Allergy Diagnostic Testing
Posted: November 2007
Dr. John Oppenheimer
Director of Clinical Research,
Pulmonary and Allergy Associates
Denville, NJ, USA
Prof. Stephen Durham
Department Allergy and Respiratory Medicine,
Imperial College, London, UK
Dr. Harold Nelson
National Jewish Medical and Research Center
Denver, CO, USA
Credit for the first skin testing goes to Charles H. Blackley, who in 1865 abraded a quarter-inch area of his skin with a lancet, applied grass pollen on a piece of wet lint, and covered the scarified area with an occlusive bandage. This resulted in intense itching and a very large cutaneous response.
Percutaneous skin test ranks first in confirming the presence of IgE-mediated sensitization in the allergist's office. This should come as no surprise, as it has many advantages. Skin testing is minimally invasive, and when it is performed correctly it has good reproducibility, is easily quantified, and allows the evaluation of multiple allergens at one session. The results correlates with in vivo challenges. in vitro testing is an alternative, usually a back up tool for diagnosing allergic illness. Skin testing alone or in combination with in vitro testing is relied upon for the evaluation of allergic rhinitis, asthma, eczema, food allergy, insect sting allergy, drug allergy (especially beta-lactam and local anesthetic allergy), occupational disease and anaphylaxis. However, the reliability of these tests depends on a number of factors. In the case of skin testing, it is important that the technician performing the skin tests and the clinician ordering or interpreting these tests are aware of the advantages and pitfalls of the type of skin testing, the device used, the location of the tests on the body, the extracts used and the potential for suppression of the skin response by medications used to treat allergies or depression. These issues have been reviewed elsewhere in greater detail.1 For in vitro testing, it is imperative that quality standards be met. These include calibration of the assay, training and experience of the technician and the use of quality allergens in the solid phase.2 As in any diagnostic test, it is of paramount importance that the clinician consider the positive and negative predictive value of the tests performed. These tests should always be considered as adjuncts to the medical history and physical exam in formulating the diagnosis in each individual case, bearing in mind that both test types can yield false positive or, less commonly, false negative results.
Skin testing may be performed using either the prick/puncture (percutaneous) or intradermal (intracutaneous) technique. Intradermal testing is far more sensitive than prick/puncture testing, which means that it requires about 1000-fold less concentrated extracts than those used for prick/puncture testing to achieve a similar response. Although direct comparisons indicate that intradermal testing is more reproducible than percutaneous testing, there are many factors that favor the routine use of percutaneous allergy tests. These include economy of time, patient comfort and patient safety. Percutaneous testing allows the use of extract in 50% glycerin, which provides greater extract stability. Intradermal testing cannot use this diluent, as it may incite a false-positive irritant response. However, the most important consideration is that results of percutaneous testing correlate better with clinical allergy. The higher sensitivity of intradermal skin tests does not usually offer added benefit, since the results of skin prick tests performed with potent extracts are of sufficient sensitivity for use in clinical practice.
Two studies reinforce this concept.3,4 Each study compared intradermal with skin prick tests by correlating their results with patients' responses to natural exposure to allergen as well as by allergen challenge testing. In the first study, three groups of patients with seasonal rhinitis were compared. These subjects were classified into 3 groups based on their degree of sensitization to Timothy grass pollen. They were either skin prick test positive, only intradermal test positive, or were negative by both skin prick and intradermal testing. Both nasal allergen provocation testing and symptom scores during the pollen season correlated best with a positive skin prick test (>60% of subjects with positive skin prick tests had symptoms on allergen exposure). The frequency of positive nasal provocation (11%) and symptom scores (21%) in subjects with positive intradermal testing alone were not different from subjects who were skin prick test and intradermal test negative. The authors conclude that under the conditions of this study, the presence of a positive intradermal skin test response to Timothy grass in the presence of a negative skin prick test did not indicate the presence of clinically significant sensitivity to this grass.
In the second study, patients were challenged with cat exposure for one hour.4 Both positive skin prick tests and in vitro tests to cat were highly predictive of the development of symptoms upon allergen exposure in the cat challenge room.4 Subjects with a negative skin prick test were just as likely to have a positive challenge result if they had a negative intradermal skin test (31%) as subjects with a positive intradermal skin test (24%). The authors conclude that, at least with regard to cat allergy, major therapeutic decisions, such as environmental control or immunotherapy, should never be based on a positive intradermal skin test alone.
Both of these studies were performed in adults and both relied upon skin testing with relatively potent allergens (Timothy grass and cat). The clinical applicability of these results to less potent allergens, such as dog, or to younger patients (especially infants) is a matter of clinical judgment, because no specific evidence is available for these groups.
Whereas intradermal skin tests are always performed using a hypodermic syringe and needle, percutaneous tests may be performed with a variety of devices. Comparisons of percutaneous devices have been reviewed elsewhere in greater detail.5 Some devices have a single stylus with one or several points, whereas others have multiple heads and allow up to 10 tests to be accomplished with one application. The degree of skin trauma created by these devices for percutaneous testing varies and so may result in differences in the size of positive reactions, and the likelihood of producing a reaction at the site of the negative control. Thus, they require different criteria for what constitutes a positive reaction (see Table 1).
