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Year : 2018  |  Volume : 15  |  Issue : 3  |  Page : 147-151

In vitro testing using immunocap provides reliable assessment of sensitizing patterns to local allergens in patients with rhinitis/asthma

Department of Allergy and Immunology, Apollo Gleneagles Hospital, Kolkata, West Bengal, India

Date of Web Publication10-Sep-2018

Correspondence Address:
Sujoy Khan
Department of Allergy and Immunology, Apollo Gleneagles Hospital, 58, Canal Circular Road, Kolkata - 700 054, West Bengal
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/am.am_18_18

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Background: Bronchial asthma affects over 300 million people worldwide and half of these patients have underlying allergic disease. Accurate identification of allergen triggers will help identify patients that may benefit from immunotherapy. Objective: The objective of this study was to understand whether in vitro testing provides accurate identification of suspected local allergens in our patients. Methods: A retrospective study was conducted on patients between 2015 and 2016 with persistent mild/moderate-severe rhinitis/asthma referred for aeroallergen sensitization tests. Total immunoglobulin E (IgE) and allergen-specific IgE levels were done using Phadia 100 (Thermo Fisher Scientific). Descriptive statistics were done using Microsoft Excel and nonparametric statistics using MedCalc software V18. Results: A total of 106 patient records (including 30 children) were analyzed. The median total IgE during 2015 (n = 38) was 402 kU/L (interquartile range [IQR] 25–75, 102–871 kU/L), while for 2016 (n = 68) was 509 kU/L (IQR 25–75, 131–1245 kU/L) (standard deviation comparison nonsignificant, P = 0.536). Thirty of 106 patients (28.3%) had no aeroallergen sensitizations. Allergen-specific IgE levels revealed that the highest sensitization rate was to house dust mites (both Dermatophagoides pteronyssinus and Dermatophagoides farinae), followed by cockroach and ragweed pollen (homologous to Parthenium hysterophorus). Median IgE level among 30 children was 511 kU/L (IQR 25–75 51.8–1688 kU/L). Almost 60% children were sensitized to house dust mites. Twelve of 30 children (40%) were polysensitized (i.e., specific IgE >0.35 kUA/L to ≥4 allergens) with 30% sensitized to P. hysterophorus. Mold sensitivity was 10% in both the years studied. Conclusions: ImmunoCAP method provides reliable assessment of sensitizations even for local allergens. House dust mite, cockroach, and Parthenium allergens appear to be important predictors of asthma morbidity not only among adults but also among children in Eastern India. Immunotherapy with house dust mites may prove beneficial in selected patients.

Keywords: Asthma, ImmunoCAP, immunoglobulin E, India, specific immunoglobulin E

How to cite this article:
Khan S. In vitro testing using immunocap provides reliable assessment of sensitizing patterns to local allergens in patients with rhinitis/asthma. Apollo Med 2018;15:147-51

How to cite this URL:
Khan S. In vitro testing using immunocap provides reliable assessment of sensitizing patterns to local allergens in patients with rhinitis/asthma. Apollo Med [serial online] 2018 [cited 2020 Jun 6];15:147-51. Available from: http://www.apollomedicine.org/text.asp?2018/15/3/147/240938

  Introduction Top

Asthma is a chronic respiratory disease that continues to affect millions of people of all ages worldwide and is now the 14th most important disorder in terms of global years lived with disability (Global Asthma Network, 2014).[1] The data provided by the 2014 report discusses ways to combat this avoidable disease in both children and adults with a focus on research and access to quality-assured essential medicines, with the latter still not available in the world's most poorest nations where the disease burden is disproportionately high.[2] Allergies are a potential factor in up to half of people with asthma, and therefore, knowledge of trigger factors that can exacerbate otherwise well-controlled asthma is very crucial in our understanding of the disease. Indoor allergens such as house dust mites have been shown to play an important role in the pathogenesis of bronchial asthma and other atopic diseases.

Mites and cockroaches can be found in almost all habitats but may differ greatly depending on the local conditions. That house dust mites (belonging to the family Pyroglyphidae) play a significant role in bronchial asthma is now well established.[3],[4],[5] Studies from asthmatic patients from one of the largest metropolis of Eastern India, Kolkata, showed that house dust mites belonging to the genus Dermatophagoides, including that of the storage mite Blomia tropicalis, were the major allergen triggering asthma in most patients.[6],[7],[8] However, the number of large reports from India still remains limited partly due to the reluctance of allergy testing or use of nonstandardized methods during testing, and therefore, do not add to the clinical knowledge or guideline development to combat the allergic disorders in the country.

