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Keywords:

  • Allergy;
  • Blomia tropicalis;
  • Dermatophagoides pteronyssinus; specific Immunoglobulin E;
  • mite allergens

Abstract

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

Background: Blomia tropicalis is a common mite found in the house dust of many tropical countries including Venezuela. The prevalence of skin test and specific serum immunoglobulin (Ig)E reactivity to B. tropicalis in Venezuela has not been previously evaluated.

Methods:  In the present study we evaluated the skin reactivity by skin prick test and specific IgE by a multiple antigen blot assay, against B. tropicalis and Dermatophagoides pteronyssinus, in a group of 115 subjects who attended the Allergy Clinic of the Institute of Biomedicine, Caracas, Venezuela, and we studied possible cross reactions between similar proteins of these two mites.

Results:  One hundred and six patients with persistent allergic respiratory symptoms showed a positive skin prick test to at least one of the mite extracts, with the frequency of positive reactions to B. tropicalis being as high as to D. pteronyssinus. Twelve patients reacted only to D. pteronyssinus and 13 different patients only to B. tropicalis. Specific IgE to each of the mite extracts was found with similar frequency, and the results coincided with the skin test reactivity.

Conclusions:  The study indicated the importance of including B. tropicalis in routine diagnostic testing in tropical and sub-tropical situations.

It is well established that house dust mites are important causes of persistent allergic diseases, such as asthma and rhinitis (1–3). Previous studies have demonstrated that Dermatophagoides pteronyssinus, Dermatophagoides farinae and Blomia tropicalis are the most common mites in the house dust of tropical and subtropical areas (4, 5). Blomia tropicalis is a domestic mite of the Glyciphagidae family that, in contrast to Dermatophagoides, is more commonly found in tropical compared with temperate climates (4, 6). Blomia tropicalis has been found in dust from homes located in many countries, including Spain, India, Taiwan, Brazil, Colombia, the Philippines and Indonesia (5–10). In Venezuela, Hurtado and Parini (11) demonstrated that B. tropicalis and D. pteronyssinus were the most frequent mites in house dust samples. However, for the clinical routine diagnosis of allergic patients, B. tropicalis has not usually been employed. The important role of this mite as an allergen that can induce allergic diseases has been previously described (7–9). The presence of positive skin tests and/or serum immunoglobulin (Ig)E antibodies to B. tropicalis has been demonstrated in allergic patients (4, 10). Moreover many authors found that there is significant cross reactivity between D. pteronyssinus and B. tropicalis, and that this is especially related to the group 5 allergens of these mites, although many other B. tropicalis allergens may also present cross reactivity with other mites (12–14). Therefore, it is of great value to determine whether the sensitization against B. tropicalis would be mainly directed to species-specific allergens.

As the reactivity of Venezuelan allergic patients to D. pteronyssinus and B. tropicalis was similar and species specific, it would be important to include B. tropicalis in the routine diagnostic test. In the present study we evaluated skin test reactivity and specific IgE in a group of Venezuelan patients with persistent allergic symptoms to B. tropicalis and D. pteronyssinus extracts, as well as possible cross reactivity between these two mites.

Subjects

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

We studied a group of 115 subjects who attended the Allergy Clinic, Institute of Biomedicine, Caracas, Venezuela. This study was performed as part of the routine evaluation of patients attending the Institute of Biomedicine after approval by the ethical committee of the Institute, ratified by the Academic Council of the Medical Faculty of Central University of Venezuela. The patients were classified according to the criteria of the International Consensus Report (15) as suffering rhinitis only, the patients with symptoms of asthma without significant rhinitis as defined by the guidelines of the WHO/NIH Global Initiative for Asthma (GINA) (16) and the patients with combined asthma and rhinitis.

The mean age of these subjects was 25 years and 67% were female. We performed skin testing with B. tropicalis and D. pteronyssinus extracts and specific serum IgE detection by a multiple antigen blot assay (MABA) (17).

We evaluated 30 subjects who reported no history of allergic disease as a control group.