Skin test results are often reported by clinicians in semiquantitative terms. They may record results only as positive or negative, or express them on a 0 to 4+ scale without any indication of the size of the reactions that these numbers represent. However, allergy patients may have to change their allergist for numerous reasons, and it is important that records of prior allergy testing be interpretable by the receiving physician. At the very least, a record of skin testing should contain sufficient information to allow another physician to interpret the results and avoid the need to repeat skin testing. Standardized forms have been developed and are available through the American Academy of Allergy Asthma and Immunology website (for an example AAAAI's Skin Test and Immunotherapy Forms).
Although the area of the wheal and erythema are the most accurate measurements, the longest diameter or two diameters at right angles to each other correlate with area (r > 0.9).6 The importance of performing such measurements is exemplified by McCann and Ownby in which allergists were asked to interpret photographs of skin test reactions. The scoring and interpretation of the skin test results varied greatly.7 The authors of this study reinforce the idea that the most reliable method of reporting a skin test reaction is to measure and record the reaction size. At the very minimum, skin test results should be graded 0 to 4+, and the criteria for each grade of reaction clearly stated along with the skin test results.
Various investigators suggest different criteria for interpreting a skin test response as positive. To assess the reliability of different means of interpreting the results of skin prick testing, Vanto and colleagues studied a group of 202 children sensitive to dogs.8 A determination of sensitivity to dog was based on a composite score derived from the history, RAST, and bronchial or conjunctival allergen challenges. Although in this study the overall efficacy was greatest with the histamine reference method (in which the allergy skin test response is compared to a histamine control, with a positive response considered to be a response at least as great as that of the histamine control), maximal sensitivity was achieved when using a cutoff of a wheal 3 mm. If a clinician wishes to maximize sensitivity, the latter criterion would be most useful; however, adjustment must be made for the device used. Therefore, the criteria for a positive test should be the larger of: 1) 3 mm mean wheal diameter or 2) equal to or greater than the 99th percentile reaction with that device at negative control sites (see Table 1).
Like all other laboratory tests, it is imperative that quality assurance standards be met to ensure that the testing technique produces accurate results. To confirm such standards, it is recommended that all technicians performing skin testing undergo evaluation of their technique.9 Certainly, it would be comforting to know that skin test technicians achieve some degree of consistency in skin test performance. Although there are no formal standards available for skin test proficiency testing, several publications suggest some possible criteria. European publications suggest a coefficient of variation of less than 20% following repeated skin test control applications, and the Childhood Asthma Management Program study requires that a coefficient of variation of less than 30% be attained with repeated testing with histamine and consistently negative reactions to saline to confirm proficiency in skin testing.11
The National Committee for Clinical Laboratory Standards recommends quality control procedures for daily performance of in vitro allergy testing, with a recommended coefficient of variation of less than or equal to 15%.2 Even with such calibration and the increased use of automation, in vitro assays still have flaws. Williams and colleagues examined the performance of 6 large commercial laboratories on tests of blinded samples of the same sera, both diluted and non-diluted.10 They found that only two of the laboratories demonstrated acceptable precision and accuracy.
The preponderance of comparative studies demonstrate skin tests to be more sensitive than in vitro tests. However, the majority of these studies were performed with earlier generation in vitro tests. The newer in vitro tests produce higher test sensitivity and specificity12 by using a matrix capsule containing antigen bound to a hydrophilic carrier to produce enhanced specific IgE binding with lower nonspecific IgE binding.2 Levels of specific IgE measured by different commercial assays are not equivalent, as each assay differs in the composition of allergen reagents, methods of measurement and standardization procedures.
The advantages of in vitro testing are largely related to use in patients with extensive dermatoses (e.g., atopic dermatitis), resulting in an inability to perform tests on unaffected skin, or in patients who are unable to discontinue medicines that block the histamine response, i.e., antihistamines or tricyclic antidepressants. The disadvantages of in vitro testing include a potential decrease in sensitivity, added cost, and lack of immediate and visible response. Performing both in vitro and in vivo tests may yield improved sensitivity.13
Although there are challenge protocols available in the research setting to confirm allergic rhinitis and asthma, the standard tool available to the clinician is a careful history and physical exam. Skin testing correlates with results of nasal challenge or bronchial challenges, when allowance is made for nonspecific airway responsiveness.
When evaluating potential food allergy, the clinical history is the initial screening, with skin testing or in vitro tests used to corroborate the history. Oral food challenges represent the "gold standard" for the confirmation of food allergy. These can be performed as open challenges or in a single- or double-blind fashion. Food challenges are not without risk and thus require that appropriate supportive care be available. Several studies demonstrate that the magnitude of the in vitro test or the skin test reaction size may be useful in determining the utility of performing a food challenge.14,15 One additional advantage of skin testing for food allergies is the ability to perform skin testing with the fresh food, "prick-prick" test. Several reports demonstrate that fresh foods provide greater sensitivity for certain foods.16,17 This is particularly important in assessing allergy to fruit; however, useful results have also been demonstrated for other foods, including seafoods, peanut, tree nuts, vegetables, milk and eggs.
