Korean J Lab Med 2010;30: DOI /kjlm Original Article Diagnostic Immunology Detection of Allergen Specific IgE by AdvanSure

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1 Korean J Lab Med 2010;30:42031 DOI /kjlm Original Article Diagnostic Immunology Detection of Allergen Specific IgE by AdvanSure Allergy Screen Test EunJee Oh, M.D., SunAh Lee, M.D., Jihyang Lim, M.D., YeonJoon Park, M.D., Kyungja Han, M.D., and Yonggoo Kim, M.D. Department of Laboratory Medicine, School of Medicine, the Catholic University of Korea, Seoul, Korea Background : In vitro serum allergenspecific IgE tests have been routinely used in the clinical diagnosis of allergic diseases. We evaluated the clinical usefulness of a newly introduced multiple antigen screen test, Advansure Allergy Screen (LG Life Science, Korea) (LGScreen) for the detection of allergen specific IgE. Methods : A total of 180 sera (80 for inhalant and 100 for food panels) were tested by LGScreen and RIDA Allergy Screen (Rbiopharm, Germany) (RIDAScreen) assays. According to the 5860 specific allergens or allergen groups, the positive rates and agreement rates were analyzed using the cut off levels of class 2. For the quantitation of total IgE and specific IgE, nephelometry and ImmunoCAP test were performed in the sera showing discrepant results between the two allergy screen assays. Results : The agreement rate and kappa value (k) of total IgE between the two allergy screen assays was 73.9% and LGScreen showed higher agreement rate with nephelometry than RIDAScreen. The positive rates to common outdoor inhalant and food allergens were significantly higher in RIDAScreen. Overall agreement rate of specific IgE between the two allergy screen assays for 58 allergens was 86.7% (6,086/7,020) (k, 0.293). In samples showing discrepant results between the two allergy screen assays, concordance rate of allergy screen assay with ImmunoCAP assay was 70.9% (449/633) for LGScreen (k, 0.585) and 29.1% (184/633) for RIDAScreen (k, 0.303). Conclusions : LGScreen showed a favorable agreement with RIDAScreen and ImmunoCAP assays, and it could be used for the detection of allergen specific IgE in the clinical laboratory. (Korean J Lab Med 2010;30:42031) Key Words : Allergens, Specific IgE assay, Allergy screen test, ImmunoCAP assay 서 혈청알레르겐특이 IgE 검사는알레르기질환의 1차적선별검사로이용되며피부검사와동시에검사되기도한다. 피부시험에비해혈청알레르겐특이 IgE 검사는환자에게위험도가적고, 투여약물의영향을받지않으며, 영유아나심한피부질환 Received : April 29, 2010 Manuscript No : KJLM10083 Revision received : June 28, 2010 Accepted : July 23, 2010 Corresponding author : Yonggoo Kim, M.D. Department of Laboratory Medicine, Seoul St. Mary's Hospital, 505 Banpodong, Seochogu, Seoul , Korea Tel : , Fax : yonggoo@catholic.ac.kr *This work was supported by the Industrial Core Technology Development Program funded by the Ministry of Knowledge Economy (No ). ISSN 론 The Korean Society for Laboratory Medicine 이있는환자에게도적용이가능하다. 또한검사결과의특이성이높으며정량적인결과보고가가능한장점이있다 [13]. 혈청알레르겐특이 IgE 검사는검사원리에따라화학발광면역측정법, 형광효소면역측정법 (fluroenzyme immunoassay), 면역블롯법 (immunoblot), 면역크로마토그래피법, 방사선면역측정법등으로분류할수있고, 검사결과형태에따라정성, 반정량, 정량검사로분류할수있다. 선별검사로는여러가지항원을동일한고체상에결합하여다양한종류의알레르겐특이 IgE 항체를동시에반정량적으로검출하는면역블롯법이흔히이용된다. 국내보고에의하면반정량면역블롯법검사를위한시약으로서 MAST CLA (Hitachi Chemical Diagnostics Inc, Mountain View, CA, USA), RIDA Allergy Screen (Rbiopharm, Darmstadt, Germany), Polycheck Allergy (Biocheck Co, Munster, Germany) 등이사용되고있으며, 한종목씩검사가가능한정량검사로는 ImmunoCAP (Phadia, Uppsala, 420