Preparation of mite extracts

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

Cultured D. pteronyssinus (Commonwealth Serum Laboratories, Parkeville, Australia and Biopol Inc, Spokane, Wash, USA) were de-fatted by Soxhlet extraction with diethyl ether, extracted for 48 h at 4°C with 0.05 M NH4HCO3 saturated with chloroform and then concentrated/dialyzed using a hollow-fiber filter with a nominal molecular weight cut off of 5 Kd (Amicon Corp., Lexington, MA, USA). Total protein concentrations were determined by the Bradford method, then the extracts were membrane filtered and sterility tested by routine bacteriological methods. Blomia tropicalis extracts were prepared as described previously (8) and lyophilized material was reconstituted at concentrations of 50 μg/ml in carbonate-bicarbonate buffer, pH 9.5 for MABA or 50% glycerol, 0.4% phenol for skin tests.

Skin prick tests

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

Skin prick tests were performed with D. pteronyssinus and B. tropicalis extracts at 0.1 mg/ml in saline solution containing 50% glycerol and 0.4% phenol. Control tests were performed with the diluent alone and with 1% w/v histamine dihydrochloride. The wheal diameters were measured after 20 min, and diameters ≥3 mm were considered positive. Anti-allergic medication was suspended for 5 days before the test.

Detection of specific IgE

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

A qualitative technique based on a dot-blot enzyme-linked immunosorbent assay (ELISA) methodology was employed, as described by Noya and Alarcon de Noya, 1998 (17), modified in our laboratory for the detection of IgE antibodies and compared with skin prick test and ELISA. Fifty microliters of the mite extracts were fixed at concentrations of 50 μg/ml in carbonate–bicarbonate buffer, pH 9.5, onto nitrocellulose membranes (Schleicher and Schuell, Keene, NH, 0.2 μm, 7.5 × 10 cm) in each channel of the Miniblotter 28 SL (Immunetics Inc, Cambridge, MA) apparatus for 2 h at room temperature. The membranes were then washed three times with phosphate-buffered saline (PBS)-0.05% Tween 20 on an orbital shaker for 10 min each. Blocking was achieved using 5% nonfat milk in PBS-0.05% Tween 20 for 2 h on an orbital shaker. Strips of 2 mm width were then cut perpendicularly to the allergen lanes, so that each strip contained both allergen preparations. These were exposed to 300 μl of the patients’ sera diluted 1 : 4 in blocking solution over night at 4°C. After washing, the strips were incubated for 2 h at room temperature with peroxidase-labeled anti-IgE (Sigma Chemical Co., St Louis, MO) diluted 1 : 1000 in blocking solution. The strips were again washed and then exposed for 10 min to the chemiluminescent substrate Luminol (ECL Detection System, Amersham, Little Chalfont, UK). Finally the strips were exposed to Hyperfilm (Amersham) in darkness and developed photographically for 5–20 s depending on the reactivity of the assayed sera. Positive reactions to the allergens are seen as small black squares on the strips. As a positive control, unlabeled monoclonal anti-IgE (Sigma) fixed directly to the membrane was used, and the negative control was the solution in which the allergens were diluted (Fig. 1).

image

Figure 1. Multiple antigen blot assay (MABA). The MABA-developed strips. Numbers correspond to strips exposed to sera from nonallergic (1) and allergic (2) individuals. Columns correspond to Positive control: monoclonal anti-IgE (A), negative control (diluent) (B), Dermatophagoides pteronyssinus (C) and Blomia tropicalis extracts (D). The serum from the allergic patient had specific immunoglobulin E to both D. pteronyssinus (C) and B. tropicalis (D) extracts.

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Cross reactivity studies

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

To evaluate the potential allergenic cross-reactivity between D. pteronyssinus and B. tropicalis, MABA inhibition was performed. Twelve positive allergic serum samples were preincubated with soluble extracts of D. pteronyssinus and B. tropicalis at concentrations of 1 mg/ml for 2 h, and then incubated with the sensitized strips. The inhibition caused by the competition between the soluble allergens and solid phase allergens was revealed as negative MABA reactions.

Data analysis

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

We calculated sensitivity and specificity of the MABA using skin prick and ELISA tests as gold standard.

The cut off of the ELISA was determined using the average optical density plus 2 SD of the 30 healthy individuals.

Patients studies and skin test reactivity

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

From a total of 115 individuals evaluated, 107 had persistent allergic respiratory symptoms upon exposure to house dust. Rhinitis was reported in 37% of the patients, and asthma was reported in 18%, with the combined prevalence of the disease being 45%.

