







































 

_____________________________________________________________________________________________________ 
 
*Corresponding author: E-mail: celso@alergoimuno.med.br; 
 
Cite as: Olivier, Celso Eduardo, Daiana Guedes Pinto, Ana Paula Monezzi Teixeira, Jhéssica Letícia Santos Santana, Raquel 
Acácia Pereira Gonçalves Santos, and Regiane Patussi Santos Lima. 2024. “Endotyping Non–IgE-Mediated Immunoreactivity 
to Polyethylene Glycol: Implications for Allergic Patients”. Asian Journal of Immunology 7 (1):100-111. 
https://journalaji.com/index.php/AJI/article/view/135. 

 
 

Asian Journal of Immunology 
 
Volume 7, Issue 1, Page 100-111, 2024; Article no.AJI.119914 
 

 
 

 

 

Endotyping Non–IgE-mediated 
Immunoreactivity to Polyethylene 

Glycol: Implications for Allergic 
Patients 

 
Celso Eduardo Olivier a*, Daiana Guedes Pinto a,  

Ana Paula Monezzi Teixeira a,  

Jhéssica Letícia Santos Santana a, 

Raquel Acácia Pereira Gonçalves Santos a  

and Regiane Patussi Santos Lima b 
 

a Instituto Alergoimuno de Americana, Brazil. 
b Lavoisier’s Laboratories, São Paulo, Brazil. 

 
Authors’ contributions  

 
This work was carried out in collaboration among all authors. Author CEO designed the study, wrote 

the protocol and the manuscript’s first draft, and managed the literature searches. Authors DGP, 
APMT, JLSS and RPSL performed laboratory research. Author RAPGS performed cutaneous  

tests. All authors read and approved the final manuscript. 
 

Article Information 
 

DOI: https://doi.org/10.9734/aji/2024/v7i1135  

 

Open Peer Review History: 

This journal follows the Advanced Open Peer Review policy. Identity of the Reviewers, Editor(s) and additional Reviewers,  
peer review comments, different versions of the manuscript, comments of the editors, etc are available here: 

https://www.sdiarticle5.com/review-history/119914 

 

 

 

Received: 01/07/2024 

Accepted: 16/07/2024 

Published: 17/07/2024 

 
 

Original Research Article 

https://doi.org/10.9734/aji/2024/v7i1135
https://www.sdiarticle5.com/review-history/119914


 
 
 
 

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ABSTRACT 
 

Background: Several publications report polyethylene glycols (PEGs) and PEGylated therapeutics 
as responsible for non–IgE-mediated allergic reactions. There is no standardized lab exam that can 
endotype (quantify the participation of these mechanisms inside each patient’s pattern of 
symptoms) besides in vivo provocation tests. 
Objective and Aim: To evaluate the Tube Titration of Precipitins (TTP) and the Leukocyte 
Adherence Inhibition Test (LAIT) to discriminate and endotype non–IgE-mediated immunoreactivity 
against a PEGs solution in patients with non–IgE-mediated allergic phenotypes. 
Research Design and Research Protocol: To retrospectively examine the medical charts of two 
cohorts (with 100 patients each) diagnosed with non–IgE-mediated allergic rhinitis, allergic 
bronchitis, asthma, allergic sinus headache, atopic dermatitis, and/or urticaria who were 
investigated with the help of TTP or ex vivo challenge tests monitored by LAIT against PEGs.  
Methodology: We identified the registered results of the semi-quantitative serum TTP against 1 
mg/mL PEGs 4000 solution, which were distributed in ranges through a cascade distribution chart 
to outline the variability of the results inside the first cohort. The LAI done with the ex vivo 
challenges with 1 mg/mL PEGs 4000 solution were distributed in ranges through a cascade 
distribution chart The statistical characteristics of the two cohorts were calculated.  
Results: The TTP showed a wide distribution range of results. The mean was estimated at 1:246; 
the median at 1:192; the standard deviation at 1:188. The LAIT showed a wide distribution range of 
results. The LAI ranged from 0% to 98%. The mean was 41.9%; the median was 45%; the standard 
deviation was 28.5%. The cascade distribution demonstrates a wide range of LAI results. 
Conclusion: Some patients showed any or low immunoreactivity during the ex vivo challenge test, 
while most displayed moderate or strong immunoreactivity, which could reflect the participation of 
PEGs antibodies in a non–IgE-mediated hypersensitivity condition. 

