Advancing drug allergy diagnosis in Tunisia using flow cytometry: a promising first step
Main Article Content
Keywords
basophils, BAT, drug allergy, flow cytometry, IgE
Abstract
Background: Hypersensitivity reactions to drugs represent a significant and growing health concern worldwide, with the potential to cause severe and life-threatening outcomes. Accurate diagnosis of drug allergies is essential for effective patient management, particularly in differentiating true allergic reactions from other adverse drug reactions.
Objective: This study aimed to evaluate for the first time the Basophil Activation Test (BAT) using flow cytometry as a reliable diagnostic tool for immediate-type drug allergies in Tunisia.
Methods: This study included 35 BATs, conducted between November 2022 and December 2024. Only one patient underwent two separate BATs on different drugs. From each patient, venous blood was drawn and BAT was assessed using flow cytometry. Serial dilutions of suspected drugs, including antibiotics (e.g., amoxicillin and cefixime), non-steroidal anti-inflammatory drugs (e.g., mefenamic acid), and others, were tested. For each sample, basophils were incubated with the drug dilutions stained with antibodies marked with different fluorescents, such as CRTH2-FITC, CD3-PC7, CD203c-PE, and then analyzed on a Navios Ex flow cytometry.
Results: Of all the performed BATs, antibiotics represented the majority of tested drugs accounting for 56%. However, only three tests were positive, in favor of a type I hypersensitivity drug reaction.
Conclusion: By applying BAT in the Immunology Department of the Military Hospital of Tunisia, this study seeks to enhance diagnostic accuracy and contribute to personalized patient care.
References
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2 Lei Y, Gregory JA, Nilsson GP, Adner M. Insights into mast cell functions in asthma using mouse models. Pulm Pharmacol Ther. 2013;26:532–9. 10.1016/j.pupt.2013.03.019
3 Johansson SGO, Hourihane JO’B, Bousquet J, Bruijnzeel-Koomen C, Dreborg S, Haahtela T, et al. A revised nomenclature of allergy: An EAACI position statement from the EAACI nomenclature task force. Allergy. 2001;56(9):813–824. 10.1034/j.1398-9995.2001.t01-1-00001.x
4 Holgate ST. The role of mast cells and basophils in inflammation. Clin Exp Allergy. 2000;30:28–32. 10.1046/j.1365-2222.2000.00093.x
5 Buhring HJ, Simmons PJ, Pudney M, Foster SJ, van der Walt J, Morrison C, et al. The monoclonal antibody 97A6 defines a novel surface antigen expressed on human basophils and their multipotent and unipotent progenitors. Blood. 1999;94(7):2343–56.
6 Ebo DG, Hagendorens MM, Bridts CH, De Knop KJ, Stevens WJ. The basophil activation test in immediate drug allergy. Acta Clin Belg. 2009;64(2):129–35. 10.1179/acb.2009.022
7 Sainte-Laudy J, Boumediene A, Touraine F, Orsel I, Cogné M. Analysis of IgE downregulation induced by basophil activation. Application to the diagnosis of muscle relaxant allergic hypersensitivity by flow cytometry. Inflamm Res. 2006;55(Supp I): S21–2. 10.1007/s00011-005-0023-3
8 Garvey LH, Ebo D, Mertes PM, et al. An EAACI position paper on the investigation of perioperative immediate hypersensitivity reactions. Allergy. 2019;74:1872–84. 10.1111/all.13820
9 Moneret-Vautrin DA, Sainte-Laudy J, Kanny G, Fremont S. Human basophil activation measured by CD63 expression and LTC4 release in IgE-mediated food allergy. Ann Allergy Asthma Immunol. 1999;82:33–40. 10.1016/S1081-1206(10)62657-9
10 Cozon G, Ferrandiz J, Peyramond D, Brunet J. Detection of activated basophils using flow cytometry for diagnosis in atopic patients. Allergol Immunopathol (Madr). 1999;27:182–7.