One hundred and six patients were skin prick test positive to at least one of the mite extracts; 81 were positive to both mite extracts, 12 reacted only to D. pteronyssinus and 13 different subjects only to B. tropicalis. The prevalence of skin prick test reactions to both mites was higher in the group of patients with combined rhinitis and asthma. Nine subjects had no allergic respiratory symptoms upon exposure to house dust, and were skin prick test negative to both mite extracts. The 30 normal subjects were negative to skin tests.

Detection of specific IgE

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

Sera of 107 patients showed specific IgE to either or both mite extracts; 80 patients showed specific IgE to both mite extracts, 13 sera reacted only to D. pteronyssinus and 14 different sera only to B. tropicalis. Eight patients did not react to either of the mite extracts. No specific IgE to D. pteronyssinus or B. tropicalis was detected in the 30 normal sera, although they did react to the anti-IgE positive control.

We compared MABA-specific IgE results with skin prick test and ELISA results, using skin prick tests or ELISA as the gold standard methods. The specificity and sensitivity of the MABA assay compared with skin prick tests as the standard when using both extracts was 94 and 88%, respectively (Table 1). Compared with ELISA the specificity and sensitivity of MABA was 96 and 91% respectively (Table 2).

Table 1.  Comparison between multiple antigen blot assay (MABA) and skin prick test results in allergic patients who were simultaneously positive or negative to both mite extracts, using the skin prick test as the standard
 Skin prick test +Skin prick test −Total
  1. Sensitivity: 76/81 = 93, 83%. Specificity: 30/34 = 88, 24%.

MABA +76480
MABA −53035
Total8134115
Table 2.  Comparison between multiple antigen blot assay (MABA) and enzyme-linked immunosorbent assay (ELISA) results in allergic patients who were simultaneously positive or negative to both mite extracts, using the ELISA methodology as the gold standard
 ELISA +ELISA −Total
  1. Sensitivity: 77/80 = 96, 25%. Specificity: 32/35 = 91, 43%.

MABA +77380
MABA −33235
Total8035115

Cross reactivity study

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

The MABA-inhibition was used to evaluate the potential allergenic cross-reactivity between D. pteronyssinus and B. tropicalis in 12 allergic sera (Fig. 2).

image

Figure 2. Cross reactivity studies. (A) Multiple antigen blot assay (MABA)-developed strips. Numbers correspond to strips exposed to sera from 12 allergic individuals without any absorption prior to incubation. Sera of patients 1, 2, 7, 10, 12 reacted to both mite extracts. Sera of patients 4, 6, 8, 11 reacted only to Dermatophagoides pteronyssinus and sera of patients 3, 5, 9 reacted only to Blomia tropicalis. Rows correspond to Positive control: monoclonal anti-IgE (A), negative control (B), D. pteronyssinus (C) and B. tropicalis extracts (D). (B). The MABA-Inhibition after previous incubation with soluble D. pteronyssinus. In sera from patients 10 and 12, soluble D. pteronyssinus inhibited binding to B. tropicalis (C). The MABA-inhibition after previous incubation with soluble B. tropicalis. In sera from patients 10 and 12, soluble B. tropicalis inhibited the binding to D. pteronyssinus.

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Five serum samples that reacted to both mite extracts (1, 2, 7, 10, 12), four that reacted only to D. pteronyssinus (4, 6, 8, 11) and three that reacted only to B. tropicalis (3, 5, 9) were preincubated with either D. pteronyssinus or B. tropicalis soluble extract as described previously.

In patients who had specific IgE only to D. pteronyssinus, soluble B. tropicalis did not inhibit the binding to D. pteronyssinus and in patients who reacted only to B. tropicalis, the soluble form of D. pteronyssinus did not inhibit the binding to B. tropicalis. In contrast, in two of five sera that reacted to both mite extracts, soluble D. pteronyssinus inhibited the binding to B. tropicalis and B. tropicalis also inhibit the binding to D. pteronyssinus. These results suggest that there was a moderate degree of cross reactivity between these two mites.