 

 
Keywords: Allergy; asthma; atopic dermatitis; bronchitis; COVID-19 vaccination; diagnosis; 

exposome-wide association study; endotype; hypersensitivity; leukocyte adherence 
inhibition test; macrogol; Non–IgE-mediated immunoreactivity; precision medicine; 
polyethylene glycol; E1521; rhinitis; sinus headache; urticaria. 

 

ABBREVIATIONS 
 

LAI : Leukocyte Adherence Inhibition 
LAIT : Leukocyte Adherence Inhibition Test 
TTP : Tube Titration of Precipitins 
PEGs : Polyethylene Glycol Polymers 
 

1. INTRODUCTION  
 
For a long time, petroleum-derived polymeric 
products were discarded as tars. The first 
polymerized structures to be identified from tars 
were the polyethylene glycol polymers (PEGs) 
after continued condensation studies in 1860 [1]. 
PEGs are polymers of parental ether monomers 
such as ethylene glycol, ethylene oxide, or 
oxyethylene, commonly available as compounds 
together with their anionic or nonionic 
derivatives, in mixtures of different chain lengths 
polymers, indicated by their average molecular 
weight (200 to over 10,000 Da) [2]. 
 
PEGs are used as solvents and emulsifying 
agents in processed food, cosmetics, medicines, 
cleansing, and personal care products. They 

may also be involved in the production of fabrics, 
plastics, resins, papers, ceramics, glasses, 
rubber, metals, wood preservatives, et cetera [3]. 
 
“Chemically designated as alpha-hydro-omega-
hydroxypoly(oxy-1,2-ethanediol) or polyethylene 
oxides, PEGs are regulated additives, allowed to 
be used as emulsifiers in industrialized food and 
food supplements (E 1521) and as plasticizers in 
film coating formulations for food supplement 
tablets and capsules” [4]. “PEGs are 
amphipathic substances soluble in water and in 
many organic solvents, including aliphatic 
ketones, alcohols, chloroform, glycol ethers, 
esters, and aromatic hydrocarbons. Acting as 
emulsifiers, they prevent the separation of 
unstable water/oil emulsions in industrialized 
food such as mayonnaise, dairy-based ice 
cream, frozen yogurts, margarine, blended 
spreads, cakes, pastries, and frozen desserts” 
[5]. When used as dietary emulsifiers, PEGs 
may increase intestinal permeability by 
interfering with the physical properties of the 
mucus layer, facilitating the growth of              
pro-inflammatory intestinal microbiota and 



 
 
 
 

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contributing to the impairment of inflammatory 
intestinal diseases and food allergies [6,7]. 
 
PEGs (also designated as macrogol in 
pharmacy) are used as vehicles for drugs and 
ointment bases, capsules, tablet and pill binders, 
suppositories, and liquid prescriptions, including 
parenteral, topical, ophthalmic, oral, and rectal 
preparations. They also prevent protein 
aggregation and inactivation of pharmaceutical 
proteins encapsulated in bioerodible polyester 
microspheres, acting as stabilizers [8]. 
PEGylation (or pegylation) is a process 
developed in the 1970s to covalent or non-
covalently attach PEG polymers to molecules, 
such as drugs, therapeutic proteins, peptides, 
antibody fragments, and vesicles, then described 
as “PEGylated” [9]. “PEGylation changes the 
physical and chemical properties of the 
biomedical molecule, such as its conformation, 
electrostatic binding, and hydrophobicity, 
resulting in an improvement of the 
pharmacokinetic behavior of the drug, reducing 
coalescence, degradation and in vivo elimination 
of the active pharmacological principles” [10-12]. 
“PEGylated products can induce anti-PEG 
antibody production associated with type III 
complement-activated immunoreactivity” [13]. 
 
“Lower molecular weight PEGs are absorbed by 
the digestive tract and excreted in the urine; 
however, high molecular weight PEGs are not 
absorbed and form hydrogen bonds with water in 
the gastrointestinal tract, where they hydrate 
stools” [14]. Long-chain PEGs are laxative 
medications in isosmotic balanced electrolyte 
preparations [15]. PEGs are also used in topical 
dermatological preparations as penetration 
enhancers [16]. PEGs and their derivatives 
(laureth, ceteth, ceteareth, steareth, oleth, 
laurate, dilaurate, stearate, distearate) do not 
readily penetrate intact skin and are added to a 
great variety of cosmetic applications as 
surfactants, cleansing agents, emulsifiers, skin 
conditioners, and humectants [17]. However, 
absorption through damaged skin of PEGs 
present in burn creams was associated with fatal 
intoxication in burned patients characterized by 
acute renal failure and metabolic acidosis with 
increased calcium, anion, and osmolar gap [18]. 
Allergic reactions due to PEGs used as 
excipients have already been reported [19]. 
 
