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1 대한수혈학회지 : 제 29 권제 2 호, 2018 The Korean Journal of Blood Transfusion Vol. 29, No. 2, , August pissn eissn Original Article 한국인헌혈자에서의 -D-/-D- 표현형빈도 이미경 1, * ㆍ정성이 1, * ㆍ김진욱 2 ㆍ김종필 1 ㆍ김동현 1 ㆍ박정란 1 대한적십자사중앙혈액검사센터 1, 대한적십자사혈액관리본부 2 -D-/-D- Phenotype Frequency among Korean Donors Mi Kyung Lee 1, *, Sung Yi Jung 1, *, Jin Uk Kim 2, Jong Phil Kim 1, Dong Hyun Kim 1, Jung Ran Park 1 Korean Red Cross Central Blood Laboratory Center 1, Seoul, Korean Red Cross Blood Services 2, Wonju, Korea Background: To improve Rh-related antigen negative blood supply effectively, the Korean Red Cross (KRC) blood centers have performed Rh phenotype screening tests of C, c, E and e antigens for all donors since April, Especially for rare -D-/-D- blood supply and donor recruitment, we have implemented Rh phenotype confirmation test for all C, c, E and e antigen negative donors. In this study, we report the test results of 7 donors with -D-/-D- phenotype. Methods: All three KRC Blood Laboratory Centers performed Rh phenotype screening tests using the automatic machine, PK7300 (Beckman Coulter, Japan), for all 876,920 donors from January 1, 2018 to April 30, We then performed the Rh phenotype confirmation test using the tube method manually, at room temperature, 37 and antihuman globulin phase. Results: Among 876,920 donors, 14 were Rh antigen C, c, E, e negative as results of Rh phenotype screening test. The results of Rh phenotype confirmation test of these 14 donors showed that 7 donors were Rh antigen C, c, E, e negative. The ratio of -D-/-D- phenotype for all donors was %. Conclusion: Our data suggests that -D-/-D- phenotype is one of the rare blood groups among Koreans. Although -D-/-D- phenotype was confirmed by serologic tests, it is necessary to re-confirm it by molecular genetic techniques. (Korean J Blood Transfus 2018;29: ) Key words: -D-/-D-, Rh phenotype, rare blood 서론 Rh 혈액형군은사람적혈구의가장복잡한항원군으로서, D, C, c, E, e 항원을포함하여현재까 지약 50여종의항원과변이형이알려져있다 [1]. 이중가장면역원성이강하고임상적으로중요한것은 D 항원으로 RhD 검사는중요한수혈전검사의필수항목이다. 그외항원은환자가감작 Received on July 11, Revised on July 27, Accepted on August 2, 2018 Correspondence to: Mi Kyung Lee Korean Red Cross Central Blood Laboratory Center, 18 Gonghang-daero 69 gil, Gangseo-gu, Seoul 07559, Korea Tel: , Fax: , mklee@redcross.or.kr, ORCID: *Mi Kyung Lee and Sung Yi Jung contributed equally as co-first authors. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License ( which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Copyright C 2018 The Korean Society of Blood Transfusion