Discussion

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

In Venezuela the prevalence of skin test reactivity to B. tropicalis has not been previously evaluated. It has been demonstrated that allergic patients are highly reactive to house-dust mites of the pyrogliphid family, which include D. pteronyssinus and D. farinae (2, 17, 18). Despite the fact that different studies have shown that B. tropicalis is commonly found in homes of many tropical countries (4), including Venezuela (11), sensitivity to this mite has been rarely evaluated in this country. In Venezuela D. pteronissynus has been used frequently as a major allergen for the evaluation of the allergic reactivity (18). In fact, in a recent questionnaire that Puccio and Di Prisco circulated to 30 allergists in Caracas, only one reported B. tropicalis extract as a usual component of their battery of inhalant allergen extracts (unpublished data).

We showed that 107 patients presented a clear history of allergic respiratory disease upon exposure to house dust and 106 showed a positive skin prick test to at least one of the mite extracts, with the frequency of positive reactions to B. tropicalis being as high as to D. pteronyssinus. The whole group of patients who had skin prick test positivity to at least one of the mite extracts had also specific IgE to one of these mites, in contrast to none of the nonallergic control group. Specific IgE to each of the mite extracts was found with similar frequency, and the results coincided with the skin test reactivity.

A high reactivity to B. tropicalis has also been demonstrated in other tropical countries. A report from Taiwan shows that asthmatic patients had positive skin tests to B. tropicalis, and it was concluded that B. tropicalis plays an important role in the pathogenesis of asthma in that country (10). In Colombia the importance of B. tropicalis as an inducer of asthma and allergic rhinitis has also been reported (8). Of interest in our results is that the patients with combined rhinitis and asthma reacted with higher frequency to both mite, in addition, some allergic patients with rhinitis and asthma reacted to B. tropicalis but not to D. pteronyssinus. Therefore patients who have unexplained allergic symptoms and are skin prick test negative to D. pteronyssinus may be sensitive to B. tropicalis, thus indicating the importance of including this nonpyrogliphid mite in routine diagnostic testing in tropical and sub-tropical situations. Also for immunotherapy, the incorporation of B. tropicalis extract may be an effective measure for treating allergic respiratory symptoms in persons sensitized to house dust.

The overall results obtained in the comparison between MABA and skin prick test, showed high specificity and sensitivity percentages, similar results were obtained when MABA was compared with ELISA. We have previously used the ELISA technique to detect specific serum IgE (19), as it has been reported as a confiable methodology by other authors (20, 21). The MABA test allows the simultaneous screening of serum with several allergens and it would represent an important method to detect rapid specific IgE to several allergens in allergic patients.

To achieve a correct diagnosis and treatment of allergic diseases, it is important to evaluate if the reactivity found in patients is allergen-specific. We found that 17 patients had detectable specific IgE against B. tropicalis and not to D. pteronyssinus; this suggests the possibility of species-specific IgE reactions. Many authors described a certain degree of cross reactivity between D. pteronyssinus and B. tropicalis (12–14) and this cross reactivity was mainly associated with the group 5 allergens: Blot 5 and Der p 5 (12).

We demonstrated by MABA-inhibition tests, that there were patients who had species-specific IgE, but in others patients we confirm the existence of some degree of cross reactivity between these two mites.

Our results are compatible with at least two possible explanations. Some patients may react to common allergens shared by both mites, and reactivity with either one may inhibit subsequent reactivity to the other in the MABA inhibition test. The other possibility is that some patients are sensitized by previous contact with both mites present at high levels in Caracas house dust rather than a single common allergen.

Acknowledgments

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References

We thank Dr Marian Ulrich and Dr Rafael Borges for their useful comments. We would also like to thank T.S.U Emperatriz Mata and Wilman Clark for their computer assistance. This work was supported by grant CONICIT S1-96000543 and by Millennium Scientific Initiative grant 4572B.