Pharmaceutical excipients are a hidden potential 
cause of drug and vaccine hypersensitivity 
reactions [20]. Allergic reactions to PEGs were 
put into evidence, particularly after Pfizer-

BioNTech® and Moderna® mRNA COVID-19 
vaccines were approved for mass vaccination 
[21,22]. 
 
Endotyping pharmaceutical excipient 
immunoreactivity has become a herculean task 
due to the variety and diversity of formulations, 
mainly after the appearance of the PEGylated 
lipid nanoparticle vaccines [23]. PEGs are the 
most popular nanomedicine components due to 
their chemical properties [24]. 
 
The IgE-mediated anaphylaxis elicited by PEGs 
excipients was the first and easiest endotype to 
diagnose [25,26]. Skin and basophil activation 
tests are the most available assays to identify 
immediate and/or IgE-mediate allergy to PEGs 
[27]. However, intradermal skin tests performed 
with PEGs may produce immediate anaphylactic 
reactions, which makes the percutaneous route 
a more secure way to perform in vivo challenges 
[28]. At our facilities, we had performed 
hundreds of skin tests with PEGs employing the 
skin scrape test technique with no systemic 
reaction, despite dozens of positive wheal-and-
flare local diagnostic reactions elicited by PEGs 
[29]. A Dual Cytometric Bead Assay (DCBA) was 
recently described for detecting anti-PEG IgG, 
IgM, and IgE in patient sera [30]. 
 
Besides the type I IgE-mediated hypersensitivity 
reaction, several other mechanisms have been 
proposed, such as IgG-mediated anaphylaxis 
(Complement-related or not) and direct mast cell 
activation [31]. The production of Anti-Poly 
(Ethylene Glycol) antibodies, besides provoking 
hypersensitivity reactions, can also be 
problematic when a PEGylated drug is 
administered, accelerating drug clearance and 
decreasing its therapeutic efficacy [32,33]. 
COVID-19 mRNA vaccination is associated with 
elevated levels of vaccine-induced anti-PEGs 
antibodies and increased systemic 
reactogenicity [34]. PEGs excipients are also 
suspected to elicit life-threatening immune-
induced organ-specific reactions such as acute 
pancreatitis, acute interstitial nephritis, and liver 
injury [35]. 
 

We routinely employ the Leukocyte Adherence 
Inhibition Test (LAIT) and the Tube Titration of 
Precipitins (TTP) in our facilities as a triage to 
evaluate non–IgE-mediated immunoreactivity 
against suspected allergens before the 
performance of more exhaustive in vivo 
provocation tests [36-42]. The present study 
hypothesizes that the LAIT and the TTP may 



 
 
 
 

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differentiate diverse endotypes and degrees of 
immunoreactivity against PEGs among patients 
suffering from the most common allergic 
phenotypes. To demonstrate diversity in immune 
responses and to evaluate the potential of the 
LAIT and the research of precipitins to 
endotyping non–IgE-mediated immunoreactivity 
against PEGs, we retrospectively compiled the 
electronic medical charts of patients with non–
IgE-mediated allergic rhinitis, allergic bronchitis, 
asthma, sinus headache, atopic dermatitis, 
and/or urticaria who were investigated with these 
procedures into our institution.  
 

2. MATERIALS AND METHODS  
 

2.1 Subjects 
 
After receiving Institutional Review Board 
approval from the Instituto Alergoimuno de 
Americana (Brazil; 05/2024), we reviewed the 
electronic chart of 9.000 outpatients who 
attended our facility from January 2018 to June 
2024. 
 
The sample selection was performed                    
by retrieving the results from the electronic 
sheets. 
 
A cohort of 100 outside patients had been 
submitted to TTP with PEGs solution 1mg/mL for 
presenting non–IgE-mediated allergic rhinitis, 
allergic bronchitis, asthma, allergic sinus 
headache, atopic dermatitis, and/or urticaria. 
This cohort counted 31 males; mean age 34.8 
years; standard deviation (SD) 21.3 years; range 
0 to 80 years; median 32.5 years; mode = 28 
years (appeared five times). 
 