2 이미경외 : 한국인헌혈자에서의 D-/-D- 표현형빈도 되어해당항원에대한항체 ( 예, 항 E+c) 가발생할경우해당항원이없는혈액을선택하여수혈해야한다. 그간의료기관으로부터대한적십자사중앙혈액원에의뢰된특정항원음성혈액요청건수를살펴보면약 46.6% 가 Rh항원 (C, c, E, e) 음성혈액공급과관련되어있다 [2]. Rh항원 (C, c, E, e) 음성혈액의원활한공급을위해대한적십자사혈액원에서는모든헌혈자에대하여 2013년 4월부터 Rh 표현형선별검사를실시하였다 (Table 1). 그런데, RhD 항원이외의 Rh항원 (C, c, E, e) 이모두존재하지않은혈액형이매우드물지만존재하는데, 이는 -D-/-D-형이다[3,4]. 이들혈액형을가진환자는 D 항원이존재하기때문에의료기관에서는환자가 Rh항원 (C, c, E, e) 에대한항체가발생하기전까지는일반수혈전검사에서는검출할수없다. 이혈액형이임상적으로중요한이유는해당항체가발생하면동일한 -D-/-D-형만수혈할수있고, 그외혈액은수혈시대부분심각한용혈이발생하기때문이다. 그래서, 일부나라에서는국가사업으로희귀혈액형등록사업을실시하여 -D-/-D-형과같이희귀한혈액형을미리검출하여헌혈자를확보하고있다. 그러나그간국내 -D-/-D-형은산발적인증례보고 [5-7] 만존재하고, 전체인구에서그빈도는보고되지않았다. 이에본연구에서는 Rh 표현형선별검사결과 Table 1. History of Rh phenotype screening test in Korean Red Cross Blood Laboratory Centers Test start date Rh antigens C, e C, E, e C, c, e C, c, E, e C, c, E, e 항원이모두음성인헌혈자를대상으로 Rh 표현형확인검사를수기법으로실시하여그빈도를구하여국내최초로이를보고하고자하였다. 대상및방법 1. 대상본연구는대한적십자사혈액관리본부생명윤리위원회의승인을받아 2018년 1월 1일부터 2018년 4월 30일까지대한적십자사중앙, 중부및남부혈액검사센터에서검사한모든헌혈자를대상으로실시하였다. Rh 표현형선별검사는총 876,920명의헌혈자를대상으로실시하였으며이중 Rh 표현형선별검사결과 C(-), c(-), E(-), e(-) 인헌혈자에대하여 Rh 표현형확인검사를실시하여위음성결과를배제하였다. 2. 검사방법 Rh 표현형선별검사는자동화마이크로플레이트시스템인 PK7300 (Beckman Coulter, Shizuoka-ken, Japan) 을이용하였다. 검사시약은 Anti-C (DIAGAST, Avenue Eugene Anivee, France), Anti-c (DIAGAST), Anti-E (DIAGAST), Anti-e (DIAGAST) 를사용하였으며적정희석배수를적용하였다. Rh 표현형확인검사는 Rh 표현형선별검사에서 C(-), c(-), E(-), e(-) 인헌혈자에대하여 Rh 표현형선별검사시와동일한시약을원액으로사용하여수기법인 tube법으로실시하였다. 대조용 tube에 22% bovine albumin과검체 (2 5% 혈구부유액 ) 을각각 1 drop씩넣고, 검사용 tube에는각각 Rh 항원별항혈청과검체 (2 5% 혈구부유액 ) 을각각 1 drop씩넣었다. 혈청원심분리기를사용하여 1,000 g에서 15초간원심한후응집유무를

3 Korean J Blood Transfus Vol. 29, No. 2, , Aug 확인했다. 이때대조용 tube가음성이고검사용 tube에서하나라도 1+이상응집을보이면양성으로판정하고검사를종료하였다. 모든 tube가음성일경우 tube내의검체와시약을잘혼합한후 37 o C 항온수조에서 30분간항온하였으며, 원침한후응집유무를확인하였다. 0.9% saline으로 3회세척한후대조용및검사용 tube에항인글로불린시약을 2 drop씩넣고잘혼합하였다. 원심후응집유무를확인하였으며, 육안판정이어려울경우현미경을이용하였다. 검사결과대조용과검사용 tube가모두음성인경우 IgG 감작적혈구를 1 drop씩넣고잘혼합한후원심후응집유무를육안으로확인하였다. 이때응집을보이면최종음성으로판정하고검사를종료하였으며, 응집이없으면재검사를수행하였다 (Table 2). 결과대한적십자사혈액검사센터에서는헌혈자 876,920명을대상으로 Rh 표현형선별검사를실시하였으며그결과 14명이 C(-), c(-), E(-), e(-) 였다 (Table 3). 이 14명에대하여 Rh 표현형확인검사를실시한결과 CDe 가 5명, cde 가 2명, C(-), c(-), E(-), e(-) 인 -D-/-D- 표현형은 7명으로확인되 어그빈도가 % 임이확인되었다 (Table 4). 선별검사와확인검사결과가불일치했던 7명은해당항원이확인검사실온단계에서 2+이하로약하게발현하는양상을보였다. 고찰 -D-/-D- 표현형은 RHCE 유전자의결핍으로인해 C, c, E, e 항원이없이 D 항원만을강하게발 Table 3. Result of Rh phenotype screening test performed by PK7300 Rh phenotype No. donors % CDe 364, CcDEe 330, cde 78, CcDe 64, cdee 29, CDEe 2, CcDE cde 1, CDE D-/-D Rh(-) 3, Total donor no. 876, Table 2. Results of Rh phenotype confirmation test Control tube result Testing tube result Room temperature phase AHG phase Coombs-control IgG phase Results Not determined* Negative - - ± 4+ Not tested Positive - ± 4+ Not tested Not tested Positive ± 4+ ± 4+ Not tested Not tested Not determined *retest; perform direct antiglobulin test. Abbreviation: AHG, antihuman globulin