2 Oh EJ, et al., AdvanSure Allergy Screen Test 421 Sweden) 이일반적으로사용되고있다 [4]. 최근국내회사인 LG 생명과학에서혈청의알레르겐특이 IgE 검출을위한면역블롯법의선별검사인 AdvanSure Allergy Screen (LG Life Science, Seoul, Korea)( 이하 LGScreen) 시약을개발하여소개하였으나현재까지평가연구에대한보고는없다. 저자들은임상검체를대상으로, 국내임상검사실에서높은빈도로사용되고있고알레르겐항원판넬구성이비슷한 RIDA Allergy Screen (RIDAScreen) 검사와 LGScreen 검사의결과를비교하고불일치검체를 ImmunoCAP 알레르겐특이 IgE 정량검사로확인하여 LGScreen 검사의임상적유용성을평가하였다. 대상및방법 1. 대상검체본원에알레르기질환이의심되어 allergy screen 검사가의뢰된검체가운데 RIDAScreen 검사에서한가지알레르겐에서라도 class 2 이상의양성이검출된 180명의혈청을대상으로하였다. 180명의연령분포는 173세였고, 남자 106명, 여자 74명을포함하였다. 동반된알레르기주질환은비염 88예, 인후염 22예, 천식 7예, 아토피 16예, 두드러기등피부질환 37예, 기타 10예였으며, 알레르기질환으로진단된환자는 56명이었다. 180예중 80 예는흡입형알레르겐패널로, 100예는음식형알레르겐패널로검사하였다. 혈청검체는 RIDAScreen 시약으로검사후 24 시간이내에분주하여 70 에냉동보관하였으며, 해동하여 LGScreen 검사를시행하였다. LGScreen 과 RIDAScreen 시약에공통으로포함된알레르겐 58종에대하여결과를분석하고일치율을구하였다. LGScreen 검사와 RIDAScreen 검사의총 IgE 또는알레르겐특이 IgE 결과를비교하여두검사간불일치를보이는검체를대상으로, 총 IgE 는면역비탁법으로재검하였고, 알레르겐특이 IgE는 Immuno CAP 방법으로재검하였다. 본연구는본원임상시험심사위원회의심의를통과하였다 ( 과제번호 ; KC08FZZZ0317). 2. 혈청총 IgE 검사 Allergy screen 검사에서제시되는총 IgE의정성적결과를 100 IU/mL 이상과미만으로구분하여결과를비교하고, 두가지 allergy screen 검사결과에불일치를보인검체를대상으로 N Latex IgE mono (Dade Behring Inc., Newark, DE, USA) 시약과 BN II system (Siemens Healthcare Diagnostics Inc., Deerfield, IL, USA) 을이용하여면역비탁법으로측정하였다. 면역비탁법검사의측정가능범위는 IU/mL, interassay CV %, intraassay CV % 였다. 제조회사에서제시한참고범위는성인의경우 100 IU/mL 미만이며, 본연구에서도 cutoff 100 IU/mL를기준으로 allergy screen 결과와정성적으로비교하였다. 3. AdvanSure Allergy Screen (LGScreen) 검사흡입형및음식형시약은각각 A, B스트립으로구성되어있으며각스트립에는 21가지의항원이부착되어있다 (Table 1). 흡입형과음식형검사시약으로각각 41종의알레르겐에대한특이 IgE와총 IgE 검출이가능하며, 총 IgE 측정결과는 <100 IU/mL, 100 IU/mL로구분되어얻어진다. LGScreen 검사는 RIDAScreen 검사에서검출가능한 39종알레르겐특이항체이외에환삼덩굴 (Japanese hop) 과고등어 (mackerel) 알레르겐에대한특이 IgE 항체의검출이가능하다 (Table 1). Profiblot 48 (Tecan Group Ltd., Mannedorf, Switzerland) immunoblot 장비를이용하였고, 각스트립에혈청 50 ml과혈청희석액 250 ml를넣고 45분동안실온에서반응시킨후세척하고, biotin이표지된항 IgE 시약 250 ml를첨가하고 30분간실온반응후세척하였다. 효소접합용액 (streptavidinalkaline phosphatase) 250 ml를넣고실온에서 30분간반응시킨후세척하고, 발색용액 250 ml를첨가하여 20분간반응시켰다. 전용판독기인 AdvanSure AlloScan을이용하여 class 06으로판독하고, 시약설명서에알레르겐특이항체양 중간 으로제시되어있는 class 2 이상 (0.7 ku/l 이상 ) 을양성으로판정하였다. 매검사마다키트내양성대조군의결과를확인하였고, 모음혈청 (pooled sera) 으로검사하여 ±1 class의범위내에드는지확인하였다. 4. Allergy Screen (RIDAScreen) 검사흡입형및음식형시약은각각 A, B스트립으로구성되어있으며각스트립에는 20가지의항원이부착되어있다. Table 1 에기재된 Allergen code 는 rbiopharm 사에서제공한설명서에기재된코드이며, 코드번호가달라도알레르겐조성이동일한경우동일한알레르겐으로간주하여결과를비교하였다. 흡입형과음식형검사시약으로각각 39종의알레르겐특이 IgE와총 IgE 검출이가능하다. 총 IgE 측정결과는 <100 IU/mL,

3 422 Korean J Lab Med 2010;30:42031 Table 1. Tested allergens in LGScreen, RIDAScreen and ImmunoCAP assays Panel LGScreen RIDAScreen ImmunoCAP Code Allergens Code Allergens Code Allergens Inhalant F17 Hazelnut * * G1 Sweet vernal grass * * G2 Bermuda grass * * G3 Orchard grass * * G6 Timothy grass * * G7 Reed * * M1 Penicillium notatum * * T11 Sycamore mix T1 Sycamore maple T11 Sycamore mix T12 Sallow willow * * T14 Poplar mix * * T15 Ash mix * * T16 Pine * * T17 Japanese cedar T24 Cedar T17 Japanese cedar T19 Acacia T36 Acacia T19 Acacia W11 Russian thistle * * W12 Goldenrod * * W14 Pigweed * * W7 Oxeye daisy * * W8 Dandelion * * Food D70 Acarus siro * * F11 Buckwheat * * F13 Peanut * * F25 Tomato * * F26 Pork * * F27 Beef * * F3 Codfish * NT F4 Wheat flour * * F40 Tuna * * F41 Salmon * NT F45 Yeast, bakers M44 Yeast, bakers F45 Yeast, bakers F47 Garlic * * F48 Onion * * F6 Barley meal * * F81 Cheddar cheese * * F83 Chicken * * F9 Rice * * Fx Citrus mix Kofx Citrus mix NT M5 Candida albicans * * Common D1 D. pteronyssinus * * D2 D. farinae * * H1 House dust Hx House dust H1 House dust E1 Cat * * E2/E5 Dog epithelium E5 Dog E2 Dog epithelium F1 Egg white * * F14 Soy bean * * F2 Milk * * F206 Mackerel * NT NT F23 Crab * * F24 Shrimp * * F95 Peach F53 Peach * G12 Rye pollens * G5 Ryegrass I6 Cockroach * * M2 Cladosporium * * M3 Aspergillus fumigatus * * M6 Alternaria alternata * * T7 Oak white * * Tx Birchalder mix TxKo Birchalder mix T2, T3 Grey alder, common silver birch W2 Ragweed, short W2 Ragweed, short W1 Common ragweed W22 Japanese hop NT NT W6 Mugwort * * *same code and allergen as LGScreen. Abbreviation: NT, not tested.