11 Hauswirth AW, Natter S, Ghannadan M, Wolf M, Krauth MT, Valenta R, Sperr WR. Recombinant allergens promote expression of CD203c on basophils in sensitized individuals. J Allergy Clin Immunol. 2002;110:102–9. 10.1067/mai.2002.125257
12 Paris-Kohler A, Demoly P, Persi L, Lebel B, Bousquet J, Arnoux B. In vitro diagnosis of cypress pollen allergy by using cytofluorimetric analysis of basophils (Basotest). J Allergy Clin Immunol. 2000;105:339–45. 10.1016/S0091-6749(00)90085-X
13 Abuaf N, Rajoely B, Ghazouani E, Bricard H, Nicolas JF, Nguyen QT. Validation of a flow cytometric assay detecting in vitro basophil activation for the diagnosis of muscle relaxant allergy. J Allergy Clin Immunol. 1999;104:411–8. 10.1016/S0091-6749(99)70386-6
14 Sanz ML, Gamboa PM, Antepara I, Fernández J, Gastaminza G, Sanz ML. Flow cytometric basophil activation test by detection of CD63 expression in patients with immediate-type reactions to betalactam antibiotics. Clin Exp Allergy. 2002;32:277–86. 10.1046/j.1365-2222.2002.01305.x
15 Kahlert H, Cromwell O, Fiebig H. Measurement of basophil-activating capacity of grass pollen allergens, allergoids and hypoallergenic recombinant derivatives by flow cytometry using anti-CD203c. Clin Exp Allergy. 2003;33:1266–72. 10.1046/j.1365-2222.2003.01756.x
16 Pâris-Köhler A, Demoly P, Persi L, Lebel B, Bousquet J, Arnoux B. In vitro diagnosis of cypress pollen allergy by using cytofluorimetric analysis of basophils (Basotest). J Allergy Clin Immunol. 2000;105:339–45. 10.1016/S0091-6749(00)90085-X
17 Bühring H-J, Simmons PJ, Pudney M, Müller R, Jarrossay D, van Agthoven A, et al. The monoclonal antibody 97A6 defines a novel surface antigen expressed on human basophils and their multipotent and unipotent progenitors. Blood. 1999;94:2343–56.
18 Buhring HJ., Streble A., Valent P. The basophil-specific ectoenzyme E-NNP3 (CD203c) as a marker for cell activation and allergy diagnosis. Int Arch Allergy Immunol. 2004;133:317–29. 10.1159/000077351
19 Mac Glashan Jr D. Expression of CD203c and CD63 in human basophils: Relationship to differential regulation of piecemeal and anaphylactic degranulation processes. Clin Exp Allergy. 2010 Sep;40(9):1365–77. 10.1111/j.1365-2222.2010.03572.x
20 Ene-Cocis M, Petrisor C, Sfichi M, et al. CD63 vs. CD203c for the diagnosis of neuromuscular blocking agents immediate-type hypersensitivity. Human Vet Med. 2020;12:65–9.
21 Santos AF, Douiri A, Bécares N, Wu SY, Stephens A, Radulovic S, et al. Basophil activation test discriminates between allergy and tolerance in peanut-sensitized children. J Allergy Clin Immunol. 2014;134:645–52. 10.1016/j.jaci.2014.04.039
22 Gernez Y, Waters J, Mirković B, Lavelle GM, Dunn CE, Davies ZA, et al. Blood basophil activation is a reliable biomarker of allergic bronchopulmonary aspergillosis in cystic fibrosis. Eur Res J. 2016;47:177–85. 10.1183/13993003.01068-2015
23 Varshney P, Jones SM, Scurlock AM, Perry TT, Kemper A, Steele P, et al. A randomized controlled study of peanut oral immunotherapy: Clinical desensitization and modulation of the allergic response.J Allergy Clin Immunol. 2011;127(3):654–60. 10.1016/j.jaci.2010.12.1111
24 Eržen R, Košnik M, Silar M, Korošec P. Basophil response and the induction of a tolerance in venom immunotherapy: A long-term sting challenge study. Allergy. 2012;67(6):822–30. 10.1111/j.1398-9995.2012.02817.x
25 Peck G, Hashim MJ, Shaughnessy C, Muddasani S, Elsayed NA, Fleischer AB. Global epidemiology of urticaria: Increasing burden among children, females and low-income regions. Acta Derm Venereol. 2021;101:adv00433.