The possible problem may be the element of apprehension among clinicians on the use of in vitro testing for allergies, especially when the allergens are derived from “nonnative” sources. However, it is important to understand that as the gold standard testing for allergy, the radio allegro solvent test is currently no longer available due to radioactivity concerns, and significant investment requirements one has to rely on alternative sensitive and specific in vitro testing methods.[9] Leading technologies such as the ImmunoCAP, Thermo Fisher Scientific, or the line blot assays (for example EUROIMMUN, Germany) use extremely purified allergens for the tests that have surpassed well above industry standards on in vitro diagnostic use requirements.[10] Allergy reports using such testing methods can be relied on provided the patient selection is appropriate, and the clinician has in-depth knowledge of the allergen and clinical manifestations, especially when multiple allergens are positive on the reports. The advantages over skin testing include (1) single-prick nonfasting blood sample; (2) no need to stop any medications; (3) can be tested even if eczema or asthma is uncontrolled; and (4) no risk to the patient. ImmunoCAP technology uses allergens that are attached to cellulose polymer, which is elastic, porous, and has a large surface area and is therefore capable of binding more than 150 times than traditional tube coating that only detects very low values of specific immunoglobulin E (IgE), giving accurate and precise detection of specific IgE from 0.10 to 100 kUA/L.[10] It measures the concentration of specific IgE, which is an indicator of allergic sensitization. It is, therefore, up to the clinician to verify whether the patient has any features of allergic disease before embarking on treatment plans and/or allergen avoidance strategies.

Accurate identification of allergen triggers for asthma will help identify patients that are candidates for immunotherapy (subcutaneous or sublingual). It is now well established that inaccurate identification or understanding of allergen triggers may lead to incorrect allergen mixes in immunotherapy protocols that would not be beneficial for the patient and may lead to Type I or III hypersensitivity reactions leading to unnecessary medical interventions.

This retrospective study was aimed at understanding the aeroallergen sensitization patterns in patients with underlying rhinitis/asthma who were referred to the Allergy and Immunology Department in a large tertiary care private hospital of Kolkata. This study would also enable to understand whether testing protocols using region-specific pollens/dust mite allergens are sufficient or a larger number of allergens are required for testing in our patients.

  Methods Top

A retrospective study was conducted on patients with persistent allergic rhinitis and persistent mild/moderate-to-severe asthma (united airways disease) on continuous treatment for at least the last year where allergen-specific IgE levels were measured. Patients with eczema, chronic urticaria, parasitic infections, and recurrent sinonasal or pulmonary infections were excluded from the study. Children were classified as asthmatic based on whether the parent reported at least one of the following symptoms for the child: (1) wheeze or (2) nocturnal cough or (3) exercise-induced wheeze, including (4) use of inhaled or oral corticosteroids over the last year.

Total IgE and allergen-specific IgE levels were measured by ImmunoCAP (Phadia 100, Thermo Fisher Scientific, and Sweden) method that uses a fluoroenzyme immunoassay platform, following manufacturer's instructions. Region-specific aeroallergens were used as a panel that included: Dermatophagoides pteronyssinus (d1 ImmunoCAP); Dermatophagoides farinae (d2 ImmunoCAP); American cockroach (i206 ImmunoCAP); grass pollen mix (gx2 ImmunoCAP) consisted of Cynodon dactylon, Lolium perenne, Phleum pratense, Poa pratensis, Sorghum halepense, and Paspalum notatum; tree pollen mix (tx3 ImmunoCAP) consisted of Juniperus sabinoides, Quercus alba, Ulmus americana, Populus deltoides, and Prosopis juliflora; food mix 1 (fx1 ImmunoCAP) consisted of peanut, hazelnut, Brazil nut, almond, and coconut; mold mix (mx1 ImmunoCAP) consisted of Penicillium chrysogenum, Cladosporium herbarum, Aspergillus fumigatus, and Alternaria alternata; animal dander mix (ex1 ImmunoCAP) consisted of cat, dog, cow, and horse dander; and weed mix (wx1 ImmunoCAP) consists of Ambrosia elatior, Artemisia vulgaris, Plantago lanceolata, Chenopodium album, and Salsola kali, following manufacturer's instructions.