4 Oh EJ, et al., AdvanSure Allergy Screen Test IU/mL, >200 IU/mL 로구분되어얻어진다. 검사는 LGScreen 검사와동일하게 Profiblot 48 (Tecan Group Ltd.) 장비를이용하여시약설명서대로검사하였다. 각스트립에혈청 250 ml를넣고 45분동안실온에서반응시킨후세척하고, Biotin 표지항IgE 시약을 250 ml를첨가하고 45분동안실온반응후세척하였다. 효소접합용액 (streptavidinalkaline phosphatase) 250 ml를넣고실온에서 20분간반응시키고세척후발색용액을첨가하여 20분간반응시켰다. CCD 카메라 (Rida Sscreen) 를이용하여 class 06으로판독하고시약설명서에 slightly increased 로제시되어있는 class 2 이상 (0.7 ku/l 이상 ) 을양성으로판정하였다. 5. ImmunoCAP 검사 IU/mL 이상과이하로구분하여정성적으로비교한결과, 두검사의일치율은 73.9% (133/180) 였고 kappa 계수 (k) 는 0.333이었다. 불일치를보여면역비탁법으로재검하였던 34건가운데 76.5% (26건) 에서면역비탁법과 LGScreen 결과가일치하였다. 80 개검체로시행한호흡형패널검사에서는 27.5% (22건) 에서불일치하였고, 이중 86.4% (19건) 에서면역비탁법과 LG Screen 결과가일치하였다. 100개검체로시행한음식형패널검사에서는불일치를보인 25건 (25%) 중검사가가능하였던 12 건의면역비탁법검사결과, 59.3% (7건) 에서 LGScreen 결과와일치하였다 (Fig. 1). 2. Allergy screen 검사에의한각종알레르겐특이 IgE 양성빈도 ImmunoCAP 검사는혈청 40 ml를사용하여 Phadia의 ImmunoCAP specific IgE 전용시약과 UniCAP100 장비로측정하였다. 검사과정은혈청을시약과실온에서 30분간반응시키고세척후 bgalactosicase가결합된항 IgE 항체를첨가하여실온에서 30분간반응시켰다. 세척후기질액과 30분간반응후반응정지용액을넣고형광강도를측정하여 ku/l 사이의농도를구하였고, allergy screen 양성확인은 allergy screen 판정기준인 0.7 ku/l 이상을기준으로하였다. 6. 통계통계분석은 SPSS 10.0 (SPSS, Chicago, IL, USA) 와 Med Calc version 11.3 (MedCalc Software, Mariakerke, Belgium) 을이용하였고 Chisquare와 ttest 방법을사용하여 P<0.05 인경우를통계적으로유의하다고판정하였다. 각검사간일치율의통계적분석은 interrater agreement 방법을이용하였으며, Kappa 계수 0.20 이하를 poor agreement, 을 fair to moderate, 0.61 이상을 good agreement로판정하였다. Allergy Screen과 ImmunoCAP 검사의 class 결과의상관관계는 Pearson 상관계수를구하고상관계수 0.5 이상을뚜렷한상관관계가있는것으로판정하였다. 결과 1. 총 IgE 결과 LGScreen과 RIDAScreen 검사의총 IgE 결과를 100 LGScreen 검사결과, 흡입형알레르겐에대해서는 Dermatophagoides farinae (d2), Dermatophagoides pteronyssinus (d1), cat (e1), dog (e2/e5) 순으로, 음식형알레르겐에대해서는 garlic (f47), tomato (f25), buckwheat (f11), peanut (f13) 순으로높은양성빈도를보였다. RIDAScreen 결과, 흡입형알레르겐에서는 D. pteronyssinus (d1), sycamore mix (t1), D. farinae (d2), Bermuda grass (g2) 의순으로, 음식형알레르겐가운데는 cheddar cheese (f81), wheat flour (f4), buckwheat (f11), pork (f26) 순으로높은양성빈도를나타내었다. LGScreen 검사에서높은빈도를보인순서대로알레르겐특이항체양성빈도를 Fig. 2에나타내었다. Tatal lge (IU/mL) by Nephelometry LGScreen (IU/mL) RIDAScreen (IU/mL) (N=6) (N=20) (N=8) > > Food Inhalant Fig. 1. Quantitative results of total IgE in 34 sera showing discrepant results between the two allergy screen assays.

5 424 Korean J Lab Med 2010;30:42031 Positive rate (%) LGScreen RIDAScreen 0.0 D. farinae (d2)* D. pteronyssinus (d1)* Cat (e1)* Dog (e2/e5)* House dust (h1, hx)* Cockroach (f6)* Acarus siro (d70) Penicillium notatum (m1) Mites, microorganisms Alternaria alternata (m6)* Aspergillus fumigatus (m3)* Cladosporium (m2)* Candida albicans (m5) A 40.0 Positive rate (%) LGScreen RIDAScreen 0.0 D. farinae (d2) Rye pollen (g12)* Ragweed, short (w2)* Bermuda grass (g2) Mugwort (w6)* Oak white (t7)* Pigweed (w14) Reed (g7) Sweet vernal grass (g1) Timothy grass (g6) Birchalder mix (tx)* Dandelion (w8) Russian thistle (w11) Pine (t16) Goldenrod (w12) Poplar mix (t14) Sycamore mix (t11, t1) Ash mix (t15) Orchard grass (g3) Oxeye daisy (w7) Sallow willow (t12) Tree pollens, grasses and weeds Acaia (t19, t36) Japanese cedar (t17, t24) B 40.0 Positive rate (%) LGScreen RIDAScreen 0.0 Garlic (f47) Tomato (f25) Buckwheat (f11) Yeast, bakers (f45, m44) Onion (f48) Peanut (f13) Barley meal (f6) Peach (f95, f53)* Crab (f23)* Rice (f9) Citrus mix (fx) Egg white (f1)* Beef (f27) Foods Shrimp (f24)* Hazelnut (f17) Soy bean (f14)* Chicken (f83) Pork (f26) Tuna (f40) Milk (f2)* Wheat flour (f4) Cheddar cheese (f81) Codfish (f3) C Fig. 2. Positive rates of allergenspecific IgE in LGScreen and RIDAScreen assays. Numbers of sera tested according to different allergens were 180(*), 100( ) and 80( ). 항원그룹을진드기 (mites), 동물항원 (animals), 진균류 (molds) 가포함된실내항원과, 수목화분 (tree pollens), 목초 (grasses), 잡초 (weeds) 가포함된실외항원, 그리고음식물항원인식품 (foods) 의세그룹으로구분하여양성빈도를비교한결과, LG Screen과 RIDAScreen의실내항원의양성빈도 (13.0±13.4% vs. 13.2±10.4%) 에는유의한차이가없었으나 (P=0.978), 실외