26 Darragi MK, Daghfous H, Ben Mansour A, Slim A, Ben Saad S, Tritar F. Investigation of drug hypersensitivity in Tunisia. Rev Fr Allergol. 2023;63(3):103539. 10.1016/j.reval.2023.103539
27 Ghozzi H, Sahnoun Z, Hammami S, Chaabane N, Ben Hassine L. Adverse effects due to antibiotics: a retrospective study over three years at the Regional Pharmacovigilance Center of Sfax. Rev Tun Infectiologie. 2011;5(1):16–21.
28 Amsler E, Soria A. Hypersensitivity reactions to beta-lactam antibiotics. Rev Med Interne. 2017;38(11):737–748. 10.1016/j.revmed.2017.06.020
29 Ponvert C, LeBourgeois M, Scheinmann P, Paupe J. Allergic reactions to commonly used anti-infective drugs. Rev Fr Allergol Immunol Clin. 1999;39(5):455–467.
30 Mayorga C, Çelik GE, Pascal M, Campana R, de Chaisemartin L, Netchiporouk E, Klossek JM, et al. Flow-based basophil activation test in immediate drug hypersensitivity. Allergy. 2024;79(3):580–600.
31 Knol EF, Mul FP, Jansen H, Calafat J, Roos D. Monitoring human basophil activation via CD63 mono-clonal antibody 435. J Allergy Clin Immunol. 1991;88:328–38. 10.1016/0091-6749(91)90094-5
32 Hoffmann HJ, Santos AF, Mayorga C, Eberlein B, Heeringa M, de Weck AL, et al. The clinical utility of basophil activation testing in diagnosis and monitoring of allergic disease. Allergy 2015;70:1393–405. 10.1111/all.12698
33 Fernandez TD, Mayorga C, Salas M, Doña I, Torres MJ, Blanca M. Evolution of diagnostic approaches in betalactam hypersensitivity. Expert Rev Clin Pharmacol. 2017;10:671–83. 10.1080/17512433.2017.1313110
34 McGowan EC, Saini S. Update on the performance and application of basophil activation tests. Curr Allergy Asthma Rep. 2013;13:101–9. 10.1007/s11882-012-0324-x
35 Sturm GJ, Kranzelbinder B, Sturm EM, Heinemann A, Groselj-Strele A, Aberer W. The basophil activation test in the diagnosis of allergy: Technical issues and critical factors. Allergy. 2009;64:1319–26. 10.1111/j.1398-9995.2009.02004.x
36 Cañas JA, Pérez-Sánchez N, Lopera-Doblas L, Moreno Aguilar C, Torres MJ, Blanca M, et al. Basophil activation test utility as a diagnostic tool in LTP allergy. Int J Mol Sci. 2022;23:4979. 10.3390/ijms23094979
37 Ebo DG, Bridts CH, Hagendorens MM, De Clerck LS, Stevens WJ. Basophil activation test by flow cytometry: Present and future applications in allergology. Cytomet B Clin Cytomet. 2008;74B:201–10. 10.1002/cyto.b.20419
38 Ebo DG, Sainte-Laudy J, Bridts CH, Mertens CH, Hagendorens MM, De Clerck LS, et al. Flow-assisted allergy diagnosis: Current applications and future perspectives. Allergy. 2006;61:1028–39. 10.1111/j.1398-9995.2006.01039.x
39 MacGlashan DW Jr. Basophil activation testing. J Allergy Clin Immunol. 2013 Oct;132(4):777–87. 10.1016/j.jaci.2013.06.038
40 Santos AF, Lack G. Basophil activation test: Food challenge in a test tube or specialist research tool? Clin Transl Allergy. 2016;6(1):10. 10.1186/s13601-016-0098-7
41 Dureuil A. Pharmaceutical industry: a difficult market to regulate. Industrie.com [online]. 2010 Dec 1. Available at: http://www.industrie.com
42 Rowe RC, Sheskey PJ, Quinn ME, editors. Handbook of Pharmaceutical Excipients, 6th ed. London, UK & Washington, DC: Pharmaceutical Press & American Pharmacists Association; 2009.
