What should be considered during epicutaneous patch testing?

Indications for epicutaneous patch testing include the detection or exclusion of allergic contact dermatitis of the skin, transitional mucosa or oral mucosa, suspected delayed-type immune reactions to ingredients of implants, and certain drug reactions (maculopapular exanthema, symmetrical drug-related intertriginous and flexural exanthema [SDRIFE], acute generalized exanthematous pustulosis [AGEP], and fixed drug eruption). When available, allergen preparations that have been pharmaceutically tested and that are approved or marketable as medicinal products should be used. Existing diagnostic gaps can be closed by testing the patient’s own materials in a suitable preparation. Interferences of ultraviolet (UV) light exposure or drugs with patch test reactions have to be considered. In addition to the reading after 48 h and 72 h, a further reading between day 7 (168 h) and day 10 (240 h) is useful, since about 15% of the positive test reactions remain undetected without this late reading. All positive patch test reactions considered allergic must be evaluated with regard to their clinical relevance based on existing exposures.


Introduction
Allergic contact dermatitis is based on a delayed-type hypersensitivity reaction (synonym type IV hypersensitivity) and is mostly caused by low molecular weight substances and metal ions [1]. Sensitizations to these contact allergens are common in the general population with a prevalence of approximately 20% [2]. The estimated 1-year prevalence of allergic contact dermatitis is 7% [3]. Important allergen groups are metals (e.g., nickel, cobalt, chromium), fragrances, preservatives/biocides, rubber ingredients or resins/adhesives [4].
The diagnostic standard for the detection or exclusion of delayed-type hypersensitivity and, thus, allergic contact dermatitis is the epicutaneous patch test. In principle, this is a provocation test in which small skin areas are exposed to the selected substances epicutaneously, usually under occlusion (test chamber and test patch). In sensitized individuals, an eczema-review tous reaction then develops in the test area with a time delay, usually after 2 or more days.
The most important indication for patch testing is the detection or exclusion of allergic contact dermatitis of the skin, adjacent mucous membranes or oral mucosa. Further indications are the suspicion of a delayed-type immune reaction to ingredients of implants or certain drug reactions (e.g., maculopapular exanthema, symmetrical drug-related intertriginous and flexural exanthema [SDRIFE], acute generalized exanthematous pustulosis [AGEP], fixed drug eruption) [5]. So-called prophetic (predictive) tests in the absence of a history of allergic symptoms to predict possible future sensitization are not recommended [5].
In March 2019, the revised evidence-based guideline on epicutaneous patch testing, which has been methodologically upgraded to S3 level, was published [5,6]. It is available on the homepage of the AWMF (Arbeitsgemeinschaft der Wissenschaftlichen Medizinischen Fachgesellschaften e. V.; https://www.awmf. org/leitlinien/detail/ll/013-018.html). Most recently, the AWMF extended the validity period of this guideline until March 2024. We take this as an opportunity to address important aspects of the S3 guideline that are relevant to practice.

Preparation of the patch test
If the clinical picture is compatible with a delayedtype allergic reaction, a thorough history taking is essential to collect information on the complaints (e.g., location of skin lesions, temporal course), exposures (occupational and non-occupational), pre-existing (skin) diseases, and (concomitant) medications [7]. If this results in an indication for patch testing, the appropriate commercial test substances should be selected. In principle, the so-called standard or baseline series should be tested in every patient. This includes the most important contact allergens that are widely distributed and/or most frequently cause sensitizations. Based on the patient's history on possible relevant exposures, further test series are selected in which the test allergens are grouped thematically (e.g., occupation, allergen group). The German Contact Dermatitis Research Group (DKG) regularly updates the composition of the different test series in cooperation with the Information Network of Departments of Dermatology (IVDK; https://dkg.ivdk.org/ testreihen.html).
If available, pharmaceutically tested allergen preparations that are approved or marketable as medicinal products should be used. The quality of these test substances should be ensured by appropriate measures (e.g., refrigeration, protection from light, storage in closed containers, observance of the expiration date). Depending on the patient's history, however, patient's own materials (e.g., cosmetics, occupational substances) should also be considered for inclusion  in the test. This is necessary because commercial test substances are not available for all allergens in question and only a few new commercial test preparations are being developed. Diagnostic gaps can therefore be closed by testing the patient's own materials in a suitable preparation. In this context, it should be noted that physicians testing patient's own materials are required to notify once the respective competent state authority (GCP inspectorate) according to Sect. 67 (1) of the German Medicinal Product Act (AMG) [8]. A sample notification letter can be found on the DKG homepage (https://dkg.ivdk.org/ leitlinien.html). Special care and expertise are required when preparing patient's own materials for testing. For example, adequate dilution, selection of a suitable test vehicle, or control of pH may be required [9]. For this purpose, assistance and instructions can be found in the literature [10][11][12] and on the DKG homepage ("Arbeitsanweisung Epikutantest mit patienteneigenem Material"; https://dkg.ivdk.org/ leitlinien.html; last accessed: 14 September 2022). Testing with substances or substance mixtures of unknown chemical identity or unknown biological effect or unknown, possibly too high concentration should be rejected accordingly [5]. Regarding the importance and performance of patch testing in the diagnostic work-up of adverse cutaneous drug reactions, reference should be made to a recent review by Barbaud et al. [13].
Prior to testing, each patient must be informed (preferably in writing) about the purpose, procedure, and risks and possible side effects of patch testing (Table 1). In principle, the possible risks and side effects are the same when testing commercial test substances or the patient's own substances. A DKG recommendation for written information and informed consent can be found on the DKG homepage (https:// dkg.ivdk.org/leitlinien.html). Patch testing should not be performed or should be postponed in the case of acute or severe inflammatory skin lesions, skin damage at the test site or pregnancy/breastfeeding [5,6].

