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A method for identifying causative chemicals of allergic contact dermatitis using a combination of chemical analysis and patch testing in patients and animal groups: application to a case of rubber boot dermatitis.

A 63-year-old woman developed allergic contact dermatitis from rubber boots. Initial investigation, by patch testing in the patient and chemical analysis of the causative rubber boots, revealed that mercaptobenzothiazole (MBT) and dibenzothiazyl disulfide (MBTS) were the causative chemicals. Subsequent investigations were performed by patch testing in animal groups. An extract of the causative rubber boots, MBT and MBTS were used for sensitization of guinea pigs by the guinea pig maximization test (GPMT). 3 animal groups, A (with the boot extract), B (with MBT) and C (with MBTS) were successfully prepared. The boot extract was fractionated by column chromatography and thin-layer chromatography (TLC). Each fraction was subjected to patch testing in the animal groups. Positive reactions in all groups would show that the active fractions contained MBT-type compounds, whereas a positive reaction in group A but negative ones in group B and C would show that the active fractions did not contain any MBT-type compounds. Each fraction was then analyzed by gas chromatography (GC), GC-mass spectrometry (GC-MS), direct inlet-MS (DI-MS) and high-performance liquid chromatography (HPLC). By this investigation, we found not only known allergens (MBT, MBTS), but also unknown allergens: S-substituted MBT-type compounds and styrenated phenol (SP). Thus, SP was shown to be a candidate as a human sensitizer even though the patient did not react to it.

Allergens↗

Comparison of irritant potential of shampoos using cultured human epidermal keratinocytes model and patch test reaction measured by laser Doppler flowmetry.

The keratinocyte culture model has previously been used as an in vitro method for testing skin irritating potential of common skin irritants. However, solubility limits its use for finished products. Shampoo is very soluble in water which should make it an ideal product category for the cell culture model. To determine the skin irritant potential of several commercial shampoos, we employed cultured human keratinocytes as an in vitro model. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide test (MTT) and lactic dehydrogenase release (LDH) test were used to document cell toxicity. 7 volunteers were patch tested and their reactions evaluated using laser Doppler flowmetry and compared with the in vitro data. MTT and LDH have a good negative correlation with each other. Patch test reaction, especially at high concentrations, correlates relatively well with the in vitro test, especially with shampoos of strong and weak irritancy. However, the rank order of the shampoos of moderate toxicity was not the same as in the in vitro data. This suggests that the cell culture technique cannot directly replace in vivo methods, and that data obtained by the cell culture method should be interpreted carefully.

Adult↗

Easier patch testing with TRUE Test.

TRUE Test, a standardized, ready-to-apply patch test system, is made from polyester covered with a film of allergens incorporated in a hydrophilic polymer. The patches are mounted on nonwoven cellulose tape with acrylic adhesive, covered with siliconized plastic, and packed in an air-tight and light-impermeable envelope. When the test strip is taped on the skin, perspiration hydrates the film and transforms it to a gel, which causes the allergen to be released. The first panel of 12 allergens and allergen mixes is standardized and tested for stability in vitro and in vivo. The accuracy of the test panel has been certified in international multicenter studies by comparing it with present patch test techniques. A second panel of 11 more allergens was completed in 1988. The two test panels include the full standard panel of the North American Contact Dermatitis Group.

Dermatitis, Contact↗

A pilot study of the usefulness and safety of a ready-to-use atopy patch test (Diallertest) versus a comparator (Finn Chamber) during cow's milk allergy in children.

BACKGROUND: Patch testing is used in the diagnosis of food allergy, especially during delayed manifestations. OBJECTIVE: A ready-to-use atopy patch test (APT), the Diallertest, was compared with another APT device, the Finn Chamber, in pediatric cow's milk allergy. METHODS: This prospective study involved 49 children (34.3 +/- 17 [mean +/- SD] months of age), with cow's milk allergy manifested by atopic dermatitis (10.2%), digestive manifestations (40.8%), or both (49%). All children underwent both APT techniques, with a reading 72 hours after application, followed by a milk elimination diet for 4 to 6 weeks and open cow's milk challenge. RESULTS: A positive result was seen in 22 (44.8%) versus 13 (26.5%) patients with the ready-to-use and the comparator APTs, respectively. No side effects were recorded. Both techniques were concordant in 67.3% of patients. Of the total 41 open cow's milk challenges, 60.9% had positive results, with 8 patients lost to follow-up. The performances of the ready-to-use and comparator APTs were as follows: sensitivity, 76% (95% CI, 59.2% to 92.7%) versus 44% (95% CI, 24.5% to 63.4%; P = .02); specificity, 93.8% (95% CI, 81.9% to 100%) versus 93.8% (95% CI, 81.9% to 100%); positive predictive value, 95% (95% CI, 85.4% to 100%; 1 false-positive result) versus 91.7% (95% CI, 76% to 100%; 1 false-positive result); negative predictive value, 71.4% (95% CI, 52% to 90.7%; 6 false-negative results) versus 51.7% (95% CI, 33.5% to 69.8%; 14 false-negative results); and test accuracy, 82.9% (95% CI, 71.3% to 94.5%) versus 63.4% (95% CI, 48.6% to 78.1%; P = .05). CONCLUSION: The ready-to-use APT exhibited a good sensitivity and specificity, with no side effects.