43 European Medicines Agency (EMA). Human regulatory – Marketing authorisation – Excipients labelling - EMA. [online]. Available at: http://www.ema.europa.eu
44 Sudheer PS, Hall JE, Read GF, Rowbottom AW, Williams PE. Flow cytometric investigation of perianaesthetic anaphylaxis using CD63 and CD203c. Anaesthesia. 2005;60:251–6. 10.1111/j.1365-2044.2004.04086.x
2 Lei Y, Gregory JA, Nilsson GP, Adner M. Insights into mast cell functions in asthma using mouse models. Pulm Pharmacol Ther. 2013;26:532–9. 10.1016/j.pupt.2013.03.019
3 Johansson SGO, Hourihane JO’B, Bousquet J, Bruijnzeel-Koomen C, Dreborg S, Haahtela T, et al. A revised nomenclature of allergy: An EAACI position statement from the EAACI nomenclature task force. Allergy. 2001;56(9):813–824. 10.1034/j.1398-9995.2001.t01-1-00001.x
4 Holgate ST. The role of mast cells and basophils in inflammation. Clin Exp Allergy. 2000;30:28–32. 10.1046/j.1365-2222.2000.00093.x
5 Buhring HJ, Simmons PJ, Pudney M, Foster SJ, van der Walt J, Morrison C, et al. The monoclonal antibody 97A6 defines a novel surface antigen expressed on human basophils and their multipotent and unipotent progenitors. Blood. 1999;94(7):2343–56.
6 Ebo DG, Hagendorens MM, Bridts CH, De Knop KJ, Stevens WJ. The basophil activation test in immediate drug allergy. Acta Clin Belg. 2009;64(2):129–35. 10.1179/acb.2009.022
7 Sainte-Laudy J, Boumediene A, Touraine F, Orsel I, Cogné M. Analysis of IgE downregulation induced by basophil activation. Application to the diagnosis of muscle relaxant allergic hypersensitivity by flow cytometry. Inflamm Res. 2006;55(Supp I): S21–2. 10.1007/s00011-005-0023-3
8 Garvey LH, Ebo D, Mertes PM, et al. An EAACI position paper on the investigation of perioperative immediate hypersensitivity reactions. Allergy. 2019;74:1872–84. 10.1111/all.13820
9 Moneret-Vautrin DA, Sainte-Laudy J, Kanny G, Fremont S. Human basophil activation measured by CD63 expression and LTC4 release in IgE-mediated food allergy. Ann Allergy Asthma Immunol. 1999;82:33–40. 10.1016/S1081-1206(10)62657-9
10 Cozon G, Ferrandiz J, Peyramond D, Brunet J. Detection of activated basophils using flow cytometry for diagnosis in atopic patients. Allergol Immunopathol (Madr). 1999;27:182–7.
11 Hauswirth AW, Natter S, Ghannadan M, Wolf M, Krauth MT, Valenta R, Sperr WR. Recombinant allergens promote expression of CD203c on basophils in sensitized individuals. J Allergy Clin Immunol. 2002;110:102–9. 10.1067/mai.2002.125257
12 Paris-Kohler A, Demoly P, Persi L, Lebel B, Bousquet J, Arnoux B. In vitro diagnosis of cypress pollen allergy by using cytofluorimetric analysis of basophils (Basotest). J Allergy Clin Immunol. 2000;105:339–45. 10.1016/S0091-6749(00)90085-X
13 Abuaf N, Rajoely B, Ghazouani E, Bricard H, Nicolas JF, Nguyen QT. Validation of a flow cytometric assay detecting in vitro basophil activation for the diagnosis of muscle relaxant allergy. J Allergy Clin Immunol. 1999;104:411–8. 10.1016/S0091-6749(99)70386-6
14 Sanz ML, Gamboa PM, Antepara I, Fernández J, Gastaminza G, Sanz ML. Flow cytometric basophil activation test by detection of CD63 expression in patients with immediate-type reactions to betalactam antibiotics. Clin Exp Allergy. 2002;32:277–86. 10.1046/j.1365-2222.2002.01305.x
15 Kahlert H, Cromwell O, Fiebig H. Measurement of basophil-activating capacity of grass pollen allergens, allergoids and hypoallergenic recombinant derivatives by flow cytometry using anti-CD203c. Clin Exp Allergy. 2003;33:1266–72. 10.1046/j.1365-2222.2003.01756.x
16 Pâris-Köhler A, Demoly P, Persi L, Lebel B, Bousquet J, Arnoux B. In vitro diagnosis of cypress pollen allergy by using cytofluorimetric analysis of basophils (Basotest). J Allergy Clin Immunol. 2000;105:339–45. 10.1016/S0091-6749(00)90085-X
17 Bühring H-J, Simmons PJ, Pudney M, Müller R, Jarrossay D, van Agthoven A, et al. The monoclonal antibody 97A6 defines a novel surface antigen expressed on human basophils and their multipotent and unipotent progenitors. Blood. 1999;94:2343–56.