References

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Subjects
  5. Preparation of mite extracts
  6. Skin prick tests
  7. Detection of specific IgE
  8. Cross reactivity studies
  9. Data analysis
  10. Results
  11. Patients studies and skin test reactivity
  12. Detection of specific IgE
  13. Cross reactivity study
  14. Discussion
  15. Acknowledgments
  16. References
  • 1
    Benaim-Pinto C. La alergia y la inmunología clínica en Venezuela como prototipo de país tropical. In: Memorias de las IV Jornadas de Alergia e Immunologia. Caracas: Impresos Gherd, 1984: 2628.
  • 2
    Lynch NR, Di Prisco-Fuenmayor MC. High allergic reactivity in a tropical environment. Clin Allergy 1984;14: 233240.
  • 3
    Platts-Mills TA, De Weck AL. Dust mite allergens and asthma – a world wide problem. J Allergy Clin Immunol 1989;83: 416472.
  • 4
    Arlian LG, Vyszenski-Moher DL, Fernandez-Caldas E. Allergenicity of the mite Blomia tropicalis. J Allergy Clin Immunol 1993;91: 10421050.
  • 5
    Fernandez-Caldas E, Puertas L, Mercado D, Lockey R, Caraballo L. Mite fauna, Der p I, Der f I and Blomia tropicalis allergen levels in a tropical environment. Clin Exp Allergy 1993;23: 292297.
  • 6
    Bronswijk JE, De Cock W, Oshima S. The genus Blomia Oudemans (Acari: Glycyphagidae). I. Description of Blomia tropicalis sp. from house dust in tropical and subtropical regions. Acarología 1974;15: 477489.
  • 7
    Arruda LK, Rizzo MC, Chapman MD, Fernández-Caldas E, Baggio D, Platts-Mills TA et al. Exposure and sensitization to dust mite allergens among asthmatic children in Sao Paulo, Brazil. Clin Exp Allergy 1991;21: 433439.
  • 8
    Caraballo L, Puerta L, Fernandez-Caldas E, Lockey RF, Martinez B. Sensitization to mite allergens and acute asthma in a tropical environment. Investic Allergol Clin Immunol 1998;85: 281284.
  • 9
    Schuhl JF. Asthma and allergy to mites in Montevideo. Preliminary study. Allergol Immunopathol 1977;5: 117122.
  • 10
    Tsai JJ, Wu HH, Shen HD, Hsu EL, Wang SR. Sensitization to Blomia tropicalis among asthmatic patients in Taiwan. Int Arch Allergy Immunol 1998;115: 144149.
  • 11
    Hurtado I, Parini M. House dust mites in Caracas, Venezuela. Ann Allergy 1987;59: 128130.
  • 12
    Caraballo L, Mercado D, Jimenez S, Moreno L, Puerta L, Chua KY. Analysis of the cross-reactivity between BtM and Der P 5, two group 5 recombinant allergens from Blomia tropicalis and Dermatophagoides pteronyssinus. Int Arch Allergy Immunol 1998;117: 3845.
  • 13
    Stanaland BE, Fernandez-Caldas E, Jacinto CM, Trudeau WL, Lockey RF. Sensitization to Blomia tropicalis: skin test and cross-reactivity studies. J Allergy Clin Immunol 1994;94: 452457.
  • 14
    Simpson A, Green R, Custovic A, Woodcock A, Arruda LK, Chapman MD. Skin test reactivity to natural and recombinant Blomia and Dermatophagoides spp. allergens among mite allergic patients in the UK. Allergy 2003;58: 5356.
  • 15
    International consensus report on the diagnosis and management of rhinitis. International Rhinitis Management Working Group. Allergy 1994;49(Suppl.):134.
  • 16
    WHO/NIH. Global initiative for asthma (GINA). Washington, DC, USA: National Institutes of Health, Publication 95-3659, 1995.
  • 17
    Noya O, Alarcon de Noya B. The multiple antigen blot assay (MABA): a simple immunoenzymatic technique for simultaneous screening of multiple antigens. Immunol Lett 1998;63: 5356.
  • 18
    Lynch NR, Di Prisco-Fuenmayor MC, Soto J. Diagnosis of atopic conditions in the tropics. Ann Allergy 1983;51: 547551.
  • 19
    Hagel I, Salgado A, Rodriguez O, Ortiz D, Hurtado M, Puccio F et al. Factores que influyen en la prevalencia e intensidad de las parasitosis intestinales en Venezuela. Gac Méd Caracas 2001;109: 8290.
  • 20
    Yi FC, Cheong N, Shek P, Wang D, Chua K, Lee B. Identification of shared and unique immunoglobulin E epitopes of the highly conserved tropomyosins in Blomia tropicalis and Dermatophagoides pteronyssinus. Clin Exp Allergy 2002;32: 12031210.
  • 21
    Odashima NS, Takayanagui OM, Figueiredo JF. Enzyme linked immunosorbet assay (ELISA) for the detection of IgG, IgM, IgE and IgA against Cysticercus cellulosae in cerebrospinal fluid of patients with neurocysticercosis. Arq Neuropsiquiatr 2002;60: 400405.