6 Oh EJ, et al., AdvanSure Allergy Screen Test 425 Table 2. Comparison of the test results between LGScreen and RIDAScreen assays Allergens N LGScreen + + RIDAScreen + + Agreement % (kappa value) D. pteronyssinus (d1) (0.481) D. farinae (d2) (0.487) Acarus siro (d70) (0.267) House dust (h1, hx) (0.215) 79.3 (0.443)* Cat (e1) (0.027) Dog (e2, e5) (0.235) 75.6 (0.123) Egg white (f1) (0.550) Buckwheat (f11) (0.313) Peanut (f13) (0.497) Soy bean (f14) (0.126) Hazelnut (f17) (0.388) Milk (f2) (0.011) Crab (f23) (0.275) Shrimp (f24) (0.661) Tomato (f25) (0.440) Pork (f26) (0.020) Beef (f27) (0.128) Codfish (f3) (NaN) Wheat flour (f4) (0.000) Tuna (f40) (0.000) Salmon (f41) (NaN) Yeast, Baker (f45, M44) (0.000) Garlic (f47) (0.019) Onion (f48) (0.519) Peach (f95, f53) (0.332) Barley meal (f6) (0.519) Cheddar cheese (f81) (0.000) Chicken (f83) (0.000) Rice (f9) (0.286) Citrus mix (fx) (0.419) 89.5 (0.220) Sweet vernal grass (g1) (0.515) Rye pollens (g12) (0.609) Bermuda grass (g2) (0.299) Orchard grass (g3) (0.104) Timothy grass (g6) (0.186) Reed (g7) (0.538) 85.6 (0.403) Cockroach (i6) (0.509) Penicillium notatum (m1) (0.385) Cladosporium (m2) (0.032) Aspergillus fumigatus (m3) (0.203) Candida albicans (m5) (0.029) Alternaria alternata (m6) (0.215) 90.9 (0.264) Sycamore mix (t11, t1) (0.057) Sallow willow (t12) (0.000) Poplar mix (t14) (0.022) Ash mix (t15) (0.233) Pine (t16) (0.221) Japanese cedar (t17, t24) (0.000) Acacia (t19, t36) (0.000) Oak white (t7) (0.427) Birchalder mix (tx) (0.198) 86.5 (0.186) Russian thistle (w11) (0.200) Goldenrod (w12) (0.078) Pigweed (w14) (0.231) Ragweed, short (w2) (0.421) Mugwort (w6) (0.364) Oxeye daisy (w7) (0.000) Dandelion (w8) (0.085) 83.6 (0.263) Total 7, , (0.293) *Agreement between LGScreen and RIDAScreen results according to the allergen group specific IgE. Abbreviation: NaN, not a number.

7 426 Korean J Lab Med 2010;30:42031 항원과음식물알레르겐에대한양성빈도는 RIDAScreen이유의하게높았다 (3.3±3.1% vs. 16.2±7.8%; P<0.0001, 3.0 ±2.9% vs. 10.8±10.1%; P=0.0007). 또한항원그룹을진드기 (mites), 동물항원 (animals), 진균류 (molds), 수목화분 (tree pollens), 목초 (grasses), 잡초 (weeds), 식품 (foods) 으로구분하여양성빈도를비교한결과, 진드기와동물항원을제외한나머지항원그룹에서는모두 RIDAScreen의양성빈도가유의하게높았다 (P<0.05). 3. LGScreen과 RIDAScreen 결과의일치율비교 LGScreen과 RIDAScreen 두가지시약에모두포함된 58종알레르겐가운데흡입형패널에만포함된 19종은 80검체에서 (1,520건), 음식형패널에만포함된 19종은 100검체에서 (1,900건), 두가지패널에모두포함된 20종은 180검체 (3,600건) 의총 7,020건에서두방법의결과를비교하였다 (Table 2). 두가지 allergy screen 검사결과의전체일치율은 86.7% (6,086/ 7,020) (k 0.293) 였다. 알레르겐종류별로는 % (k ) 의일치율을보였고, 두방법간결과의일치율이 70% 이하, 7080%, 8090%, 90100% 인알레르겐을 Table 3 에나열하였다. 4. LGScreen과 RIDAScreen 검사에서불일치를보인 633건에서시행한 ImmunoCAP 검사결과두가지 allergy screen 검사결과, 58종알레르겐에대한특이 IgE 검사 7,020건중 934건에서불일치결과를보였고이가운데 633건 (67.8%) 에서 ImmunoCAP 검사가가능하였다. ImmunoCAP 검사를시행한 633건에는 LG와 RIDAScreen 검사결과에불일치예가없거나검체부족으로 ImmunoCAP 검사가불가능하였던 salmon (f41), codfish (f3), citrus mix (fx), ash mix (t15) 를제외한 54종알레르겐에대한특이 IgE 검사를포함하였고, 알레르겐종목별로 12±8 건이시행되었다. LGScreen 검사는 633건중 449건 (70.9%) (kappa 0.585) 에서, RIDAScreen 검사는 184건 (29.1%) (k, 0.303) 에서 ImmunoCAP 검사결과와일치하였다 (Table 4). LGScreen 양성 /RIDAScreen 음성이었던 137건중 75건 (54.7%) 에서 ImmunoCAP 양성이었고, LGScreen 음성 /RIDAScreen 양성이었던 386건중 374건 (96.9%) 에서 ImmunoCAP 음성이었다. ImmunoCAP 검사와 LGScreen 검사의알레르겐그룹 Table 3. Specific allergens according to the agreement between LGScreen and RIDAScreen assay results <70% Agreement between LGScreen and RIDAScreen assay 7080% 8090% 90100% Allergens Cat D. farinae Dog Penicillium notatum Cheddar cheese D. pteronyssinus House dust Alternaria alternata Wheat Acarus siro Cockroach Pork Cladosporium Candida albicans Milk Buckwheat Aspergillus fumigatus Sycamore mix Beef Peanut Timothy grass Rice Soy bean Goldenrod Peach Yeast, bakers Bermuda grass Tomato Egg white Sallow willow Crab Onion Birchalder mix Garlic Barley meal Orchard grass Citrus mix Oxeye daisy Hazelnut Dandelion Shrimp Russian thistle Tuna Ragweed, short Chicken Mugwort Rye pollens Pigweed Poplar mix Acacia Reed Oak white Pine Ash mix Sweet vernal grass Japanese cedar Codfish Salmon