Child↗

Six-hour versus 48-hour patch testing with varying concentrations of potassium dichromate.

This study examines the possibility of using patch contact times shorter in duration than the standard 48 hours. Using varying concentrations of potassium dichromate (K2Cr2O7), this study analyzed results from two sets of patches applied to the backs of 11 subjects for durations of 6 and 48 hours, respectively. Results showed that after the 48-hour application period, all subjects reacted to K2Cr2O7 at some concentration. For the patches applied for 6 hours, 7 of the 11 subjects (64%) reacted to K2Cr2O7 at some concentration. Minimum elicitation thresholds (METs), the lowest concentration at which a reaction was observed, were established for both the 6-hour and 48-hour application times. The ratio of an individual's 6-hour MET to their 48-hour MET was calculated to evaluate the feasibility of patch testing with a higher concentration of an allergen for a shorter time period. Although the results clearly indicated that a higher concentration of allergen is required in order to elicit a reaction at 6 versus 48 hours, a fair amount of interindividual variability is exhibited by these 6-hour to 48-hour MET ratios. This observed variability would seem to preclude the use of 6-hour duration patch contact times for routine patch testing with K2Cr2O7.

Dermatitis, Contact↗

Immunohistochemical studies in mite antigen-induced patch test sites in atopic dermatitis.

The role of mite allergen in atopic dermatitis is still unclear. In this study, we investigated whether an eczematous reaction could be induced by patch testing with dust mite antigen. We succeeded in reproducing an eczematous lesion and the mite RAST-positive AD group showed a positive reaction much more than the RAST-negative group. Many mite antigen-bearing Langerhans cells, also possessing IgE molecules, were found by the use of an immuno-double labelling technique. By using immunoelectron microscopy, it was observed that the mite antigens were trapped by some macrophages, which were apposed to lymphocytes. To investigate the time-course of the reaction, the patch test reactions were read and biopsied after 1 h, 6 h, 24 h and 48 h. An eczematous reaction developed 24 h after patch testing. The mite antigen-bearing Langerhans cells were seen exclusively in the epidermis after 6 h, and mainly in the dermis after 24 h and 48 h. These results suggested that IgE-mediated contact hypersensitivity to mite antigen may develop and play an important role in AD.

Adolescent↗

Percutaneous absorption of gold sodium thiosulfate used for patch testing.

Contact allergy to gold has been shown to be correlated to wearing of golden jewellery as well as to the presence of dental gold. The prerequisite for sensitization and elicitation of an allergic contact dermatitis to gold is ionization and percutaneous absorption of the allergen. The present study was undertaken with the aim to demonstrate gold in blood after an occlusive patch testing with gold sodium thiosulfate (GSTS). Patients were patch tested with GSTS 0.5, 2.0 and 5.0% in petrolatum. Venous blood was sampled before testing as well as on day 3 (D3) or day 7 (D7). Blood gold levels were analysed by inductively coupled plasma mass spectrometry. In all 66 patients, the blood level of gold was significantly higher in the 2nd blood sample than in the 1st (median 0.34 versus 0.03 micro g/l). The increase was similar in patients allergic and not allergic to gold. There was no difference between gold levels on D3 and on D7 after epicutaneous application. Presumably, other allergens may also be taken up in blood during patch testing.

Adult↗

Sodium lauryl sulphate for irritant patch testing--a dose-response study using bioengineering methods for determination of skin irritation.