Execution of patch testing
Patch testing is usually performed on the back. Testing on the upper arms or thighs may be considered. However, it has been shown that test reactivity de-78 What should be considered during epicutaneous patch testing?  [14]. Testing should ideally be performed only after resolution of acute dermatitis lesions; otherwise the occurrence of an "angry-back" phenomenon or flare-up reactions is to be expected.
In view of the higher sensitivity (fewer false-negative test reactions), it is recommended to leave the test chambers on the skin for 48 h [15]. In individual cases, exposure for only 24 h may be considered (e.g., when testing particularly irritant substances to prevent false-positive test reactions). The first reading is usually done 15-30 min after patch removal, i.e., usually on day 2 (in case of a 48 h-occlusion period) and on day 1 (in case of a 24 h-occlusion period). A further reading on day 3 (72 h) or day 4 (96 h) is mandatory. If such a reading is missing, the test is not viable. If a reading is performed only after 48 h, there is a risk that the patch test will be false-negative, as subsequent positive reactions are not recorded, and that irritant reactions (positive test reaction after 48 h, but negative after 72 h) will be incorrectly interpreted as allergic. An additional reading between day 7 (168 h) and day 10 (240 h) is useful, as some reactions appear very late. Approximately 15% of positive test reactions are missed if such a late reading is not done [16]. Late reactions are particularly common with the following allergens: corticosteroids, antibiotics (neomycin), formaldehyde, formaldehyde releasers, formaldehyde resins, p-phenylenediamine, (dental) metals [5,6]. Therefore, late reading is recom- Fig. 1 Positive patch reactions to different test substances, each read after 72 h. a Weak positive reactions "+", erythema, infiltration, papules, but no vesicles; b strong positive reaction "++", erythema, infiltration, papules, and vesicles; c extreme positive reaction "+++", erythema, infiltration, and confluent vesicles mended for these allergens. However, a recent study by the IVDK shows that late reactions are also frequent with other allergens, including mercaptobenzothiazole, butyhydroxytoluene, chlorhexidine, chloracetamide, and sorbic acid [17].

Reading of the patch test
For the classification of the test reactions, reference is made to the DKG reading criteria (Table 2; [5]). They are based on the reading criteria of the International Contact Dermatitis Research Group (ICDRG) [18]. The skin reaction is classified based on morphological criteria. Positive test reactions are divided into three grades (+, ++, +++) according to their strength (Fig. 1). Especially the differentiation of (non-allergic) irritant, questionable and only weakly positive (allergic) reactions can be challenging. Therefore, a reading training is offered free of charge on the DKG homepage to familiarize oneself with the different grades (https://ablesetraining.ivdk.org/).

Interpretation of test reactions
During the last reading, the test reactions are interpreted with respect to their etiopathogenesis (allergic or irritant). A negative patch test result usually excludes sensitization to the tested substance if the test was performed properly. For individual allergens, reactions morphologically classified as positive may also represent irritant reactions; this is especially true for K What should be considered during epicutaneous patch testing? 79 review Table 3 Problem allergens for which false-positive test reactions must be considered. Modified according to [19] Allergen Test concentration (%)

Preservatives and disinfectants
Benzalkonium chloride 0. weakly positive test reactions. A rapid decrescendo of a positive reaction is suggestive of an irritant origin, whereas an increasing reaction (crescendo) is more typical of an allergic reaction. Particular care must be taken in the interpretation of test reactions to socalled problem allergens (Table 3), as these have an inherent potential for skin irritation [19]. Accordingly, in patch testing of problem allergens, questionable or irritant reactions occur more frequently than positive reactions (low reaction index) or the proportion of weakly positive reactions (+) outweighs all positive reactions (high positivity rate) [6]. Therefore, falsepositive test reactions must be considered when testing problem allergens. It is also helpful to additionally test the obligatory irritant sodium lauryl sulfate (SLS; 0.25% aq.), which is therefore also part of the DKG standard series [20]. If erythema occurs in the test field where sodium lauryl sulfate is tested, this indicates nonspecific increased skin irritability at the time of testing at the test site. This should lead to critically questioning weak positive test reactions, especially to problem allergens, and rather to evaluate them as suspected false-positives. A confirmation test (e.g., repeated open application test [ROAT]) may help to clarify the situation further in individual cases [21]. The irritant reaction to sodium lauryl sulfate should not be entered in the allergy ID card, as this can then be misinterpreted as a sensitization.

