Journal of Clinical Otolaryngology Head and Neck Surgery

The Busan, Ulsan, Gyeoungnam Branch of Korean Society of Otolaryngology-Head and Neck Surgery

J Clin Otolaryngol Head Neck Surg 2007; 18(2):170-177

pISSN: 1225-0244, eISSN: 2713-833X

DOI: https://doi.org/10.35420/jcohns.2007.18.2.170

원저

자가면역성 감각신경성 난청 환자에서 냉동보관법을 이용한 내이 조직 항원에 대한 T세포 반응성 검사의 유용성

백무진¹^,^*

Efficacy of Enzyme Linked Immunospot Assay Using Cryopreserved Cell in Detection of Inner Ear Antigen Specific T Cells in Patients with Autoimmune Sensorineural Hearing Loss

Moo Jin Baek¹^,^*

¹인제대학교 의과대학 부산백병원 이비인후과학교실

¹Department of Otolaryngology Head and Neck Surgery, College of Medicine, Inje University, Busan Paik Hospital, Busan, Korea

^*교신저자: 백무진, 614-735 부산광역시 부산진구 개금동 633-165 인제대학교 의과대학 부산백병원 이비인후과학교실 전화： (051) 890-6311·전송： (051) 892-3831 E-mail： mjbaek@ijnc.inje.ac.kr

© Copyright 2007 The Busan, Ulsan, Gyeoungnam Branch of Korean Society of Otolaryngology-Head and Neck Surgery. 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.

Received: Aug 18, 2007; Accepted: Oct 19, 2007

Published Online: May 31, 2020

ABSTRACT

Objective: Cryopreservation is very useful method for long term storage of various samples. Author tried to use the cryopreservation technique with ELISPOT assay to exam the T cell responsiveness to inner ear specific antigen (Cochlin) and evaluate its efficacy. Methods: Isolated peripheral blood mononuclear cells (PBMC) were suspended with freezing medias (A & B) in cryotube. After pre-chilling for 24 hours, transferred to liquid nitrogen tank. Cell viability and functionality were evaluated by viable cell count with trypan blue stain, proliferation assay with methyl-³H thymidine and the frequencies of IFN- γ and IL-5 producing T cells using ELISPOT assay with anti CD3 antibody before and after freezing. And also compared the frequencies of cytokine producing T cells in response to human Cochlin using ELISPOT assay with frozen cells between patient and control subjects. Results: The rate of viable cell count and stimulation index of feezed cells showed about 75%, 70% respectively when compared with before freezing cells. The frequencies of IFN- γ and IL-5 producing T cells in response to anti CD3 were 68% and 71% of before freezing condition. And the frequencies of IFN- γ and IL-5 producing T cells response to human recombinant cochlin were significantly higher in patients than control subjects using cryopreserved cell. Conclusion: The functional responsiveness and cell number of cryopreserved cells were restored nearly before freezing level. So the cryopreservation technique could be very useful tool in ELISPOT assay for determination of the frequencies of cytokines. (J Clinical Otolaryngol 2007;18:170–177)

Keywords: 냉동보관법; 자가면역성 감각신경성 난청; ELISPOT assay

Keywords: Cryopreservation; Autoimmune sensorineural hearing loss; ELISPOT assay