Letter to the Editor Diagnostic Immunology Ann Lab Med 2016;36:498 http://dx.doi.org/10.3343/alm.2016.36.5.498 ISSN 2234-3806 • eISSN 2234-3814
Reply to the Letter to the Editor by Dr. Chirumbolo Dong Il Won, M.D. Department of Clinical Pathology, Kyungpook National University School of Medicine, Daegu, Korea
Dear Editor, We are grateful to Dr. Chirumbolo for the letter in response to our paper describing a basophil activation test for chronic urticaria [1, 2]. We agree with most of the issues raised by Dr. Chirumbolo, but some points need to be addressed. The non-atopic donors in our study were adults, not infants. Among 3-5 healthy individuals with blood group O, one with a high basophil count was selected for each experiment. The average basophil count was 0.55% by flow cytometric analysis, 0.75% by manual cell counting on a peripheral blood smear, and 0.81% by the complete blood count analyzer (data not shown in the article). Therefore, the purity of basophils in gate 3 appeared to be sufficiently high for our indirect flow basophil activation test. In order to retrieve as many cellular events as possible during flow cytometry, data acquisition was continued until non-cellular events appeared in the time/side scatter plot
(plot not shown in the article). The contaminating non-cellular events were excluded in the subsequent data analysis. Gate 3 for purified basophils was the intersection area of the CD123bright and CCR3pos populations. These two combined basophil identification markers may reciprocally help to exclude T lymphocytes (CD123neg) and monocytes (CCR3neg), even without additional exclusion markers. The sufficiently high purity of basophils in gate 3 was confirmed by the CD63 FITC vs. CD203c APC plot, where most events were either CD203cdim or CD203cbright.
Received: May 24, 2016 Revision received: May 24, 2016 Accepted: May 31, 2016
© The Korean Society for Laboratory Medicine.
Corresponding author: Dong Il Won Department of Clinical Pathology, Kyungpook National University School of Medicine, 130 Dongdeok-ro, Jung-gu, Daegu 41944, Korea Tel: +82-53-200-5291, Fax: +82-53-426-3367, E-mail: [email protected]
498 www.annlabmed.org
REFERENCES 1. Kim Z, Choi BS, Kim JK, Won DI. Basophil markers for identification and activation in the indirect basophil activation test by flow cytometry for diagnosis of autoimmune urticaria. Ann Lab Med 2016;36:28-35. 2. Chirumbolo S. Basophil activation test for chronic urticaria. Ann Lab Med 2016; accepted.
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
http://dx.doi.org/10.3343/alm.2016.36.5.498
Reply to the Letter to the Editor by Dr. Chirumbolo Dong Il Won, M.D. Department of Clinical Pathology, Kyungpook National University School of Medicine, Daegu, Korea
Dear Editor, We are grateful to Dr. Chirumbolo for the letter in response to our paper describing a basophil activation test for chronic urticaria [1, 2]. We agree with most of the issues raised by Dr. Chirumbolo, but some points need to be addressed. The non-atopic donors in our study were adults, not infants. Among 3-5 healthy individuals with blood group O, one with a high basophil count was selected for each experiment. The average basophil count was 0.55% by flow cytometric analysis, 0.75% by manual cell counting on a peripheral blood smear, and 0.81% by the complete blood count analyzer (data not shown in the article). Therefore, the purity of basophils in gate 3 appeared to be sufficiently high for our indirect flow basophil activation test. In order to retrieve as many cellular events as possible during flow cytometry, data acquisition was continued until non-cellular events appeared in the time/side scatter plot
(plot not shown in the article). The contaminating non-cellular events were excluded in the subsequent data analysis. Gate 3 for purified basophils was the intersection area of the CD123bright and CCR3pos populations. These two combined basophil identification markers may reciprocally help to exclude T lymphocytes (CD123neg) and monocytes (CCR3neg), even without additional exclusion markers. The sufficiently high purity of basophils in gate 3 was confirmed by the CD63 FITC vs. CD203c APC plot, where most events were either CD203cdim or CD203cbright.
Received: May 24, 2016 Revision received: May 24, 2016 Accepted: May 31, 2016
© The Korean Society for Laboratory Medicine.
Corresponding author: Dong Il Won Department of Clinical Pathology, Kyungpook National University School of Medicine, 130 Dongdeok-ro, Jung-gu, Daegu 41944, Korea Tel: +82-53-200-5291, Fax: +82-53-426-3367, E-mail: [email protected]
498 www.annlabmed.org
REFERENCES 1. Kim Z, Choi BS, Kim JK, Won DI. Basophil markers for identification and activation in the indirect basophil activation test by flow cytometry for diagnosis of autoimmune urticaria. Ann Lab Med 2016;36:28-35. 2. Chirumbolo S. Basophil activation test for chronic urticaria. Ann Lab Med 2016; accepted.
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
http://dx.doi.org/10.3343/alm.2016.36.5.498