Relevance assessment of patch test reactions
All positive patch test reactions judged to be allergic should be evaluated for clinical relevance. A distinction should be made between current and previous relevance. Clinical relevance exists when the identified sensitization "explains" current or previous episodes of allergic contact dermatitis due to exposure to that particular allergen. Identification of such exposures often requires a targeted additional history taking at the conclusion of patch testing. It is not uncommon to discover sensitizations unrelated to the current symptoms that led to testing. In addition, sensitizations are also frequently detected that are not associated with any clear current or previous relevance even after a detailed history taking (unclear relevance). An allergy ID card should be issued for allergens that have elicited an allergic reaction in the patch test. The patients should be informed in detail about the occurrence of the allergens and possibilities of avoidance. This represents the basis for a sustainable treatment success.

Interference of UV exposure or drugs with patch test reactions
In case of intensive ultraviolet (UV) light exposure of the entire body surface and/or the test site (leisure time, occupation, UV therapy) within the last 4 weeks, testing should be postponed, since UV-induced reduction of allergen-presenting dendritic cells in the epidermis [22] may impair the test reactivity.
Certain drugs may interfere with the patch test reaction. This is particularly true for drugs that have an impact on the immune system. Therefore, if possible, glucocorticoid doses of ≥ 20 mg/day prednisolone equivalent and topical glucocorticoids in the test area should be discontinued one week prior to patch testing. Execution of patch testing can be considered if administered glucocorticoid doses are < 20 mg/day prednisolone equivalent. If discontinuation of immunosuppressive drugs is not possible, patch testing can still be performed. However, it should then be noted that false-negative results may occur. Since second-generation H1 antihistamines may also lead to an attenuation of the test reactivity, these should also be discontinued, if possible, at a sufficient interval before the start of the patch test. Empirically, a latency of five half-lives (depending on the active substance, approximately 2-5 days) can be considered sufficient. The half-lives of commonly used antihistamines are listed 80 What should be considered during epicutaneous patch testing? K review in the AWMF guideline for performing patch testing [6]. Azathioprine does not affect patch test reactivity; performing patch testing can be recommended despite taking azathioprine [6]. Also, a recent systematic review indicates that a large number of positive patch test reactions are reproducible when retested under treatment with various immunosuppressive or immunomodulatory agents (e.g., dupilumab, tumor necrosis factor [TNF]-α blockers). This suggests that it may be useful to perform a patch test even under such therapies [23].

Special patient groups
For children over 12 years of age, the same recommendations apply for patch testing as for adults. For children under 12 years of age, testing of the adapted "DKG standard series for children" (currently 12 test substances; https://dkg.ivdk.org/testreihen.html) is recommended. In addition, it can be considered to shorten the test exposure duration to 24 h (1 day) [6]. Children under 6 years of age should be tested with single, selected contact allergens only in exceptional cases of clear clinical suspicion of contact allergy.
For general safety reasons, patch testing should not be performed in pregnant or breastfeeding women. Patch testing can also be performed in cases of nondrug-induced immunosuppression (e.g., HIV infection). However, the individual immune status must be taken into account when interpreting the results [6].

Modified testing
In the case of patch test reactions not interpreted as positive or considered to be allergic but persistent suspicion of contact allergy, and in the case of test substances with poor penetration of the corneal layer or reduced penetration capacity at the test site, a strip patch test may be useful [24]. In this case, the uppermost skin layers in the test area are removed by tearing off adhesive tape before application of the patch test [6,21]. Commercial and patient-specific test substances are suitable for this purpose, provided that they are not expected to cause an irritant reaction under the conditions of the regular patch test. Other variants of patch testing are the photopatch test, the open and semi-open patch test and the repetitive open application test (ROAT), which will not be discussed separately here [21]. In addition, reference is made to the recommendations published by the DKG on the performance of the photopatch test [25].
Funding Open Access funding enabled and organized by Projekt DEAL.
Conflict of interest R. Brans and V. Mahler declare that they have no competing interests. The following applies to V. Mahler: The contents and positions expressed in this minireview reflect the personal expert opinion of the author and must not be interpreted or quoted as if they had been commissioned by or reflected the position of the competent national higher federal authority, the European Medicines Agency, or one of their committees or working groups.
Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.