Evaluation of gelatin particle agglutination assay for the detection of anti-PGLI antibodies. Comparison with ELISA method and applicability on a large scale study using blood collected on filter paper.

Given the technical difficulties of the ELISA method, a gelatin particle agglutination test (MLPA) has been developed recently for the detection of anti-PGLI antibodies. The purpose of this study was to compare these 2 tests. MLPA was found to be less specific than ELISA (91% versus 98%, chi 2 = 66.8, p less than 0.001). The sensitivity of both tests was of 95% for the diagnosis of multibacillary patients. In the case of paucibacillary patients. MLPA was found to be less sensitive than ELISA (21% versus 35%, chi 2 = 6.98, p greater than 0.01). The agreement between the 2 tests for a positive or a negative result was satisfying (85% to 100%), except for the weakly seropositive individuals (71%). The correlation between OD obtained with ELISA and antibody titre obtained with MLPA was statistically significant (r = 0.70, p less than 0.001). Conversely to ELISA, MLPA was not applicable on blood samples absorbed on filter paper without a serious loss of sensitivity. In conclusion, this study demonstrated that the MLPA test can only reliably detect anti-PGLI antibodies in multibacillary cases.     

Clinical usefulness of determining the antibodies to soluble nuclear antigens in various collagen diseases

The purpose of the study was evaluation of the clinical usefulness of determination of antibodies to soluble nuclear antigens. The study was carried out in 71 cases of various collagen diseases. Antibodies dsDNA (IIF method with Crithidium luciliae as substrate) were found only in patients with SLE and renal involvement. RNP antibodies (double immunodiffusion method) were demonstrated in 83.3% of cases of mixed connective tissue disease, and Sm antibodies in 8% of SLE patients. It is worth stressing that in the presented material Sm antibodies were present only in association with RNP antibodies. Antibodies Ro and/or La were present most often in the sera of patients with SCLE, while Scl 70 antibodies were a marker of systemic sclerosis, more frequent in patients with diffuse scleroderma, while their demonstration in acroscleroderma suggested a more severe course of the disease. The study showed a high diagnostic and prognostic value of antibodies to soluble nuclear antigens in collagen diseases.     

Utility of gelatin particle agglutination test (MLPA) for rapid serodiagnosis of leprosy in a hyperendemic area.

The anti-PGL M. leprae specific antibodies were estimated by MLPA test in 79 patients of leprosy, 8 contacts of lepromatous cases and 10 healthy controls in a hyperendemic area. The results indicated an over all seropositivity of 50.6% in leprosy patients. Three of the eight contacts and five of the controls also gave positive results. Higher seropositivity rates were noted in multibacillary patients (73% in lepromatous, 53.6% in borderline, 40% each in tuberculoid and indeterminate and 10% in pure neuritic types). The practical application of MLPA test in its present form as a serodiagnostic procedure for screening subclinical or clinical infections in leprosy patients appear to be of limited value in hyperendemic areas. Further studies involving large series of subjects are necessary for reaching definite conclusions.     

Anti-sperm antibodies in homosexual men: prevalence and correlation with sexual behaviour.

The sera of 60 homosexual males were examined for the presence of antibodies to sperm using an indirect immunobead test (IBT). Six of 60 (10%) had antibodies of IgG isotype; in addition two of the six had antibodies of IgA isotype. The presence of antibodies was associated with the practice of unprotected receptive anal intercourse in the previous six months. Antibodies were not found in homosexual men who were celibate, or who practised only oral intercourse during the same period. There was no correlation between the presence of anti-sperm antibodies and antibodies to human immunodeficiency virus (HIV), or numbers of T lymphocytes. These preliminary results lend support to the hypothesis that antigen presentation in the lower gut may be a source of sensitisation against sperm. The possibility that anti-sperm antibodies may be a marker of receptive anal intercourse merits further investigation.     

Evaluation of Mycobacterium leprae particle agglutination test, using eluates of filter paper blood spots.

A comparison of the ELISA test with the newly-developed MLPA test was carried out, using eluates of blood spots from filter paper for the detection of the anti-PGL-I antibody. A very good positive correlation was observed between these two tests. The concordance rate was found to be 92.6%, ranging from 71.4% to 100%. This nonconcordance was not found when freshly-collected samples were used. The MLPA test is simple and reliable. The use of eluates from blood spots collected on filter paper further simplifies the test in the collection and transportation of blood samples. This accurate and rapid method makes the MLPA test logistically feasible for large-scale screening. With our modification of MLPA with eluates more samples can be screened with the kit than with sera.     