The dose-response relationship in patch testing with sodium lauryl sulphate (SLS) was studied. The irritant skin response was quantified by visual scoring as well as by the following noninvasive methods: measurement of transepidermal water loss (TEWL) by an evaporimeter, measurement of skin color by a colorimeter, measurement of superficial blood flow by laser Doppler flowmetry, and measurement of edema in the skin by ultrasound A-scan. Twelve volunteers were patch tested with 0.12, 0.25, 0.50, and 1.00% SLS, and the skin response was evaluated after 24 and 48 h, respectively. We found a statistically significant linear dose-response relationship between dose of SLS and skin response evaluated by measurement of TEWL, skin color, superficial blood flow, and edema. Statistical evaluation by regression analysis proved measurement of TEWL to be the method best suited overall for quantification in relation to patch testing with SLS, whereas colorimetry was found to be the least sensitive of the applied methods. Ultrasound A-scan was found to be a promising method for quantification of the inflammatory response, being consistently more sensitive than measurement of skin color.

Adult↗

Visual and laser Doppler perfusion scanning assessments of Kathon CG patch test reactions.

BACKGROUND: Testing procedure for Kathon CG (Diagnostics AB, Tygelsjö, Sweden) is not standardized. OBJECTIVE: To improve test results for Kathon CG. METHODS: Eleven subjects were tested with Kathon CG patch tests applied from 6 to 96 hours. Tests were read visually and by a laser Doppler perfusion scanning technique at time points ranging from 6 to 240 hours. RESULTS: There were large differences in the perfusion profiles of subjects tested in identical fashion. Beginning reactions were detected earlier with the laser Doppler scanning than with the visual assessment technique. There was no obvious agreement between peak perfusion values of reactions and application times. Both assessment techniques were effective at distinguishing between positive and negative subjects at an adequate dose and application time. This distinction was possible using a shorter reading time with the laser Doppler scanning for all subjects. Six-hour applications were sufficient to detect all positive subjects so tested regardless of assessment technique. With that application time, positive and negative subjects were most effectively identified using long reading times regardless of assessment technique. CONCLUSION: The shortest application time used on all subjects, 24 hours, was effective and 6-hour applications with long reading times for all subjects so tested. Doses lower than 0.0040 mg/cm2 resulted in false-negative tests.

Adult↗

Adhesion molecules in atopic dermatitis: upregulation of alpha6 integrin expression in spontaneous lesional skin as well as in atopen, antigen and irritative induced patch test reactions.

Atopic dermatitis is a lymphocyte-mediated skin disease. We studied the expression of the adhesion molecule alpha6 integrin by immunohistochemistry in spontaneous atopic inflammation as well as during the eliciting phase of atopen (Dermatophagoides pteronyssinus), antigen (nickel sulfate) and irritative (anthralin) induced patch test reactions in atopic skin. Results were compared with nickel sulfate patch test reactions in normal skin. A role of the alpha6 integrin, expressed at the luminal side of blood vessels, for T cell migration in lesional atopic skin was supposed. In normal human skin the alpha6 integrin was weakly expressed by blood vessels and by basal epithelial cells of the epidermis. In acute and chronic lesional skin of patients with atopic dermatitis dramatic upregulation of alpha6 integrin expression was observed on endothelial cells and in the epidermis. The similar pattern of upregulated suprabasal alpha6 integrin expression was established in the patch test reactions 48 h after atopen and antigen application or irritation of the skin without differences in dependence on the eliciting substance. No difference of alpha6 integrin expression was seen between atopic and normal skin. Tumor necrosis factor alpha, interleukin-1, interleukin-4 and interferon gamma play a role in atopic inflammation. Tumor growth factor beta and interleukin-6 are mitogenic/growth factors for keratinocytes. For this reason the effect of these cytokines and of phorbol-12-myristate-13-acetate on the expression level of alpha6 integrin was tested in short-term skin organ culture of normal and atopic skin as well as in keratinocyte cultures. In these assays no cytokines had an effect on alpha6 integrin expression suggesting another mechanism which regulates this integrin. However, the increased expression of alpha6 integrin in the suprabasal epidermis is associated with a T cell influx into the epidermis. We speculate that the alpha6 integrin expression may lead to an epidermotropism of T cells during inflammation.

Anthralin↗

The frequency of fragrance allergy in a patch-test population over a 17-year period.