8 Oh EJ, et al., AdvanSure Allergy Screen Test 427 Table 4. Analysis of discrepant results between LGScreen and RIDAScreen assays compared with ImmunoCAP assay (N=633) Allergens N LGScreen RIDAScreen ImmunoCAP Agreement % (kappa value) between ImmunoCAP & LG RIDA D. pteronyssinus(d1) 27 2 (7.4)* 8 (29.6) 2 (7.4) 15 (55.6) D. farinae (d2) 31 3 (9.7) 21 (67.7) 3 (9.7) 4 (12.9) Acarus siro (d70) 13 (0.0) 1 (7.7) 5 (38.5) 7 (53.8) House dust (hx, h1) 28 4 (14.3) 6 (21.4) 8 (28.6) 10 (35.7) (0.127) (0.138) Cat (e1) (50.0) 9 (25.0) 8 (22.2) 1 (2.8) Dog (e2,e5) (68.2) 6 (27.3) 1 (4.5) (0.0) (0.094) (0.172) Egg white (f1) 6 (0.0) (0.0) 5 (83.3) 1 (16.7) Buckwheat (f11) 9 (0.0) (0.0) 5 (55.6) 4 (44.4) Peanut (f13) 3 (0.0) 1 (33.3) 1 (33.3) 1 (33.3) Soy bean (f14) 7 (0.0) (0.0) 1 (14.3) 6 (85.7) Hazelnut (f17) 3 (0.0) (0.0) 3 (100) (0.0) Milk (f2) 24 (0.0) (0.0) 22 (91.7) 2 (8.3) Crab (f23) 16 2 (12.5) 1 (6.3) 12 (75.0) 1 (6.3) Shrimp (f24) 2 (0.0) 1 (50.0) (0.0) 1 (50.0) Tomato (f25) 6 (0.0) (0.0) 2 (33.3) 4 (66.7) Pork (f26) 15 (0.0) (0.0) 15 (100) (0.0) Beef (f27) 13 (0.0) (0.0) 13 (100) (0.0) Wheat flour (f4) 17 (0.0) (0.0) 6 (35.3) 11 (64.7) Tuna (f40) 1 (0.0) 1 (100) (0.0) (0.0) Yeast, Bakers (f45, M44) 5 3 (60.0) 2 (40.0) (0.0) (0.0) Garlic (f47) 9 1 (11.1) 8 (88.9) (0.0) (0.0) Onion (f48) 5 (0.0) 3 (60.0) (0.0) 2 (40.0) Peach (f95, f53) 18 (0.0) (0.0) 11 (61.1) 7 (38.9) Barley meal (f6) 3 (0.0) 1 (33.3) 2 (66.7) (0.0) Cheddar cheese (f81) 24 (0.0) (0.0) 24 (100) (0.0) Chicken (f83) 1 1 (100) (0.0) (0.0) (0.0) Rice (f9) 9 (0.0) (0.0) 3 (33.3) 6 (66.7) (0.271) (0.160) Sweet vernal grass (g1) 5 (0.0) 1 (20.0) 3 (60.0) 1 (20.0) Rye pollens (g12, g5) 6 1 (16.7) 2 (33.3) 3 (50.0) (0.0) Bermuda grass (g2) 6 (0.0) (0.0) 6 (100) (0.0) Orchard grass (g3) 13 (0.0) (0.0) 11 (84.6) 2 (15.4) Timothy grass (g6) 17 (0.0) (0.0) 17 (100) (0.0) Reed (g7) 5 (0.0) (0.0) 4 (80.0) 1 (20.0) (0.496) (0.117) Cockroach (i6) 13 1 (7.7) 2 (15.4) 3 (23.1) 7 (53.8) Penicillium notatum (m1) 7 1 (14.3) (0.0) 5 (71.4) 1 (14.3) Cladosporium (m2) 15 3 (20.0) (0.0) 11 (73.3) 1 (6.7) Aspergillus fumigatus (m3) 3 1 (33.3) (0.0) 2 (66.7) (0.0) Candida albicans (m5) 3 (0.0) (0.0) 3 (100) (0.0) Alternaria alternata (m6) 8 2 (25.0) 1 (12.5) 5 (62.5) (0.0) (0.069) (0.025) Sycamore mix (t11, t1) 22 (0.0) (0.0) 22 (100) (0.0) Sallow willow (t12) 7 (0.0) (0.0) 6 (85.7) 1 (14.3) Poplar mix (t14) 7 1 (14.3) (0.0) 5 (71.4) 1 (14.3) Pine (t16) 6 1 (16.7) (0.0) 5 (83.3) (0.0) Japanese cedar (t17, t24) 2 (0.0) (0.0) 2 (100) (0.0) Acacia (t19, t36) 4 (0.0) (0.0) 4 (100) (0.0) Oak white (t7) 14 (0.0) (0.0) 10 (71.4) 4 (28.6) Birchalder mix (tx, t2, t3) 18 (0.0) (0.0) 12 (66.7) 6 (33.3) (0.045) (0.009) Russian thistle (w11) 13 (0.0) (0.0) 12 (92.3) 1 (7.7) Goldenrod (w12) 17 (0.0) (0.0) 17 (100) (0.0) Pigweed (w14) 9 1 (11.1) (0.0) 8 (88.9) (0.0) Ragweed (w2, w1) 17 (0.0) (0.0) 15 (88.2) 2 (11.8) Mugwort (w6) 17 (0.0) (0.0) 11 (64.7) 6 (35.3) Oxeye daisy (w7) 13 (0.0) (0.0) 9 (69.2) 4 (30.8) Dandelion (w8) 13 1 (7.7) (0.0) 11 (84.6) 1 (7.7) (0.037) (0.007) Total (9.8) 75 (11.8) 374 (59.1) 12 (19.3) 70.9 (0.585) 29.1 (0.303) *Number of cases(%); Agreement between LGScreen and RIDAScreen results according to the allergen group specific IgE. Abbreviations: +, positive;, negative, NaN, not a number.

