Subject enrolment
Subjects (the treatment and control groups) were enrolled in this research at The First Affiliated Hospital of Guangzhou Medical University. The inclusion criteria were as follows: 1. Subjects were less than eighteen years old. 2. Subjects had skin prick tests that were positive for Der p1 and Der f1 allergens. ImmunoCAP was performed to confirm the presence of Der p-specific IgE and Der f-specific IgE in positive subjects (defined as ≥ 0.35 KU/L). 3. According to the 2018 Global Strategy for Asthma Management and Prevention issued by the Global Initiative for Asthma (GINA) [9], asthma can be classified according to severity. Subjects enrolled in this study were diagnosed with mild asthma (acute exacerbations could be alleviated by a short term β2 receptor agonist without the need for maintenance medication). 4. During the study, the subjects were not allowed to change their place of residence. Subjects needed to adhere well to the instructions provided pertaining to the experiment (the treatment group needed to use the air purifier, complete the questionnaire, return for a follow-up visit, and allow the collection of samples; the control group needed to complete the same steps, with the exception of using the air purifier). All subjects signed an informed consent form.
Grouping and processing
This research utilized a random number sequence to assign individuals to the treatment group and control group; the treatment group used air purifiers, while the control group did not.
Air purifier intervention in the treatment group
This research provided a new HEPA air purifier (BA1030/1045, BRI air purifier, Xiamen, China) to every subject in the treatment group; it was placed in the bedroom. Subjects were asked to utilize the air purifier for six consecutive months (August 2016 to February 2017). Before the subjects used the air purifiers, the subjects were taught how to use the air purifier correctly by one of the researchers. Researchers checked the air purifiers when they collected samples each month. The filter of each air purifier was cleaned after three months.
Air purifier usage
The air purifier was located at the head of each subject’s bed. The air flow was carefully noted to prevent obstruction, and humidifiers were not permitted to be used at the same time. The doors and windows were kept closed when the air purifier was functioning. Moreover, the subjects were asked to use the air purifier every day and were not allowed to clear the air filter themselves. The mean duration of daily HEPA air purifier use was 9.6±3.3 hours and fairly consistent during the course of the study.
Return visit
The researcher confirmed the air purifier placement and evaluated the subjects' correct usage of the air purifier on a monthly basis. Incorrect usage was corrected. The control group did not receive any environmental interventions.
Sample collection
The researcher collected dust samples from the treatment and control groups before the air purifiers were provided and on a monthly basis after the use of the air purifiers was implemented.
Bedding dust sample collection
Glass fibre filters and vacuum cleaners (Haier, ZW1401B) were the main tools used to collect dust samples from bedding. The glass fibre filter was set on the top of the collector and fixed there with a rubber band; the vacuum cleaner was used directly on the bedding. The duration of the sample collection was 15 min.
Static dust collection
Through natural subsidence of dust particles, dust samples were collected in a static location by a glass fibre filter. Each glass fibre filter was placed in a 60-mm culture dish. The dish was placed in a location with open exposure to the room and could not be moved or covered during the month.
PM data collection
The researchers utilized a PM detector (DT-9881M, Huasheng Qi Xieco. Ltd., Shenzhen, China) to collect indoor and outdoor PM (2.5, 10) concentrations. The machine was used for five points collections (four corners of the room and the middle) indoors. Outdoor collections were performed in triplicate on the balcony or out of the window. The PMindoor/outdoor ratio was recorded.
Sample processing and extraction
Every dust sample was weighed and recorded when it was collected. Then, the glass fibre filter was cut into pieces and placed on a 10 ml syringe. Then, 1 ml (containing 1% BSA (bovine serum albumin) and 0.05% tween-20) of PBST was added, and the sample was extracted overnight at 4℃ with shaking. The extracted mixture was removed from the injector and centrifuged at 4°C and 3000×g for 30 min. The supernatant was removed and stored at -20°C.
HDM allergen content in dust
This research utilized a double antibody sandwich enzyme-linked immunoassay (Indoor Biotechnologies, Charlottesville, VA, USA-ELISA) to test for two key HDM allergens, Der p1 and Der f1. According to the manufacturer’s instructions, the concentrations of Der p1 and Der f1 were tested in each sample. The weight per gram of Der p1 and Der f1 were calculated from the concentrations of Der p1 and Der f1 and the weight of HDM.
Evaluation of subjects
ACT and C-ACT were used to evaluate the subjects' asthma control, while FENO was used to evaluate the ability of the air purifiers to improve inflammation levels in the asthma patients.
ACT
ACT, which was designed by Nathan RA [10], is commonly used in asthma control research. It is an ideal method of evaluating asthma control. The questionnaire was completed by the subjects or their parents during each dust collection visit; ACT is only valid for adolescents aged 12 years or above.
C-ACT
C-ACT, designed by Liu AH [11], is often used in asthma control research. The questions are answered by the subjects' parents. The higher the score, the better the level of asthma control is. A score greater than 20 indicates that asthma is well controlled, while a score less than 20 indicates that it is not well controlled.
FENO
FENO is a marker of airway inflammation. Subjects underwent FENO tests (NIOX, Aerocrine, Sweden) [12] at The First Affiliated Hospital of Guangzhou Medical University.
Statistical analysis
SPSS 19.0 (SPSS INC, Chicago, IL, USA) was utilized to record and analyse data from the treatment and control groups. Descriptive analysis was used to analyse the subjects' basic characteristics (gender, age, sIgE levels), the monthly HDM allergen concentrations, the PM levels and the ACT and C-ACT scores. Normally distributed data are expressed as the mean±SD, and non-normally distributed data are expressed as the median (P50 (P25, P75)). The Kolmogorov-Smirnov test was utilized to analyse the HDM allergen results. For normally distributed data, we used two independent samples for a nonparametric test analysis. According to the data distribution model, a two-tailed t test or non-parametric test was used to compare the HDM allergen concentrations and FENO rates before and after using an air purifier. Moreover, repeated ANOVA was used to analyse the monthly HDM allergen concentrations, PM ratios, ACT scores and C-ACT scores. P<0.05 was considered statistically significant.