The range of measurement of IgE level using Phadia was 2–5000 kU/L, with >5000 kU/L truncated at 5000 during statistical analyses. Sensitization level for specific IgE was considered as ≥0.35 kUA/L for all allergens.

Statistical analyses

Measures of descriptive statistics were used for initial analysis (Microsoft Excel 2007). Nonparametric statistical data were calculated and graphical images generated using the MedCalc Statistical Software version 18 (MedCalc Software bvba, Ostend, Belgium; http://www.medcalc.org; 2018).

  Results Top

Of 1084 samples analyzed between 2015 and 2016, 106 patients (57 males and 49 females) with allergic rhinitis and asthma (united airways disease) were studied over the 2 years that included 30 children (age ≤12 years) who had total IgE and allergen-specific IgE levels measured. The mean age of all patients was 25.6 years (range 2–68 years), with children aged between 2 and 12 years (mean age 6.93 years). The total IgE level showed an increasing trend across most seasons for the 2nd year (2016) among the patients studied as compared to the 1st year [Figure 1]. As the data for total IgE were not normally distributed, the median levels with interquartile ranges (IQRs) were used in the analyses. The median IgE level among all patients was higher in 2016 (n = 68) at 509 kU/L (IQR 25–75, 131–1245 kU/L) than in 2015 (n = 38) at 402 kU/L (IQR 25–75, 102–871 kU/L), as shown in the box-and-whisker plot comparison data graph [Figure 2]. Comparison of standard deviations between total IgE levels of 2015 and 2016, however, was not statistically significant (F-statistic 1.1860, P = 0.536). There was also no statistically significant difference between total IgE level and gender.
Figure 1: Median immunoglobulin E level (breakdown) across 4 seasons (2015–2016). Median total immunoglobulin E level across 4 seasons between the years 2015 and 2016 show an overall high immunoglobulin E level among all patient age groups with united airways disease

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Figure 2: Box-and-whisker plot comparing data of total immunoglobulin E for 2015 and 2016

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Aeroallergen sensitivity profiles among patients showed that maximum patients were sensitized to D. pteronyssinus and D. farinae with almost 60% of patients (adults and children) being positive to both species of house dust mites (dual positivity). Cockroach allergen was the second highest aeroallergen for both the years. Thirty of 106 patients (28.3%) had no sensitization to any of the aeroallergens (i.e., nonallergic asthma), with range of total IgE 5.93–1463 kU/L and median IgE level at 86.35 kU/L (25–75 IQR 39.5–160). [Table 1] lists the allergen sensitization profiles (median levels) with percentage positivity by year. Almost 26.3% and 30.9% of patients were sensitized to w1 ImmunoCAP (common ragweed that shows cross sensitivity to Parthenium hysterophorus) during 2015 and 2016, respectively. [Figure 3] shows the percentage sensitization rates of allergens in patients for the year 2016, with 48% of the pie chart taken up by the three major sensitizing allergens, namely, D. pteronyssinus, D. farinae, and cockroach.
Table 1: Comparison of median total IgE and specific IgE values among 106 patients with united airways disease between 2015 and 2016

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Figure 3: Pie chart showing the % sensitization rate of allergens for 2016 (allergen codes in text)

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Of the 30 children tested, total IgE ranged from 16.5 to >5000 kU/L (median 511 with IQR 25–75 51.9–1688 kU/L) and 60% of children were predominantly sensitized to the same house dust mites as adults. [Table 2] shows the breakdown of allergen sensitization with grass and tree pollen sensitization (seasonal allergens) that is on a similar scale as cockroach (perennial allergen). Between 2015 and 2016, 12 of 30 children (40%) were found to be polysensitized (i.e., specific IgE >0.35 kUA/L to ≥4 allergens) with 30% (9 of 30) sensitized to common ragweed (similar to P. hysterophorus) and 7% (2 of 30) were sensitized to mold mix. Of 10 children in 2016 whose total IgE levels ranged from 37.5 to 2628 kU/L, 20% of children were polysensitized with 60% children sensitized to dust mites. Mold sensitivity was 10% in both the years studied.
Table 2: Median total IgE and specific IgE levels among children ≤12 years age