18 Buhring HJ., Streble A., Valent P. The basophil-specific ectoenzyme E-NNP3 (CD203c) as a marker for cell activation and allergy diagnosis. Int Arch Allergy Immunol. 2004;133:317–29. 10.1159/000077351
19 Mac Glashan Jr D. Expression of CD203c and CD63 in human basophils: Relationship to differential regulation of piecemeal and anaphylactic degranulation processes. Clin Exp Allergy. 2010 Sep;40(9):1365–77. 10.1111/j.1365-2222.2010.03572.x
20 Ene-Cocis M, Petrisor C, Sfichi M, et al. CD63 vs. CD203c for the diagnosis of neuromuscular blocking agents immediate-type hypersensitivity. Human Vet Med. 2020;12:65–9.
21 Santos AF, Douiri A, Bécares N, Wu SY, Stephens A, Radulovic S, et al. Basophil activation test discriminates between allergy and tolerance in peanut-sensitized children. J Allergy Clin Immunol. 2014;134:645–52. 10.1016/j.jaci.2014.04.039
22 Gernez Y, Waters J, Mirković B, Lavelle GM, Dunn CE, Davies ZA, et al. Blood basophil activation is a reliable biomarker of allergic bronchopulmonary aspergillosis in cystic fibrosis. Eur Res J. 2016;47:177–85. 10.1183/13993003.01068-2015
23 Varshney P, Jones SM, Scurlock AM, Perry TT, Kemper A, Steele P, et al. A randomized controlled study of peanut oral immunotherapy: Clinical desensitization and modulation of the allergic response.J Allergy Clin Immunol. 2011;127(3):654–60. 10.1016/j.jaci.2010.12.1111
24 Eržen R, Košnik M, Silar M, Korošec P. Basophil response and the induction of a tolerance in venom immunotherapy: A long-term sting challenge study. Allergy. 2012;67(6):822–30. 10.1111/j.1398-9995.2012.02817.x
25 Peck G, Hashim MJ, Shaughnessy C, Muddasani S, Elsayed NA, Fleischer AB. Global epidemiology of urticaria: Increasing burden among children, females and low-income regions. Acta Derm Venereol. 2021;101:adv00433.
26 Darragi MK, Daghfous H, Ben Mansour A, Slim A, Ben Saad S, Tritar F. Investigation of drug hypersensitivity in Tunisia. Rev Fr Allergol. 2023;63(3):103539. 10.1016/j.reval.2023.103539
27 Ghozzi H, Sahnoun Z, Hammami S, Chaabane N, Ben Hassine L. Adverse effects due to antibiotics: a retrospective study over three years at the Regional Pharmacovigilance Center of Sfax. Rev Tun Infectiologie. 2011;5(1):16–21.
28 Amsler E, Soria A. Hypersensitivity reactions to beta-lactam antibiotics. Rev Med Interne. 2017;38(11):737–748. 10.1016/j.revmed.2017.06.020
29 Ponvert C, LeBourgeois M, Scheinmann P, Paupe J. Allergic reactions to commonly used anti-infective drugs. Rev Fr Allergol Immunol Clin. 1999;39(5):455–467.
30 Mayorga C, Çelik GE, Pascal M, Campana R, de Chaisemartin L, Netchiporouk E, Klossek JM, et al. Flow-based basophil activation test in immediate drug hypersensitivity. Allergy. 2024;79(3):580–600.