Fragrances are widely encountered in our daily environment and are known to be a common cause of allergic contact dermatitis. We have reviewed our patch test data from 1980 to 1996 to establish whether the pattern of fragrance allergy has changed with time. During this period, 25,545 patients (10,450 male, 15,005 female) were patch tested with the European standard series. The mean annual frequency of positive reactions to the fragrance mix was 8.5% in females (range 6.1-10.9) and 6.7% in males (range 5.1-12.9). Females were 1.3 times more likely to be allergic to fragrance (P < 0.001, 95% confidence interval, CI 1.17-1.41). Males with fragrance allergy were older than females by 5.6 years (mean age 48.2 vs. 42.6 years; P < 0.001, 95% CI 3.9-7.3). The incidence of a concomitant positive patch test to balsam of Peru in fragrance-sensitive patients showed wide variation, suggesting that it is not a reliable marker of fragrance allergy. There was a positive correlation between the isomers isoeugenol and eugenol. Oak moss remained the most common overall allergen throughout the study, positive in 38.3% of females and 35.6% of males who were tested to the constituents of the fragrance mix. During the period of the study the incidence of positive tests to oak moss increased by 5% yearly (P = 0.001, 95% CI 2.2-8.7). The frequency of allergic reactions to eugenol and geraniol remained relatively constant. Isoeugenol and alpha-amyl cinnamic aldehyde sensitivity increased and hydroxycitronellal showed a slow decline. There was a striking reduction in the frequency of sensitivity to cinnamic aldehyde (by 18% yearly; P < 0.001, 95% CI 14.3-21.0) and cinnamic alcohol (by 9% yearly; P < 0.001, 95% CI 5.2-12.9); these are now uncommon fragrance allergens. These data show temporal trends which may reflect the frequency of population exposure to individual fragrances.

Acrolein↗

Oral challenge with nickel and cobalt in patients with positive patch tests to nickel and/or cobalt.

During a 3-year period, 146 patients with positive patch tests to nickel and/or cobalt, for whom a systemic cause of dermatitis was suspected, were challenged orally with a single dose of 2.5 mg nickel given as nickel sulphate and 1 mg cobalt given as cobalt sulphate using a double-blind, placebo-controlled method. No following-up was possible for 2 patients. Seventy-five of the remaining 144 (52%) had flares of their dermatitis after challenge with nickel and/or cobalt, while 45 patients (31%) had no reaction and 24 (17%) reacted only to the placebo or to the placebo in combination with nickel or cobalt. Seven out of 13 patients with positive patch tests to cobalt experienced flares of their dermatitis only after challenge with cobalt.

Administration, Oral↗

Immunophenotyping of the cutaneous cellular infiltrate after atopy patch testing in cats with atopic dermatitis.

Cats with spontaneously occurring atopic dermatitis have clinical and immunocytochemical characteristics compatible with these in humans with atopic dermatitis (AD). The atopy patch test (APT) has proven to be a valuable tool in elucidating the disease process in humans. Additionally, the APT is very specific and bypasses the problem of conflicting results due to differences in chronicity of lesions of AD patients. We adapted the APT for use in cats to explore the suitability of the APT as a tool to study the onset of allergic inflammation in cats with atopic dermatitis. APT were performed in AD cats (n = 6) and healthy cats (n = 10). All cats were patch tested with two allergens in three different dilutions and a diluent control. The allergens for the APT were selected from positive intradermal test and /or prick test results and consisted of: Dermatophagoides farinae, D. pteronyssinus, Tyrophagus putrescentiae, and a grass pollen mixture. APT were read after 10, 24 and 48 h, and punch biopsies for immunohistochemical evaluation were collected at these time points. Macroscopically positive APT reactions were observed in three out of six cats at 24 and/or 48 h with allergen concentrations of 25,000 and 100,000 NU/ml. Reactions were not observed at negative control sites and neither in control animals. A significantly increased number of IL-4+, CD4+, CD3+, MHC class II+ and CD1a+ cells was found in one AD cat with positive APT reactions. Five out of six AD cats had significantly increased IL-4+ T cell numbers at 24 and/or 48 h. Our data indicate that in cats, macroscopically positive patch test reactions can be induced, which have a cellular infiltrate similar to that in lesional skin. We found a high specificity and a macroscopically positive APT reaction in half of the cats, which is similar to what is seen in humans. Hence, the APT in cats might be a useful tool in studying the immunopathogenesis of feline atopic dermatitis.

Allergens↗

Patch testing with markers of fragrance contact allergy. Do clinical tests correspond to patients' self-reported problems?