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  Discussion Top

Our study shows that ImmunoCAP can reliably detect aeroallergen sensitizations in the majority of patients with suspected allergic asthma. About 60% of patients were sensitized to house dust mite, and almost 50% of total sensitizations were a combination of house dust mite and cockroach allergens. About 80% of children in our study had asthma diagnosed below the age of 10 years with 1 in 5 children polysensitized (aeroallergens implicated were house dust mites, cockroach, weed pollens, and tree and grass pollens). Skin testing to B. tropicalis in some of the patients was also positive (data not shown), which emphasizes that adding in vitro testing for B. tropicalis may be helpful in some settings.[11],[12] In the study sample overall, sensitizations to dust mite, cockroach, and Parthenium appeared to be important aeroallergen or environmental risks in our patients with moderate-to-severe asthma thus reinforcing the importance of controlling indoor exposure to these allergens. This is particularly relevant to children when early sensitization to these perennial allergens, coupled with excessive atmospheric pollutants and a family history of atopy puts them at a quite high risk of developing asthma below the age of 10 years.

The total IgE level did not show any correlation to either age, gender, or the presence of polysensitization, thus confirming the presence of additional factors such as atopy, interleukin 5 polymorphisms, socioeconomic, exposure to passive smoke, or environmental factors like pollutants that determine the IgE level in an individual. Skin testing in patients also does not confirm any correlation to wheal and flare response to the histamine control (i.e., mm histamine release) and underlying IgE level (personal experience) that has been partially examined in animal models.[13] It is, therefore, probably unnecessary to measure total IgE level in allergic patients when specific IgE levels are measured at the same time, and certainly, measuring only the IgE level without allergen-specific IgE has no value at all.[14]

In this study, we also noticed that ragweed sensitization (molecular homology to P. hysterophorus) appeared to be increasing across all age groups, and almost throughout the year thus becoming a perennial weed allergen of significance in India. A previous report from Northern India on the identification of common allergens in patients with united airway disease by skin prick test showed sensitization rates to house dust mite and Parthenium at 60% and 45%, respectively, with cockroach allergen at 18.75%.[11] The study also showed that aeroallergen sensitization rates in a North Indian city like Allahabad, Uttar Pradesh, is now comparable to increasing sensitization patterns throughout the country, which is also a disturbing trend all over the world. We had previously reported on increasing Type I hypersensitivity to weed pollens, especially P. hysterophorus (weed pollen) and showed that 25% of patients were sensitized[15] that was similar to the study by Verma et al. on seasonal variation in Type IV (contact) hypersensitivity to Parthenium where the mean titer of contact hypersensitivity correlated with increased sensitivity to Parthenium in summer.[16]

The region-specific pollens available on testing using ImmunoCAP appear to be sufficient in addressing the allergic concerns of patients with rhinitis/asthma. In the past 5 years, several patients have benefited from use of the highly standardized and locally relevant allergens for allergy testing and clinically relevant panel profiles have been created to address specific needs of clinicians. Thus, having a knowledge of seasonal pollens (grass, tree, or weed), perennial pollens (dust mite species, cockroach) or fungal allergens is critical to understand whether specific lung problems are due to Type I (allergies) or Type III/IgG-related responses. This not only prevents indiscriminate testing of several allergens but also helps decide on specific immunotherapy protocols.

The limitations to the study were as follows: (1) absence of data collection related to family history of atopy, socioeconomic status, exposure to passive smoke and other related demographics that may have provided with clues as to why IgE levels were high in some patients who did not have positive allergen-specific IgE; (2) single point data collection without follow-up to see improvement in allergen-specific IgE when management was altered based on allergen profiles; and (3) there was no correlation with FeNO (fraction exhaled nitric oxide) in children/pulmonary function tests (FEV1/FVC in adults) or peripheral eosinophilia with IgE/specific IgE level to identify asthma phenotypes.

  Conclusions Top

Sensitizations to dust mite, cockroach, and Parthenium allergens are important predictors of asthma morbidity not only among adults but also among children in Eastern India. Immunotherapy with house dust mite standardized extracts will therefore prove beneficial in these patients with allergic asthma.

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Conflicts of interest

There are no conflicts of interest.

  References Top

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