A different cohort of 100 outside patients had 
been submitted to an ex vivo allergen challenge 
test with PEGs solution 1mg/mL monitored with 
LAIT for presenting non–IgE-mediated allergic 
rhinitis, allergic bronchitis, asthma, allergic sinus 
headache, atopic dermatitis, and/or urticaria. 
This cohort counted 27 males; mean age 39.9 
years; SD 20.5 years; range 3 to 84 years; 
median 39.5 years; modes = 24 and 41 years 
(each appeared four times). 

 
This study did not include patients under 
biological and/or systemic anti-inflammatory 
therapy. These procedures were offered to 
patients with clinical suspicion of PEGs’ 
hypersensitivity who demonstrated a non-
reactive or inconclusive allergic skin scrape test 
against PEGs’ solution [29]. 

2.2 PEGs Solution  
 
The PEGs solution was prepared with powdered 
PEG 4000 (acquired from ACS Científica) diluted 
with isotonic saline solution at 1 mg/mL to 
perform the allergic skin scrape tests, TTP, and 
LAIT.  
 

2.3 Ex vivo Investigation: Leukocyte 
Adherence Inhibition Test 

 
2.3.1 Procedure for allergen ex vivo 

challenging  
 
We performed the LAIT as previously described 
[43-51]. Shortly, each donor’s fresh plasma was 
divided into two parts and used in paralleled ex 
vivo challenging tests with PEGs solution 1 
mg/mL and the unchallenged plasma assay. We 
collected the plasma with high leukocyte content 
(buffy coat) from the heparinized tube after one 
hour of sedimentation at 37°C. Then we 
distributed aliquots of 100 μL into Eppendorf 
tubes kept under agitation for 30 minutes (200 
rpm at 37°C) with PEGs solution (10μL of a 
solution with 1mg/mL and pH 7.5) or without 
PEGs solution (when used as control). 
 
2.3.2 Procedure for Adherence Assay  
 
After incubation, the plasma was allocated into a 
standard Neubauer hemocytometer counting 
chamber with a plain, non-metallic glass surface 
and left to stand for 2 hours at 37 °C in the 
humidified atmosphere of the covered water bath 
to allow leukocytes to adhere to the glass. Next, 
we counted the leukocytes, removed the 
coverslip, and washed the chamber by 
immersion in a beaker with PBS at 37°C. Then, 
we added a drop of PBS to the hemocytometer’s 
chamber and allocated a clean coverslip over it. 
The remaining cells were counted in the same 
squares as previously examined.  
 
2.3.3 Procedure for calculating the 

adherence inhibition  
 
The percentage of Leukocyte Adherence (LA) of 
each assay was estimated as: (the number of 
leukocytes observed on the hemocytometry 
chamber after washing divided by the number of 
leukocytes observed on the hemocytometry 
chamber before washing) and multiplied by 100 
(%). The Leukocyte Adherence Ratio (LAR) was 
estimated based on the ratio between the LA 
from the antigen-specific challenged plasma and 
the LA from the unchallenged control plasma: 



 
 
 
 

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LAR = LA of the challenged sample divided by 
LA of unchallenged control plasma multiplied by 
100 (%). To further calculate the Leukocyte 
Adherence Inhibition (LAI), we subtracted the 
LAR from 100 (%). We employed the LAI results 
for the cascade distribution chart and the 
statistics calculations, both performed with the 
help of the Microsoft Excel® statistical package. 
 

2.4 In vitro Investigation: Tube Titration 
of Precipitins (TTP) 

 

As previously reported, the semi-quantitative 
TTP against the PEGs solution was performed in 
a transparent vitreous tube [52]. Shortly, the 
patient’s blood was collected in a clot-activator 
collecting tube. After separation, the serum was 
centrifugated at 2,000 rpm for 10 minutes. The 
allergen extracts were allocated in sets of eleven 
glass tubes at progressive duplicated serum 
dilutions. The progressive dilutions were 
combined with the 15 μL of the antigen (PEGs 
solution 1 mg/mL) with 250 μL of the patient’s 
serum, progressively diluted into physiological 
saline solution (NaCl 0,9%) in the dilution ratios 
of 1:1; 1:2; 1:4; 1:8; 1:16; 1:32; 1:64; 1:128; 
1:256; and 1:512. One tube was a blank control 
done with the water and serum to observe 
occasional spontaneous precipitation (Sia Test). 
After 24 hours, one of us examined the tubes, 
and the titers (the highest dilution factor that 
yields a positive visual reading) were recorded 
[53]. 