9 428 Korean J Lab Med 2010;30:42031 별일치율 (kappa) 은진드기 (mites) 알레르겐 54.5% (0.127), 동물 (animals) 알레르겐 41.4% (0.094), 음식 (foods) 알레르겐 73.0% (0.271), 목초화분 (grasses) 알레르겐 90.4% (0.496), 진균류 (molds) 75.0% (0.069), 수목화분 (tree pollens) 82.5% (0.045), 잡초화분 (weeds) 83.8% (0.037) 였다. 알레르겐종류와그룹별로두가지 allergy screen 검사와 ImmunoCAP 검사의일치율을 Table 4에나열하였다. 두 allergy screen 검사결과의불일치검체에대하여 allergy screen 검사의 class 결과와 ImmunoCAP의 class 결과의상관관계를정량적으로비교하면, LGScreen (r=0.277) 과 RIDAScreen (r=0.149) 의 ImmunoCAP과의상관관계는낮았다. 고찰알레르기질환진단을위한총 IgE의임상적유용성은알레르겐특이 IgE 항체검사보다낮은데, 12 종류의알레르기항원에만감작된경우는혈청총 IgE가정상범위일수있고, 여러종류의알레르기항원에감작되어천식, 아토피피부염, 비염등의복합적증상을보이는경우에는농도가크게증가하기때문이다 [5]. Allergy screen 검사에서총 IgE는 100 또는 200 IU/mL 을기준으로정성결과가보고되는데, 알레르겐특이 IgE 검사결과의확인과판정에보조적으로이용될수있다. 본연구에서비교한두가지 allergy screen 검사의총 IgE 결과는흡입형패널과음식형패널로비교한검체들에서각각 27.5% 와 25% 의불일치결과를보였다. 비록불일치를보였던모든검체를대상으로면역비탁법정량검사를시행하지는않았으나, LGScreen 검사의총 IgE 검사결과와면역비탁법검사결과와의일치율이 RIDAScreen에비해유의하게높았다. 총 IgE 결과가 allergy screen 검사결과의확인에이용될수있으나, 알레르겐특이 IgE와총 IgE 검사결과가불일치를보이는경우는다른방법으로정량적총 IgE 검사결과의확인이필요하다고판단된다. 검사실에서의혈청알레르겐특이 IgE 측정은알레르기질환의 1차적선별검사나피부검사의보조적수단으로이용되고있다. 국내에도 allergy screen 검사시약으로서 MAST CLA, RIDA Allergy Screen, Polycheck Allergy 등의시약이도입되어널리이용되고있으며, 반정량법인 allergy screen 검사를이용한알레르기선별검사후에정량적검사를실시함으로써보다정확한진단이가능하게되었다 [68]. LGScreen은국내회사에서개발된시약으로서한국에서흔한 60종의알레르겐을선별하여패널을구성하였고, 기존의수입 allergy screen 시약에는포함되지않았으나한국인에서감작률이높은알레르 겐으로보고된 [9, 10] 환삼덩굴 (Japanese hop) 과고등어 (mackerel) 항원을포함하고있다. 또한시약에포함된알레르겐가운데약30여종의알레르겐은자체정제하였으므로, 수입항원의구조와차이가있을수있으나, 한국인의알레르기질환의진단과기전규명에더유용한정보를줄수있을것으로생각한다. 반정량검사인 allergy screen에대한기존의평가는알레르겐특이 IgE 항체검사법의민감도나특이도를생체내검사 ( 피부검사및유발시험 ) 나병력과비교한연구가일반적이다 [69]. 또한연구자에따라대표적인알레르기항원에대해서만평가하였고, allergy screen에포함된모든종류의알레르겐특이항체에대하여평가한보고는드물다. 그러나피부시험, 유발시험과같이반응세포의활성화에의한다클론성 (polyclonal) 면역반응을검출하는생체내 (in vivo) 검사는알레르겐특이 IgE 항체를검출하는생체외 (in vitro) 검사의절대적인민감도와특이도의참고검사가될수없다 [11]. 피부시험에서음식물알레르기의진단적가치는호흡기알레르기에비해떨어진다고알려져있으며, 생체내검사는알레르기항원노출후검사까지의시간, 알레르기항원의종류, 환자의연령, 대상장기에따라시약의민감도가다르게나타날수있고, 생리적인면역반응이환자에따라차이가나거나교차반응도나타날수있으며 [13], 판독이주관적일수있다. 따라서본연구에서는 LGScreen 검사의유용성평가를위해, 동일한알레르겐을포함하고있고국내사용빈도가높은 RIDAScreen 검사와비교하고, 불일치예에서 allergy screen 검사에포함된대부분의알레르겐에대한특이항체를대상으로 ImmunoCAP 검사로확인하였다. 기존보고에의하면 RIDAScreen 검사의 ImmunoCAP 검사와의일치율, 양성일치율, 음성일치율은 81.8%, 58.2%, 94.6% 로, 피부검사와의일치율, 양성일치율, 음성일치율은 89.2%, 59.1%, 90.0% 로보고되어있다 [6, 7]. 본연구에서는 allergy screen 검사결과 class 2 (0.7 IU/mL) 이상을양성으로판정하였고, 정성적인판단결과에차이를보인경우, 즉두가지 allergy screen 결과가 class 2 이상과이하로다른경우에 ImmunoCAP으로확인검사를시행하였다. Class 2 이상을양성으로설정한이유는, allergy screen는반정량검사이므로 ±1 class의검사결과는차이를보일수있고특이 IgE 항체를선별검사로이용할경우위양성결과를보고하는경향이있다고보고되어있기때문이다 [1, 12]. 또한음식물에대한낮은농도의 IgE는임상적으로의미없는경우가많은데, 계란, 우유, 땅콩, 생선은 class 3 이상의알레르겐특이 IgE 농도인 632 IU/mL가존재하는경우에만유발시험양성과관계가있다는보고가있으므로 [13], 기존평가보고 [8, 14] 에서와같