4 이미경외 : 한국인헌혈자에서의 D-/-D- 표현형빈도 Table 4. Result of Rh phenotype confirmation test for 14 donors performed by standard tube tests Rh phenotype No. donors % of total donors (876,920) CDe cde D-/-D 현하는것을특징으로하는희귀한혈액형이며백인, 아시아인, 히스패닉, 아프리카등다양한인종에서보고되고있다 [3,4,8]. 그러나국내에서는 1998년이후 3건의사례 [5-7] 외에는 -D-/-D- 표현형빈도에대한연구보고는없었다. 국내 -D-/-D- 표현형빈도에대한보고로서는대한적십자사혈액원의헌혈자총 876,920명을대상으로한본연구가최초이며 4개월간의비교적짧은연구기간이었음에도불구하고 Okubo 등 [9] 이일본헌혈자 692,000명중 7명의 D-/-D- 표현형발견사례를보고한일본인헌혈자에서의 -D-/-D- 표현형빈도와유사한결과를보여주었다. 지연성용혈성수혈반응을초래하는항체로 Rh 항체및 Kidd, Duffy, Kell 항체등이알려져있으며이로인해수혈을받은환자가사망에이른보고가있었다. 그래서만성적수혈이예상되는환자에게 ABO, RhD 이외의혈액형항원음성혈액을우선적으로공급하는방안이고려되고있다 [10,11]. 특히 -D-/-D- 표현형을가진사람은임신이나수혈에의해감작될경우, CcEe 항원복합체에대하여반응하는항체인항 Hro(Rh17) 이종종형성되며, 산모의혈청에존재하는항 Hro는다양한임상상의신생아용혈성질환을유발할수있다 [12-14]. 이처럼 -D-/-D- 표현형을가진수혈자는결핍된유전자에대한면역항체가생길수있기때문에 -D-/-D- 표현형혈액을수혈받아야하므로 -D-/-D- 표현형을가진헌혈자관리가매우 중요하다. 우리나라에서실제 -D-/-D- 표현형을가진환자에대한수혈사례는두차례로서 2004 년과 2017년에지면을통해알려진바있는데두사례모두일본적십자사에보관된냉동적혈구를공급받아수혈한사례로우리나라에서도희귀혈액형관리에대한필요성이제기된바있다 [2]. 대한적십자사혈액원에서는현재 rare blood donor registry program을통해다양한희귀혈액형헌혈자를확보하고자노력하고있으며 [15] 본연구결과가도움이되리라생각한다. 본연구는혈청학적검사법으로만진행되어적혈구에적은양의항원만발현하는경우는검출하기어려운한계를가지고있다 [16]. Rh 항원의경우이미그다양성으로인해표현형 (phenotype) 과유전자형 (genotype) 의불일치에대해서국내외에서많은연구가있어왔고 [17-19] 특정항원음성혈액검사의경우에도최근외국에서는많은수의헌혈자를대상으로대량으로검사할수있는유전형검사시스템이도입되고있다 [20,21]. 본연구에서도선별검사와확인검사결과가불일치했던 7명 (CDe 5명, cde 2명 ) 은해당항원이확인검사실온단계에서 2+이하로약하게발현하는양상을보여항원성이약할경우에는선별검사에서위음성의가능성이있음을보여주었다. 따라서국내헌혈자에있어서 -D-/-D- 표현형에대한보다더정확하고유용한정보와통계를얻기위해서는유전학적검사를포함한추가연구가필요하다고생각한다. 요약배경 : Rh 항원 (C, c, E, e) 음성혈액의원활한공급을위해대한적십자사혈액원에서는 2013년 4 월부터모든헌혈자에대하여 Rh 표현형선별검