3. RESULTS 
 
As a retrospective survey, there was no research 
protocol; therefore, we report the incidental 
immune investigation as registered in the digital 
medical charts. 
 
The TTP showed a wide distribution range of 
results. There were two negative results. Most 
positive results concentrated on the higher 
dilutions (Fig. 1). The mean was estimated at 
1:246; the median was 1:192; the SD was 
estimated at 1:188; the mode was 1:512 
(appeared 30 times). All Sia tests were          
negative. 
 
The LAIT showed a wide distribution range of 
results. The LAI ranged from 0% to 98%. The 
mean was 41.9%; the median was 45%; the SD 
was 28.5%; the mode was 0% (appeared seven 
times). The cascade distribution demonstrates a 
wide range of distribution of LAI results (Fig. 2). 
Six patients ignored the presence of the allergen 
on the plasma and presented no inhibition of 
leukocyte adherence (LAI = 0%) after contact 
with the PEGs extract (6% of the tests). Some 
patients showed any or low immunoreactivity 
during the ex vivo challenge test, while most 
displayed moderate or strong immunoreactivity, 
which could reflect the participation of         
PEGs antibodies in a non–IgE-mediated 
hypersensitivity condition. 

 

 
 

Fig. 1. Cascade distribution chart of the Tube Titration of Precipitins (TTP on the x-axis %) 
resulting from the PEGs solution against the serially diluted serum of a cohort of 100 

tests/subjects (y-axis) 



 
 
 
 

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105 

 

 
 
Fig. 2. Cascade distribution chart of the range groups of Leukocyte Adherence Inhibition (LAI) 
results (x-axis %) of ex vivo PEGs solution monitored by the Leukocyte Adherence Inhibition 

Test (LAIT), according to the respective number of outcomes over a cohort with 100 
tests/subjects (y-axis) 

 

4. DISCUSSION 
 
More and more patients are being diagnosed 
with PEG allergy as an increasing number of 
processed foods, cosmetics, household 
products, vaccines, and medicines add PEG to 
their composition [54]. The development of new 
PEGylated compounds is advancing, especially 
among biological immune therapeutics [55]. 
PEGs are the polymer of choice for the delivery 
of small drugs, proteins, oligonucleotides, and 
liposomes due to their biocompatibility and ability 
to improve pharmacokinetics; however, PEGs 
are a mixture of different polymers that can 
stimulate unwanted immunogenic reactions, 
leading to efforts to develop                             
synthetic (monodisperse) more uniform PEGs 
[56]. 
 
PEG hypersensitivity is associated with 
medicine-induced anaphylaxis, and PEGylated-
drug reactions should prompt PEGs 
hypersensitivity investigations, especially by use 
of skin allergic tests with high molecular weight 
PEGs [57]. Diagnosing PEG allergy is 
challenging since the traditional allergy testing 
approaches are insufficient to demonstrate all 
kinds of hypersensitivity reactions [58]. Besides 
the IgE-mediated hypersensitivity reaction, the 
production of specific IgM and IgG antibodies 
and Complement activation were also described 

[59]. Since a methodology for specific-IgE 
research has not yet been described, the 
feasibility of in vitro challenge tests, such as the 
Basophil Activation Test, is being studied [60]. 
Brand new allergic diagnostic techniques (such 
as performing tests with the help of PEGylated 
liposomes) are currently being designed to 
endotype PEGs allergies [61]. 
 
The precise diagnosis of the phenotypes and 
endotypes of hypersensitivities to PEGylated 
medications is paramount for prescribing a 
successful PEGs desensitization [62]. 
 
This retrospective compilation of our data 
showed a large distribution of results when we 
ascertained the results of TTP and TIAL to 
explore humoral and cellular immunoreactivity 
against PEGs. These immunoassays do not 
identify the exact mechanisms responsible for 
the clinical condition. Instead, they provide clues 
about sensitization and immunoreactivity 
distributed into an extensive spectral range 
between immune tolerance and symptomatic 
hypersensitivity. 
 