10 Oh EJ, et al., AdvanSure Allergy Screen Test 429 이 class 2 이상을양성으로간주하였다. 저자들은알레르기질환이의심되어의뢰된검체중 RIDA Screen 검사에서한가지항원에서라도양성으로검출된검체를대상으로양성빈도를구하였으므로기존의한국인을대상으로한감작률보고결과 [9] 와직접적인비교는불가능하다. 따라서 LGScreen 검사에서높은빈도로검출되는알레르겐을나열하고 (Fig. 2), 두 allergy screen 검사간항원그룹의양성빈도를비교하였는데, 진드기와동물항원을제외한모든항원그룹에서 RIDAScreen의양성빈도가유의하게높았다. RIDA Screen과다른 allergy screen 검사의비교에대한기존보고에의하면비록통계적유의성은분석되지않았으나 RIDA Screen는 D. farinae, cat 항원특이항체양성빈도가 82% 와 14% 로서, 다른 allergy screen 시약의양성빈도인 D. farinae 86%, cat 28% 보다다소낮았고, birch, ragweed, timothy grass, acacia, oak white 등의목초, 수목항원의양성빈도는높아본연구결과와일치하였다 [7]. 본연구에서 LGScreen 검사에만포함된환삼덩굴은 180검체중 2예 (1.1%) 에서양성이었고, 고등어항원에대한양성검체는없었는데, 대상군의연령과검사방법, 항원등의차이에의해기존보고 (2.75.4%, 4.2%) [9, 10] 와다른결과를나타낸것으로판단된다. 58종알레르겐에대한 LGScreen과 RIDAScreen 검사결과의전체일치율은 86.7% (6,086/7,020) (k, 0.293) 였고, 알레르겐종류에따라 % (k, ) 의다양한일치율을나타냈는데, 이또한시약에포함된알레르겐원료의차이, 추출방법의차이, 알레르겐부착방법의차이등에의하는것으로생각된다 [13]. 본연구에서는비교한시약사이에동일한알레르겐코드가없거나코드번호가다른 10여가지의알레르겐에대해, 알레르겐의명칭이같은경우동일한알레르겐으로간주하고결과를비교하였다. 코드번호의차이는시약제조회사마다알레르겐의원료와추출방법의차이에의한것으로생각되며, 이또한불일치결과의원인으로작용하였을가능성이있다. 저자들이불일치예에대한확인검사로이용한 Immuno CAP 검사는국내뿐아니라세계적으로널리이용되고있는알레르겐특이 IgE 정량검사로서임상적유용성의체계적인평가가가장많이보고된검사이며 [2], 알레르기환자로확진된대상군에서민감도와특이도가 91100% 와 6273% 로보고되어있다 [15]. 본연구에서두가지 allergy screen 검사에불일치를보인 633건의 ImmunoCAP 확인검사결과 70.9% 에서 LG Screen 검사와 ImmunoCAP 검사결과가일치하였다 (k 0.585). 알레르겐종류별로불일치원인을분석하면, LGScreen 검사는 dog (e2/e5), chicken (f83), yeast (f45) 항원에대한양성, soy bean (f14), tomato (f25), wheat flour (f4), rice (f9) 에대한음성이주된불일치원인이었고, RIDAScreen 검사는음식물알레르겐, 목초화분, 진균류, 수목화분알레르겐에서의양성과 D. farinae (d2), tuna (f40), garlic (f47), onion (f48) 에대한음성이불일치의원인이었다. RIDAScreen과 Immuno CAP의일치율에대한이전보고 [6] 에의하면, D. farinae (83.0%), dog (80.6%), house dust (62.5%), cockroach (81.0%), ragweed (80.6%), mugwort (65.8%), rye (100%), birchalder (86.4%) 등이나, 본연구에서는각특이알레르겐에대한일치율이 kappa 계수 0.2 이하로낮았는데, 저자들은두가지 allergy screen 검사에불일치를보인검체만을대상으로 ImmunoCAP 검사를시행하였으므로, 이전보고결과와직접적인비교는어렵다. 또한일치율과 kappa 값을모두구하여제시하였는데, kappa 값은통계분석시 2 2 분할표에서주변합이심하게불균형되어있어 kappa 값이낮게추정되었으므로, 일치율과 kappa 값에차이가있는경우해석의주의가필요하다 [17]. RIDAScreen과 ImmunoCAP 불일치의주요한원인인실외항원에대한양성은유사한항원에대한교차반응에의한것으로판단되며목초, 진균류등에서는항원차이에의해검사실간에차이가많다는이전보고와일치한다 [16]. 일반적으로정제하지않은추출물 (crude extract) 이알레르기항원으로사용되므로, 항원에는여러종류의알레르기항원이포함되어있거나항원성이없거나미미한다양한단백질도포함될수있다. 또한알레르기를유발하는물체는대게여러알레르기항원의복합체이며각각의알레르기항원은다수의항원결정인자를가지고있으므로, 알레르겐특이 IgE는교차반응을일으킬수있다 [1820]. 본연구에서분석된 RIDAScreen의교차반응은 RIDAScreen 검사가민감도가높은대신특이도가낮았다고보고한이전연구결과 [7] 와일치한다. 그러나 allergy screen 검사는일반적으로 ImmunoCAP 검사나피부시험에비해민감도가낮다고보고되어있고, 교차반응이알레르기질환의진단과치료에도움이된다는보고가있으므로 [20], 정확한민감도, 특이도분석을위해서는불일치검체가아닌알레르기질환환자를대상으로한추가평가가필요하다. 또한 dog, rye pollens, ragweed의경우 allergy screen (E2/E5, E5, G12, W2) 과 ImmunoCAP (E2, G5, W1) 에서사용된알레르겐코드가달랐는데, 항원종류와추출방법의차이가결과의차이를유발하였을가능성이있으므로, 추후알레르겐코드가동일한시약으로비교를하면본연구결과와다른결과를보일수있을것이다.