5 Korean J Blood Transfus Vol. 29, No. 2, , Aug 사를시행하여왔고희귀혈액형 (-D-/-D-) 공급및헌혈자확보를위해 2018년 1월 1일부터는 Rh 표현형선별검사결과 C, c, E, e 항원이모두음성인헌혈자를대상으로 Rh 표현형확인검사를수기법으로실시하고있다. 본연구에서는 Rh 표현형확인검사결과 -D-/-D- 표현형 7명이발견되어이를보고하고자한다. 방법 : 2018년 1월 1일부터 4월 30일 ( 헌혈일기준 ) 까지대한적십자사 3개혈액검사센터에서검사한모든헌혈자 876,920명을대상으로자동화장비 PK7300 을사용하여 Rh 표현형선별검사를시행하였고선별검사결과 C(-), c(-), E(-), e(-) 인헌혈자를대상으로 tube법을이용하여수기로 Rh 표현형확인검사를하였다. 결과 : Rh 표현형선별검사에서 876,920명중 14명이 C(-), c(-), E(-), e(-) 의결과를보였다. 이 14명에대하여 Rh 표현형확인검사를실시한결과 7명이 C(-), c(-), E(-), e(-) 인 -D-/-D- 표현형으로확인되었으며, 전체헌혈자대비 -D-/-D- 표현형의비율은 % 였다. 결론 : -D-/-D- 표현형은한국인의매우그빈도가낮은희귀혈액형중하나임이확인되었다. 비록혈청학적방법에의해 -D-/-D- 표현형이확인되었지만향후유전학적검사를추가로실시하여재확인이필요할것이다. 감사의글본연구를위해많은도움을주신조덕교수님 ( 성균관의대삼성서울병원 ) 께감사의뜻을표합니다. References 1. Fung MK, American Association of Blood Banks. Technical manual. 18th ed. Bethesda, Md: American Association of Blood Banks, 2014: Lee MK, Choi BC, Oh DJ, Jeong OJ, Kim MJ. A study to establish rare blood type donor registration system. Korean J Blood Transfus 2005;16: Daniels G. Human blood groups. 2nd ed. Malden, MA: Blackwell Science, Kemp TJ, Poulter M, Carritt B. A recombination hot spot in the Rh genes revealed by analysis of unrelated donors with the rare D--phenotype. Am J Hum Genet 1996;59: Whang DH, Kim HC, Hur M, Choi JH, Park JS, Han KS. A successful delivery of a baby from a D--/D-- mother with strong anti- Hr0. Immunohematology 2000;16: Hwang JY, Suh IB, Shim JY, Cho JS, Lee DH. A case of fetal hydrops associated with maternal -D-/-D- phenotype. Korean J Obstet Gynecol 2006;49: Um TH, Cho CR, Whang JH, Whang DH, Woon MS, Han KS. A case of D-/Dphenotype associated with moderate hemolytic disease of the newborn. Korean J Blood Transfus 2007;18: Westhoff CM, Vege S, Nickle P, Singh S, Hue-Roye K, Lomas-Francis C, et al. Nucleotide deletion in RHCE*cE (907delC) is responsible for a D--haplotype in Hispanics. Transfusion 2011;51: Okubo Y. Pretransfusion testing with special reference to blood groups in Japanese. Tokyo: Ishiyaku Shuppan, 1991: Kim S, Hur M, Lee KM, Chun W. A case of delayed hemolytic transfusion reaction in a patient with anti-c, anti-e, and anti-jk(b). Korean J Blood Transfus 2009;20: Kim KH, Kim BR, Choi JL, Woo KS, Kim JM,

6 이미경외 : 한국인헌혈자에서의 D-/-D- 표현형빈도 Ham JY. Difference of Rh phenotype between irregular antibody positive patients and RhD positive population in Korea. Korean J Blood Transfus 2014:25: Han KS, Park KU, Song EY. Transfusion medicine. 4th ed. Seoul: Korea Medical Book Publisher, 2014: Fung MK, American Association of Blood Banks. Technical manual. 18th ed. Bethesda, Md: American Association of Blood Banks, 2014: Salamat N, Bhatti FA, Hussain A, Ziaullah. Anti-Rh17 (anti-hr0): a rare diagnostic and management problem. J Pak Med Assoc 2004;54: Lim AH, Kim TE, Park MH, Kim YB, Min HK. Study for the establishment of donor-based rare blood management system. Korean J Blood Transfus 2018;29(S):S Chang HE, Hwang SM, Hong YJ, Han M, Park JS, Park KU, et al. Genotyping for RhD and RhCEce antigens using free circulating nucleic acids in plasma and serum. Korean J Blood Transfus 2014;25: Tax MG, van der Schoot CE, van Doom R, Douglas-Berger L, van Rhenen DJ, MaaskantvanWijk PA. RHC and RHc genotyping in different ethnic groups. Transfusion 2002;42: Yang HS, Lee MY, Park TS, Cho SY, Lee HJ, Lim G, et al. Primary anti-d alloimmunization induced by Asian type RHD (c.1227g>a) DEL red cell transfusion. Ann Lab Med 2015;35: Flegel WA, von Zabern I, Wagner FF. Six years' experience performing RHD genotyping to confirm D- red blood cell units in Germany for preventing anti-d immunizations. Transfusion 2009;49: Döscher A, Vogt C, Bittner R, Gerdes I, Petershofen EK, Wagner FF. RHCE alleles detected after weak and/or discrepant results in automated Rh blood grouping of blood donors in Northern Germany. Transfusion 2009;49: Flegel WA, Gottschall JL, Denomme GA. Implementing mass-scale red cell genotyping at a blood center. Transfusion 2015;55:2610-5; quiz

석사논문.PDF

석사논문.PDF ABO Rh A study on the importance of ABO and Rh blood groups information in Public Health 2000 2 1 ABO Rh A study on the importance of ABO and Rh blood groups information in Public Health 2000 2 2 ABO Rh