The semi-quantitative study of the serological 
reactions through the titration of precipitins was 
the most basic laboratory exam upon which the 
fundamentals of Immunology were constructed 
[63]. Precipitating antibodies are classically 



 
 
 
 

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associated with a robust immune humoral 
response against an antigen or allergen [64]. 
The LAIT is an ex vivo challenge test performed 
with a viable leukocyte buffy coat that can 
theoretically explore most known immune 
pathways as it allows the interaction of all 
immune-circulating participants with the allergen 
[65]. Similarly, with the TTP, the LAIT did not 
indicate which pathways are involved in 
producing the immune cellular response 
demonstrated by the leukocyte adherence 
inhibition [66-69]. 
 
TTP and LAIT must be interpreted as humoral 
and cellular markers of the immune response 
after contact with a specific antigen, configuring 
themselves as techniques to endotype the 
exposome phenomenon, as proposed by the 
exposome-wide association study [70]. TTP and 
LAIT are complementary triage tests used at our 
facilities to select worthwhile antigens to proceed 
with more laborious in vivo provocation tests 
when the specific IgE is undetectable. None of 
our patients presented an exclusive reaction to 
PEGs. Every patient was simultaneously tested 
with several chemical and biological allergens, 
demonstrating positive results for some of them. 
Our clinical experience suggests that                   
reactive allergic patients may impair their 
symptoms by an additional immunoreactivity 
against PEGs. 
 
This preliminary retrospective survey 
demonstrated an extensive range of results from 
the TTP and the ex vivo challenge test 
monitored by LAIT against PEGs in two cohorts 
of patients with various allergic symptoms. The 
preliminary results support that the TTP and 
LAIT performed with 1 mg/mL PEGs 4000 
solution may discriminate diverse degrees of in 
vitro and ex vivo immunoreactivity in patients 
suffering from diversified allergic phenotypes. It 
is worth carrying out more in-depth studies to 
evaluate the usefulness of TTP and LAIT in 
endotyping non–IgE-mediated hypersensitivity to 
PEGs. 
 

5. CONCLUSION 
 

Our society is now flooded with chemical 
derivatives through industrialized foods, 
cosmetics, cleansing products, and medicines 
producing unknown immunoreactivity [71]. Our 
preliminary results show that the LAIT and TTP 
may differentiate diverse degrees of 
immunoreactivity against PEGs in patients 
clinically diagnosed with non–IgE-mediated 

allergies. This methodology can provide a 
socioeconomic impact since the technology to 
perform TIAL and TTP is inexpensive and can 
be achieved in a single lab room attached to the 
clinical facility with minimum laboratory 
equipment. However, the propaedeutic meaning 
of these results and the possibility of              
interferents must be better established [72]. 
Studies designed to establish diagnostic               
cutoffs focused on the quality-by-design 
approach with prospective larger double-blind 
cohorts need to evaluate the potential 
contribution of LAIT and TTP for endotyping 
immunoreactivity in patients suspected of 
symptomatic hypersensitivity against PEGs 
[73,74].  
 

6. LIMITATIONS 
 
This study is a retrospective analysis of data 
collected over six years. There was no protocol 
research, and the subject’s data were limited to 
the essentials available on our electronic sheets. 
Therefore, we could not establish a cross-
comparison between positive and negative 
controls to validate the results. The number of 
subjects is appropriate for a preliminary study; 
however, future studies must be more 
comprehensive and include patients’ follow-up 
information. The lack of a research protocol 
implies the possibility of a bias produced by the 
physician’s point of view, which indicated the 
exam (CEO) based on a clinical suspicion led by 
the anamnesis, physical examination, and in vivo 
provocation tests. The study lost many of these 
patients to follow-up, so assuring the                    
relationship between the immunoassays’ results 
and the patient’s clinical outcome is impossible. 

 
DISCLAIMER (ARTIFICIAL INTELLIGENCE) 

 
Authors hereby declare that no generative AI 
technologies such as Large Language Models 
(ChatGPT, COPILOT, etc) and text-to-image 
generators have been used during the writing or 
editing of manuscripts. 

 
CONSENT 
 
It is not applicable. 

 
ETHICAL APPROVAL 
 
The authors have collected and preserved 
written ethical approval per international 
standards. 



 
 
 
 

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107 

 

ACKNOWLEDGEMENTS 
 
The Instituto Alergoimuno de Americana funded 
this work. We want to thank our new laboratory 
technician, Alessandra Vieira de Oliveira, for 
participating in the exams. 
 

COMPETING INTERESTS 
 
Authors have declared that no competing 
interests exist. 
 

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