic pattern has been reported. Twenty-seven renal biopsy specimens with proven amyloidosis were studied by direct immunofluorescence using monospecific antisera against IgG, IgA, IgM, IgD, IgE, kappa and lambda light chains, C3, C4 properdin, alpha-2 macroglobulin, albumin, transferrin, and fibrinogen. Ten biopsy specimens were negative. The remaining specimens showed principally a diffuse immunofluorescent staining of the mesangial areas with various of the above fluorescein-labeled antisera except IgE, albumin, and transferrin antisera, which gave a consistently negative reaction. IgD, properdin, or fibrinogen were weakly identified in only one case. Immunoglobulins and C3 were observed in 14 cases. Kappa and/or lambda light chains were found in nine of ten biopsy specimens so tested. Positive immunofluorescent staining of other elements of the nephron was rarely observed. These results suggest that various immunoreactants, in addition to light chains, can be frequently detected in renal amyloidosis and that passive nonselective absorption of plasma proteins does not explain these immunomicroscopic findings.

Bullous Eruption of Diabetes Mellitus

Diabetes mellitus is a systemic disorder with cutaneous manifestations that need not correlate with gross derangements of intermediary metabolism. The lesions may be bullous, necrobiotic, or atrophic and hyperpigmented, as in diabetic dermopathy. 1 In a recent study of a patient with the bullous eruption of diabetes mellitus, 2 findings from both direct and indirect immunofluorescence microscopy examinations of the skin were negative. We have found positive direct perivascular immunofluorescence in involved tissue from a patient with the bullous eruption of diabetes mellitus. Report of a Case A 73-year-old woman had a two-year history of adult-onset diabetes mellitus that was controlled with 25 units/day of isophane insulin suspension. Complicating her disease were peripheral neuropathy and diabetic retinopathy. She had no other medical problems and was taking no additional medication. One month before examination, an asymptomatic bullous eruption had developed, which had first manifested as a 2 × 2-cm bulla on