Antinuclear antibodies during Puva therapy.

During PUVA therapy 7 patients out of 34 with severe psoriasis developed circulating antinuclear antibodies (ANA) (21%). Before treatment only 3 patients of 50 (6%) considered for PUVA had detectable ANA. The ANA titres were usually low. Antibodies against native DNA as studied with the Crithidia luciliae test, were not found, and blood and urinary screening for collagenosis was negative. All 7 patients responded well to the PUVA treatment. The significance of these findings remains to be determined.     

Serologic studies in patients with lupus erythematosus and psoriasis

We present four patients with coexistent lupus erythematosus (LE) and psoriasis. This is an unusual combination. All four patients had antibodies to Ro, which were absent in twenty-four control psoriatics. Antibodies to Ro occur in only 25% to 30% of unselected SLE patients, but occur in approximately 60% of "antinuclear antibody (ANA)-negative" LE patients, many of whom are highly photosensitive. The increased frequency of anti-Ro in our patients suggests that this may be a specific serologic marker for the LE/psoriasis overlap. Also, since anti-Ro correlates with photosensitivity, LE/psoriasis overlap patients may be at increased risk for photosensitivity, which occurred in two of our patients, one of whom developed severe systemic disease following ultraviolet (UVB) phototherapy. Screening for anti-Ro antibodies may be appropriate in psoriatics prior to UVB phototherapy.     

Oral lichen planus and HCV infection: a clinical evaluation of 263 cases

Background Hepatitis C virus (HCV) infection induces variable dermatologic manifestations. Our purpose was to determine whether there is an association between HCV infection and oral lichen planus (OLP). Methods Antibodies to HCV were determined in patients with OLP (263 patients; 156 women and 107 men, with a mean age of 55.5 years) and in a control population. Results Seventy six cases (28.8%) were positive for HCV antibodies with the second-generation enzyme-linked immunosorbent assay (ELISA II) test. All of these cases were confirmed with the second-generation recombinant immunoblot assay (RIBA II) test. In 61 cases (23.1%), high levels of serum transaminase were found. Positivity for hepatitis B virus (HBV) markers was found in 31 patients (11.7%) and for hepatitis A virus (HAV) markers in 43 patients (16.3%). None had positivity for hepatitis D virus (HDV) markers. As a control group, we used 100 patients (58 women and 42 men, with a mean age of 55.3 years) referred to the School of Dentistry of the University of Naples “Federico II,” and treated for general dental caries. In the control group, HCV antibody positivity was found in three cases. Conclusions The high prevalence of HCV antibody in this group of patients with OLP, higher than in the healthy population, suggests a link (p = 1.423 × 10^–7, chi-squared test) between these two diseases. These findings stress the importance of liver examination in OLP patients, and the need for other studies on the high susceptibility to hepatitis viruses in the population in the southern part of Europe.     

Subacute cutaneous lupus erythematosus: Clinical, serologic, and immunogenetic studies of forty-nine patients seen in a nonreferral setting

Subacute cutaneous lupus erythematosus has been clearly recognized as a distinct cutaneous manifestation of lupus erythematosus. Two forms have been described, an annular erythema and a papulosquamous variant. Previous data have suggested that these patients have a high incidence of mild to moderate systemic disease, anti-Ro (SS-A) antibodies, and human lymphocyte antigen (HLA)-DR3, particularly the annular form. We studied forty-nine patients with subacute cutaneous lupus erythematosus seen in local private practices in our area. Lesions of chronic cutaneous lupus erythematosus were seen in 34.8% of our patients. Twenty-five patients (51%) fulfilled the American Rheumatism Association criteria for the classification of systemic lupus erythematosus, and renal disease was present in nine of these patients (including 3 with decreased function). Antibodies to Ro (SS-A) and/or La (SS-B) were present in only sixteen patients, and HLA-DR3 was found in only seventeen patients. Twenty-two patients had inactive cutaneous disease at follow-up. We concluded that our patient population with subacute cutaneous lupus erythematosus skin lesions is less distinctive than previous literature suggests. The serologic and immunogenetic correlates were not demonstrated. The full range of lupus erythematosus-related disease was seen, although most patients follow a benign course.     