31 Knol EF, Mul FP, Jansen H, Calafat J, Roos D. Monitoring human basophil activation via CD63 mono-clonal antibody 435. J Allergy Clin Immunol. 1991;88:328–38. 10.1016/0091-6749(91)90094-5
32 Hoffmann HJ, Santos AF, Mayorga C, Eberlein B, Heeringa M, de Weck AL, et al. The clinical utility of basophil activation testing in diagnosis and monitoring of allergic disease. Allergy 2015;70:1393–405. 10.1111/all.12698
33 Fernandez TD, Mayorga C, Salas M, Doña I, Torres MJ, Blanca M. Evolution of diagnostic approaches in betalactam hypersensitivity. Expert Rev Clin Pharmacol. 2017;10:671–83. 10.1080/17512433.2017.1313110
34 McGowan EC, Saini S. Update on the performance and application of basophil activation tests. Curr Allergy Asthma Rep. 2013;13:101–9. 10.1007/s11882-012-0324-x
35 Sturm GJ, Kranzelbinder B, Sturm EM, Heinemann A, Groselj-Strele A, Aberer W. The basophil activation test in the diagnosis of allergy: Technical issues and critical factors. Allergy. 2009;64:1319–26. 10.1111/j.1398-9995.2009.02004.x
36 Cañas JA, Pérez-Sánchez N, Lopera-Doblas L, Moreno Aguilar C, Torres MJ, Blanca M, et al. Basophil activation test utility as a diagnostic tool in LTP allergy. Int J Mol Sci. 2022;23:4979. 10.3390/ijms23094979
37 Ebo DG, Bridts CH, Hagendorens MM, De Clerck LS, Stevens WJ. Basophil activation test by flow cytometry: Present and future applications in allergology. Cytomet B Clin Cytomet. 2008;74B:201–10. 10.1002/cyto.b.20419
38 Ebo DG, Sainte-Laudy J, Bridts CH, Mertens CH, Hagendorens MM, De Clerck LS, et al. Flow-assisted allergy diagnosis: Current applications and future perspectives. Allergy. 2006;61:1028–39. 10.1111/j.1398-9995.2006.01039.x
39 MacGlashan DW Jr. Basophil activation testing. J Allergy Clin Immunol. 2013 Oct;132(4):777–87. 10.1016/j.jaci.2013.06.038
40 Santos AF, Lack G. Basophil activation test: Food challenge in a test tube or specialist research tool? Clin Transl Allergy. 2016;6(1):10. 10.1186/s13601-016-0098-7
41 Dureuil A. Pharmaceutical industry: a difficult market to regulate. Industrie.com [online]. 2010 Dec 1. Available at: http://www.industrie.com
42 Rowe RC, Sheskey PJ, Quinn ME, editors. Handbook of Pharmaceutical Excipients, 6th ed. London, UK & Washington, DC: Pharmaceutical Press & American Pharmacists Association; 2009.
43 European Medicines Agency (EMA). Human regulatory – Marketing authorisation – Excipients labelling - EMA. [online]. Available at: http://www.ema.europa.eu
44 Sudheer PS, Hall JE, Read GF, Rowbottom AW, Williams PE. Flow cytometric investigation of perianaesthetic anaphylaxis using CD63 and CD203c. Anaesthesia. 2005;60:251–6. 10.1111/j.1365-2044.2004.04086.x
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Mar 1, 2026
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Mzoughi, R., Ben Houria, B., Tazeghdanti, A., Ben Azaiz, M., Kochkar, R., & Ghazouani, E. (2026). Advancing drug allergy diagnosis in Tunisia using flow cytometry: a promising first step. Allergologia Et Immunopathologia, 54(2), 99–108. https://doi.org/10.15586/aei.v54i2.1512
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How to Cite
Mzoughi, R., Ben Houria, B., Tazeghdanti, A., Ben Azaiz, M., Kochkar, R., & Ghazouani, E. (2026). Advancing drug allergy diagnosis in Tunisia using flow cytometry: a promising first step. Allergologia Et Immunopathologia, 54(2), 99–108. https://doi.org/10.15586/aei.v54i2.1512