The aim of the present study was to investigate the relationship between patients' own recognition of skin problems using consumer products and the results of patch testing with markers of fragrance sensitization. Eight hundred and eighty-four consecutive eczema patients, 18-69 years of age, filled in a questionnaire prior to patch testing with the European standard series. The questionnaire contained questions about skin symptoms from the use of scented and unscented products as well as skin reactions from contact with spices, flowers and citrus fruits that could indicate fragrance sensitivity. A highly significant association was found between reporting a history of visible skin symptoms from using scented products and a positive patch test to the fragrance mix, whereas no such relationship could be established to the Peru balsam in univariate or multivariate analysis. Our results suggest that the role of Peru balsam in detecting relevant fragrance contact allergy is limited, while most fragrance mix-positive patients are aware that the use of scented products may cause skin problems.

Adolescent↗

Patch test results in 542 patients with suspected contact dermatitis in Turkey.

In this retrospective study, patch test results of 542 patients (303 female and 239 male), referred to our clinic between June 1996 and July 1999, were evaluated. All patients were patch tested with 32 allergens of an extended European standard series, 140 patients were tested with supplemental series, and 246 patients with their own substances. 280 (51.7%) patients had 1 or more positive results but allergic contact dermatitis, according to clinical relevance, was diagnosed in only 190 (67.9%) of these. Nickel sulfate was the most frequent sensitizer (19.1%), followed by potassium dichromate (11.8%), palladium chloride (9.4%), cobalt chloride (8.5%), and thiuram mix (7.7%). Frequencies of contact allergy to fragrance mix (5.5%), balsam of Peru (Myroxylon Pereirae resin) (2%), quaternium-15 (0.6%), and Kathon CG (methylchloroisothiazolinone+methylisothiazolinone) (0.2%) were relatively low, while no positive reactions were obtained to paraben mix in any case. Current clinical relevance to the present dermatitis was most frequently seen with carba mix (96.2%), thiuram mix (95.2%) and potassium dichromate (78.1%), whereas this rate was low for nickel (37.5%). In 85.8% of patients with allergic contact dermatitis, the responsible allergen was detected by testing with the extended standard series alone (18.4% identified by testing with the additional 10 allergens), in 11.6% by testing with the patients' own substances, and in 2.6% by testing with the supplemental series. Occupational contact dermatitis was diagnosed in 77 of the patients with positive reactions (27.5%), most of these being construction workers and house painters who showed relevant sensitizations to potassium dichromate, cobalt chloride, thiuram mix and carba mix. There was no statistically significant difference in the total contact sensitization rate between atopics and non-atopics, but contact sensitivity to nickel sulfate and palladium chloride was significantly higher among atopics. MOAHL index in % was: 44.1/14.2/24.9/ 54.4/2.4. In conclusion, we believe that similar large series of patients would be helpful to delineate the exact profile of contact dermatitis in Turkey.

Adolescent↗

Papulo-vesicular count for the rating of allergic patch test reactions. A simple technique based on polysulfide rubber replica.

Silicone rubber replica of patch test reactions were obtained from 12 patients with allergic contact dermatitis. 85 positive or doubtful reactions to allergens included in a standard battery were studied. The number of papulo-vesicles and sweat pores as seen in the replica were counted under a stereomicroscope. The papulo-vesicular count increased significantly in relation to clinical reading of the reactions of increasing strength. The number of sweat pores of positive reactions was decreased. This was probably due to cutaneous oedema with compression of sweat gland ducts. Papulo-vesicles are also consequences of the inflammatory oedema. The replica method was concluded to be a useful tool for the grading of allergic patch test reactions. It has the virtues of reproducibility, simplicity and low cost, and it can be evaluated under standardized laboratory conditions. Replica may also be the basis for more sophisticated methods of quantitation such as computer-aided image analysis.

Adult↗

Routine patch testing with paraben esters.

Paraben esters are the most widely used preservatives in cosmetics and topical medicaments. Their sensitization potential is low, based on both experimental and human experience. A paraben mixture is included in the ICDRG standard series, and in patch test studies, approximately 1% of eczema patients react to it. The present study confirms this frequency in 8020 patients patch tested consecutively. Testing with the individual paraben esters was employed as confirmation, which makes it unlikely that the excited skin syndrome is a significant problem in this context. It remains undetermined whether the present paraben mixture is the optimal patch test material for diagnosing paraben sensitivity.

Female↗