11 430 Korean J Lab Med 2010;30:42031 Allergy screen 검사와 ImmunoCAP 검사의 class 결과를정량적으로비교한이전연구에서는두가지검사사이의양호한상관관계 (R=0.60.8) 를보고하였는데 [6, 14], 본연구에서는두가지 allergy screen 검사모두 ImmunoCAP 결과와의상관성이매우낮았다 (r=0.277, r=0.149). 이는본연구에서두가지 allergy screen 검사의결과에불일치를보인일부검체 ( 전체검체중 13.3%) 만을대상으로 ImmunoCAP 검사를시행하였기때문이며, 불일치검체가상관성비교에많이포함되어낮은상관성결과를보인것으로판단된다. 따라서전체검체를대상으로 class 결과에따른정량적비교를한다면상관관계는향상될것으로생각된다. 국내개발된 LGScreen 검사의적용은추후추출정제된항원과수입항원에의한결과분석을통해알레르기질환의인종적차이나기전규명연구에도움이될수있을것으로생각한다. 또한국내에서개발된검사법이므로검사실의요구에대한제조사의긴밀한피드백이가능하리라예상되며, 본연구에서일치율에차이를보였던알레르겐에대해서항원정제나시약제조방법등의최적화가이루어진다면보다우수한성능의시약으로업그레이드도가능할것이다. 현재국내에는알레르기관련검사에대한외부정도관리사업이없고, 알레르기관련시약이고가이며, 정도관리물질의표준화및참고검사방법이부재하여검사실간비교도어려운실정이다 [4]. 앞으로혈청알레르겐특이항체검사에대한질향상과표준화를위한국내외부정도관리프로그램의도입이요구되며, 정도관리물질의조제와공급에대해국내제조회사와의상호협조를기대해본다. 결론적으로 LGScreen 검사는기존에사용되고있던 allergy screen 검사법과양호한일치율을보였고, 불일치검체를대상으로시행한 ImmunoCAP 정량검사와도기존검사에비해우수한일치율을보였으므로 (k, vs ) 알레르겐특이 IgE 검출을위한검사로서유용할것이라생각한다. 요약배경 : 혈청알레르겐특이 IgE 측정은알레르기질환의진단을위해임상검사실에서흔히이용된다. 최근국내에서혈청알레르겐특이 IgE 검출을위한 allergy screen 검사인 AdvanSure Allergy Screen (LG Life Science, Korea) 시약 (LGScreen) 이개발되어소개되었기에 LGScreen 검사의임상적유용성을평가하였다. 방법 : 총 180예의혈청검체를이용하여 LGScreen과 RIDA Allergy Screen (Rbiopharm, Germany) (RIDAScreen) 검사 를시행하였으며, 80예는호흡형패널로 100예는식품형패널로검사하였다. 5860종의알레르겐또는알레르겐그룹에따라양성빈도와두가지 allergy screen 검사결과의일치율을분석하였으며, class 2 이상을양성으로판정하였다. 두가지 allergy screen 검사에서총 IgE와알레르겐특이 IgE 검사결과가불일치를보이는검체에대하여면역비탁법과 ImmunoCAP 방법으로정량검사를시행하였다. 결과 : LGScreen과 RIDAScreen 검사의총 IgE 검사결과의일치율은 73.9% (133/180), kappa 계수 (k) 는 0.333이었다. 불일치검체를대상으로시행한면역비탁법검사와는 LG Screen이 RIDAScreen에비해높은일치율을보였다. RIDA Screen 검사는실외호흡형항원과음식물항원에대해유의하게높은양성률을보였다. 58종알레르겐특이 IgE 검출을위한두가지 allergy screen 검사결과의전체일치율은 86.7% (6,086/ 7,020) (k, 0.293) 였다. Allergy screen 검사에서불일치결과를보여 ImmunoCAP 검사를시행한 633건중 70.9% (449건 ) 는 LGScreen 결과와, 29.1% (184건 ) 는 RIDAScreen 결과와일치하였다 (k, vs ). 결론 : LGScreen 검사는기존에사용되고있던 allergy screen 검사법과양호한일치율을보였고, 불일치검체를대상으로시행한 ImmunoCAP 정량검사와도양호한일치율을보였으므로알레르겐특이 IgE 검출을위한검사로서유용할것이라생각한다. 참고문헌 1. Hamilton RG. Clinical laboratory assessment of immediatetype hypersensitivity. J Allergy Clin Immunol 2010;125:S Cox L, Williams B, Sicherer S, Oppenheimer J, Sher L, Hamilton R, et al. Pearls and pitfalls of allergy diagnostic testing: report from the American College of Allergy, Asthma and Immunology/American Academy of Allergy, Asthma and Immunology Specific IgE Test Task Force. Ann Allergy Asthma Immunol 2008;101: Steckelbroeck S, BallmerWeber BK, Vieths S. Potential, pitfalls, and prospects of food allergy diagnostics with recombinant allergens or synthetic sequential epitopes. J Allergy Clin Immunol 2008;121: Lim HS, Kim HS, Oh H. Current status of serum allergen tests in Korea. Korean J Lab Med 2008;28:1249. ( 임환섭, 김현수, 오흥범. 국내혈청알레르기항원검사법의현황. Korean J Lab Med 2008;28:1249.) 5. CLSI. Analytical performance characteristics and clinical utility of

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