Chlamydial genital infection in Ibadan, Nigeria. A seroepidemiological survey.

Sera from patients attending a sexually transmitted diseases (STD) clinic, a family planning clinic, and an antenatal clinic in Ibadan, Nigeria, as well as from male blood donors from the same area were tested for the presence of type specific antichlamydial antibodies using a modified micro-immunofluorescence test. Among men and women attending the STD clinic the exposure rates to Chlamydia trachomatis serotypes D to K (genital pathogens) were 18.7% and 26.7% respectively. Antibody titres suggesting active disease in these men and women were found in 11.8% and 22.7% respectively. The highest rate of exposure (35%) was among women attending the family planning clinic; of these women 25% had antibody suggesting active disease. Titres of IgG antibody in this study were similar to those found among men and women with chlamydial genital infections in the United Kingdom. Antibodies to serotypes D to K were also detected in 10.3% of women attending an antenatal clinic and in 9.9% of male blood donors. The prevalence of antibodies to C trachomatis serotypes A to C and lymphogranuloma venereum serotypes was low. These results suggest that the prevalence of chlamydial genital infections in Ibadan, both among STD patients and especially among those individuals not seeking treatment (family planning and antenatal clinic patients), is high. Since serious sequelae can follow chlamydial genital infections it is imperative to carry out further investigations in this area.     

Detection of Antibodies to Human Melanoma Cells in Vitiligo and Alopecia Areata by Western Blot Analysis

Antibody reactivity to human melanoma cells (SK-Mel-23) was investigated in 48 patients with vitiligo, 14 with alopecia areata (AA), and 35 normal control individuals by Western blot analysis. Antibodies to SK-Mel-23 were found in 44 (92%) of the patients with vitiligo, in 7 (50%) of the patients with AA, and in 14 (40%) of the normal control individuals. Significant differences between patients with vitiligo and normal controls were found in the incidence and distribution of antibodies, but no significant differences were found between patients with AA and normal controls. The antibodies were predominantly directed to one or more antigens of approximately 110 KD, 103 KD, 88 KD, 70 KD, 56 KD, 46 DK, or 41 KD. The most common responses were to 110 KD, 88KD, and 70 KD antigens. These antibodies were present in 60%, 60%, and 73% of the patients with vitiligo; 7%, 14% and 35% of the patients with AA; and 11%, 11% and 40% of normal control individuals, respectively. There were no statistical differences in the incidence of antibodies to pigment cells between segmental and non-segmental vitiligo. These findings suggest that autoreactivity to pigment cells occurs mostly in patients with vitiligo and might be a secondary immune reaction to destroyed pigment cells.     

Seroprevalence of human herpesvirus 8 and its association with Kaposi sarcoma in Brazil

BACKGROUND: Epidemiologic studies have indicated that human herpesvirus 8 is implicated in the development of Kaposi sarcoma in different geographic areas worldwide. GOAL: To provide information on the prevalence of human herpesvirus 8 in Brazil and its association with Kaposi sarcoma. STUDY DESIGN: An immunofluorescence assay was performed to test 1,044 serum samples from 747 blood donors, 73 patients presenting to casualty departments, and 224 patients attending sexually transmitted disease/AIDS clinics. The sexually transmitted disease group was composed of 88 patients with HIV (40 Kaposi sarcoma positive, 48 Kaposi sarcoma negative) and 136 patients without HIV. RESULTS: Antibodies to human herpesvirus 8-latent nuclear antigens were found in 34 blood donors (4.6%), and in seven casualty patients (9.6%). The highest frequency of human herpesvirus 8 antibodies was found in the sexually transmitted disease group: 32 HIV-positive patients with Kaposi sarcoma (80%) and seven patients without Kaposi sarcoma (14.6%). CONCLUSION: The presence of human herpesvirus 8 in patients with HIV was strongly associated with Kaposi sarcoma (odds ratio, 23.4; 95% CI, 7.7-71.4), male gender, homosexual or bisexual orientation, and hepatitis B virus infection, but not with the other sexually transmitted diseases that were investigated.     