Conclusion Diagnostic testing remains an essential tool for the evaluation of the allergic patient. Several variables should be controlled to produce more reliable skin test results and improve the predictive values of allergy skin testing. It is also imperative that allergists ensure that the results of skin testing are reliable by conducting proficiency testing. In addition, the results must be properly documented to make them easily understandable by others. Similar standards must be applied to in vitro testing; as in the case of skin testing, it is imperative that the ordering physician be familiar with the operating characteristics that the in vitro lab employs.
- Dreborg S. ed. Skin tests used in type I allergy testing Position paper. Allergy 1989;44:s1-59.
- Yunginger, J. MD a; Ahlstedt, S; Eggleston, P. et al. Quantitative IgE antibody assays in allergic diseases JACI 2000;105:1077-84
- Nelson HS, Oppenheimer JJ, Buchmeier A, et al. An assessment of the role of intradermal skin testing in the diagnosis of clinically relevant allergy to timothy grass. J Allergy Clin Immunol 97:1193-1201, 1996.
- Wood RA, Phipatanakul W, Hamilton RG, Eggleston PA. A comparison of skin prick tests, intradermal skin tests, and RASTs in the diagnosis of cat allergy. J Allergy Clin Immunol 103:773-9, 1999.
- Oppenheimer J. Devices for epicutaneous skin testing. in Skin Testing Dolen W (ed): Immunology and Allergy Clinics of North America Philadelphia, WB Saunders 2001, p 263-72.
- Ownby DR. Computerized measurement of allergen-induced skin reactions. J Allergy Clin Immunol 69:536-8, 1982;
- McCann WA, Ownby, DR. The reproducibility of the allergy skin test scoring and interpretation by board-certified/board-eligible allergists. Ann All Asthma Immunol 2002;89:368-71
- Vanto T. Efficacy of different skin test methods in diagnosis of allergy to dogs. Ann All 1982:49:340-4.
- Turkeltaub P. Performance standards for allergen skin testing: An approach to proficiency testing in Skin Testing Dolen W (ed): Immunology and Allergy Clinics of North America Philadelphia, WB Saunders 2001, p321-8.
- Williams, PB ; Barnes, J; Szeinbach, S; Sullivan, T Analytic precision and accuracy of commercial immunoassays for specific IgE: Establishing a standard J All Clin Immunol 2000;105:1221-30
- Adkinson NF Jr. The radioallergosorbent test in 1981--limitations and refinements Jl All Clin Immunol 1981;67:87-9
- Oppenheimer J, Nelson HS. Skin Testing. Ann All Asthma Immunol. 2006;96:S6-12.
- Droste JH, Kerkhof M, de Monchy JGR et al. Association of skin test reactivity, specific IgE, total IgE, and eosinophils with nasal symptoms in a community-based population study J All Clin Immunol 1996;97:922-32
- Sampson H Update on food allergy Jl All Clin Immunol 2004;113: 805-819
- Sporik, R0, Hill DJ, Hosking, CS0 Specificity of allergen skin testing in predicting positive open food challenges to milk, egg and peanut in children. Clin and Exp All 2000;30:1540-6
- Ortolani C, Ispano M., Pastorello EA.,. Ansaloni R, Magri GC Comparison of results of skin prick tests (with fresh foods and commercial food extracts) and RAST in 100 patients with oral allergy syndrome Jl All Clin Immunol 1989;83:683-90
- Rosen JP, Selcow JE, Mendelson LM et al. Skin testing with natural foods in patients suspected of having food allergies: Is it a necessity? Jl All Clin Immunol 1994;93:1068.
Wheal size indicating a positive response to skin tests using various devices.a
|Devices for which a 3-mm
wheal would be significant
|Devices for which a wheal >3 mm
should be used as significant
|Device||99th Percentile of reactions at the negative control sites||Device||99th Percentile of reactions at the negative control sites|
|Quintest (HS) b puncture||0 mm||DuoTip (Lincoln) twist||3.5 mm|
|Smallpox needle (HS) prick||0 mm||Bifurcated needle (ALO) prick||4.0 mm|
|DuoTip (Lincoln) prick||1.5 mm||MultiTest ( Lincoln ) Puncture||4.0 mm|
|Lancet (HS) puncture||2.0 mm||Bifurcated needle (ALO) puncture||4.5 mm|
|Lancet (ALK) Puncture||3.0 mm||Quick Test (Pantrex)||4.0 mm|
|DermaPICK II (Biomedixs) Prick or puncture?||0 mm||Greer Track (Greer)||3.5 mm|
a Positive response is defined as a wheal greater than 99% of wheals generated by the administration of saline to the subject's back by the same operator. Adapted from ref. 12.
b HS = Hollister Steir, Greer = Greer laboratories, Lincoln = Lincoln Diagnostics, ALK = ALK America, ALO = Allergy Labs of Ohio