Sensitivity of indirect immunofluorescence and ELISA in detecting intercellular antibodies in endemic pemphigus foliaceus (Fogo Selvagem)

BACKGROUND: Since 1967 dermatology has used the classic technique of indirect immunofluorescence (IFI) for the detection of autoantibodies against antigens of the skin in diseased people with endemic pemphigus foliaceus. Thirty years later enzyme-linked immunosorbent assays--ELISA (rDsg1 and rDsg3) appeared as a viable option. A group of highly recognized researchers have concluded that ELISA is a simple, sensitive and highly specific method, allowing for diagnostic differentiation between pemphigus vulgaris (PV) and endemic pemphigus foliaceus (EPF). Scientific literature certifies that both ELISA and IIF bear high sensitivity in spite of the fact that a direct comparison between the ELISA and IIF tests has never been performed. OBJECTIVES: This study was conducted to compare the sensitivity of these tests in detecting antibodies in the EPF. MATERIAL AND METHODS: Thirty-two serum samples were collected from patients with EPF. The control serum of 15 healthy individuals was tested to detect the presence of antibodies of EPF by indirect immunofluorescence and ELISA (rDsg1 and rDsg3). The IIF was performed, taking human skin as a substrate. RESULTS: Antibodies in patients with EPF were detected more commonly by the ELISA (rDsg1) (91%) compared with IIF (81%). CONCLUSIONS: The ELISA (rDsg1) is slightly more sensitive than IIF in detecting antibodies related to EPV. However, according to our results, we do not currently possess a test with 100% accuracy in differentiating EPF from PV. Although previous studies have associated Dsg3 with PV, the tests performed during this study showed that 12% (4/32) of patients with EPF (cutaneous diseases only) also had Dsg3 antibodies.     

Sexually transmitted diseases, antibodies to human immunodeficiency virus, and subsequent development of acquired immunodeficiency syndrome. Visitors of homosexual sauna clubs in Copenhagen: 1982-1983

Sera from 260 men from Denmark and elsewhere attending two Copenhagen sauna clubs for homosexual men during nine months of 1982-1983 were investigated for markers for syphilis, hepatitis A and B, and human immunodeficiency virus (HIV). Five per cent (12 men) had active syphilis, and another 35% (92) had a history of and/or serologic markers for syphilis. Ninety-four men (36%) were positive for antibodies to hepatitis A virus, ten (4%) were positive for hepatitis B surface antigen (HBsAg), and 153 (59%) were positive for antibodies to HBsAg. Antibodies to HIV were found in 45 (20%) of the 220 men investigated for this marker. Markers for hepatitis A and B and for syphilis were more frequent in the HIV antibody-positive individuals, but the association was significant only for markers for hepatitis B (relative risk = 2.0). Thus STD markers had little predictive value for seropositivity for antibodies to HIV. Among 37 men investigated more than once, a seroconversion rate of 3% per month for antibodies to HIV was found, but this estimate must be taken with reservation. The rate of seropositivity for antibodies to HIV among men from Denmark was 23%, and three (8%) of the 40 HIV-positive Danish men developed the acquired immunodeficiency syndrome (AIDS) during the four years following the initial investigation. This study shows that by 1982-1983 HIV had spread considerably in the Danish high-risk group, although there were only seven reported cases of AIDS in the country at that time.     

Anti-phenolic glycolipid 1 IgM antibodies in leprosy patients and in their household contacts

Though they have no apparent protective action, the specific antibodies are important markers of the infection with Mycobacterium leprae. For their detection we employed an ELISA method using as substrate a synthetic immunodominant disaccharide of phenolic glycolipid 1 antigen of M. leprae, conjugated with bovine serum albumin (D-BSA). Increased levels of anti-D-BSA antibodies of the IgM class were detected in 61.5% of the 13 leprosy patients and in 13.3% of their 53 household contacts, whereas they were not found in any of the 37 normal blood donors. A strong correlation (r = -0.846) was found between the antibody levels and the duration of the disease among the 12 patients with lepromatous leprosy. These preliminary data demonstrate the usefulness of this method for epidemiological studies and for the detection of cases with subclinical infection.     

A Clinicopathological Study of Pemphigus in Eastern India with Special Reference to Direct Immunofluorescence

Background:Pemphigus is a group of chronic autoimmune vesico-bullous disorders in which the epidermis and the basement membrane zone are the focus of attack resulting in cutaneous and mucosal blister formation. Direct immunofluorescence (DIF) test is a very sensitive test for the diagnosisAim:To study the clinico histopathological patterns of pemphigus in eastern India. The study also aims to correlate DIF with clinical and histologic findings as well as severity of skin involvement [scoring systems].Results:In our study Pemphigus vulgaris (PV) was the predominant type with 32 cases followed by 8 cases of pemphigus foliaceus (PF) and a single case of IgA pemphigus. Mean age at presentation was late middle age. Majority of the patients, 26 (63.41%) initially had cutaneous involvement followed by mucosal involvement. In this study group 36 (87.80%) patients showed acantholytic cells on histopathological examination. Most patients of PV showed suprabasal blister 20 (62.50%) followed by intraspinous 5 (15.62%) and subcorneal 5 (15.62%) blister. In majority 28 (87.50%) of the PV patients IgG and C3 antibodies were deposited throughout the epidermis. The strength of antibody positivity was strong in most of the patients (71.87%). In cases of PF mostly IgG 6 (75%) antibodies were deposited in the upper epidermis. DIF intensity had poor correlation with disease activity/severity except in PF.Conclusion:Almost 85.36% cases of pemphigus were diagnosed clinicopathologically. But 6 cases couldn’t be diagnosed accurately on clinicopathological basis and in them DIF was confirmatory. Two cases of pure mucosal PV and 1 case of IgA pemphigus was confirmed by DIF. Two cases of bullous pemphigoid clinico-histologically mimicking PV were also excluded by DIF. So it appears from our study that DIF is confirmatory for diagnosis of pemphigus in all cases.     

Antibodies to UV DNA and photosensitivity

Antibodies to ultraviolet light denatured DNA (UV DNA) have been measured in the sera of patients with systemic lupus erythematosus, discoid lupus erythematosus, and light sensitive skin lesions. Antibodies were found in significant levels in the patients with SLE but not in the other groups. It appears that although UV denaturation of dermal DNA occurs in vivo this is not sufficient to induce antibodies to UV DNA in patients with a normal DNA repair mechanism. This may not be the case in patients with SLE in whom a break in tolerance to Native (N-) DNA has already occurred and where antibody to N-DNA will cross react with UV DNA.     

Anti-collagen antibodies in granuloma annulare and necrobiosis lipoidica

Antibodies to native collagens I-V were measured by enzyme-linked immunosorbent assay (ELISA) in sera from 32 patients with granuloma annulare (GA), 32 patients with necrobiosis lipoidica (NL) and 31 healthy controls. Antibodies to all collagen types were detected in all patient groups and in normal controls. No difference in antibody levels between patient and control groups was found except for a significant reduction in collagen type III antibodies in patients with GA compared with normal control subjects.     

Spectrum of antinuclear and anti-cytoplasmic antibodies in dermatomyositis and polymyositis overlap syndromes

Antinuclear antibodies (ANA) of the IgG class were detected in 60% of patients with dermatomyositis. Only in 16% could we demonstrate precipitating antibodies against the nuclear Mi-2 antigen. These antibodies are considered to be serological markers of dermatomyositis. The ANA spectrum in polymyositis-overlap syndrome was considerably more heterogeneous: patients with polymyositis/progressive systemic scleroderma overlap had antibodies against the nucleolar PM-Scl antigen or the nuclear Ku antigen. Cytoplasmic antibodies to Jo-1 were present in polymyositis associated with Sj?gren's syndrome and pulmonary fibrosis. Antibodies against ribosomal ribonucleoprotein were found in polymyositis with systemic lupus erythematosus and antibodies to nuclear ribonucleoprotein in polymyositis associated with mixed connective tissue disease. The investigation demonstrates that the characterization of ANA specificities helps to differentiate between dermatomyositis and distinct forms of polymyositis-overlap syndrome.     

CSF diagnosis in syphilis

Treponemal antibodies in CSF are often to be found by FTA-ABS and TPHA, also in cases without syphilis cerebrospinalis. The checking of the CSF barrier allows--together with the TPHA-index--the diagnostic of the syphilis cerebrospinalis. Antibodies against treponema pallidum have been proved in 15 patients at different stages of syphilis until 30 years after treatment, but the titres were low. Only in one case we have found an active syphilis cerebrospinalis by the help of the TPHA- and IgG-index.