Unexpected Antibody Screening & Identification

제품 소개

주식회사 유니온랩

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DIAGNOSTIC GRIFOLS

• Spain 에 본부를 둔 약 50 년 역사의 수혈의학관련 제품 및 면역혈청진단 및 치료제 전문기업입니다 .

• 1998 년 , 세계 최초로 전자동 혈액은행 검사 장비 출시하였습니다 .

WADiana 를 • 혈액은행검사 자동화 분야에 있어서는 가장 앞서 있는 선두주자입니다 . • DiaMed 사에는 ID Gel Station 이란 이름으로 , J&J Ortho 에는 라는 이름으로 자동화 장비를 OEM 공급하고 있습니다 . ProVue • 2006 년 현재 , 전세계에 • WA Diana 는 미국 장비입니다 .

1,700 여대의 장비가 가동 중에 있습니다 . FDA 의 승인을 받은 유일한 혈액은행검사 자동화

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Gel Column Technique

• • • • • • • •

안정한 반응 표준화 기법 높은 감도 보다 분명한 결과 간편한 취급 항글로불린검사시 세척과정 불필요 적은 검체량 자동화 용이

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210310 210311 210313 210312 210314 210315 210316 210319 210320 210321 210317 210323 210318 210324 210322 6 7 1 2 3 4 5 8 9 10 11 12 13 14 15

Grifols DG Gel

®

Cards

종류

DG Gel Configuration

Cat #

DG Gel ABO/Rh DG Gel ABO/Rh (2D) DG Gel ABO-CDE DG Gel Confirm DG Gel Anti-D DG Gel Rh DG Gel Rh+Kell DG Gel Coombs DG Gel Neutral DG Gel Neutral/Coombs DG Gel Neonatal DG Gel DC Scan DG Gel CT DG Gel AB DG Gel IgG A B AB D CDE Ctrl A1 B A B AB D D' Ctrl A1 B A B AB D C E A1 B 2 x (A B D Ctrl) 4 x (D' Ctrl) 2 x (C c E e) D' C Cw c E e Kell Ctrl 8 x (AHG) 8 x (N) N N N N AHG AHG AHG AHG A B AB D D' Ctrl IgG AHG 2 X (AHG IgG CD3 Ctrl) A B D Ctrl N N AHG AHG 4 X (A B) 8 x (IgG) Marketing GI Immunohematology

•

Unexpected Antibody Test

용 진단제품

DG Gel Coombs Card

– – 마이크로 컬럼법을 사용하는 혈액형판정용 카드제품중 Sephadex Gel column 방식은 glass bead column 방식보다 specificity 가 뛰어납니다 . 또 , Gel column 방식은 Micro Glass bead column 방식에 비해 없습니다 .

false positive 율이 거의 카드 한 개당 8 개의 구성되어 있습니다 .

column (4 환자분 ) 으로

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•

Unexpected Antibody Test

용 진단제품

SeraScan Diana

-

비예기항체 선별용 Screen cells • • • 제품 사용기간이 제조일로부터 8 주로서 타제품들 (6 주 ) 보다 사용기간이 길어 손실율이 적습니다 .

제품 자체가 LISS (Low Ion Strength Solution) 처리되어 있어 항체 검출율을 높이고 별도의 LISS 용액시약이 필요없으며 따라서 검사과정도 간단합니다 .

검출 가능한 비예기항체의 종류는 국제표준인 23 종의 항체를 포함합니다 . D,C,E, c, e, K, k, Kp b , Js b , M, N, S, s, Fya, Fyb, Jk a , Jk b , Le a , Le b , Lu b , P 1 , Xg a

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•

Unexpected Antibody Test

용 진단제품

SeraScan Diana

-

비예기항체 선별용 Screen cells • • C w 항체를 항상 포함하고 있습니다 .

가장 흔히 검출되는 망라하고 있습니다 .

Rh, Duffy, MNS, Kidd 시스템을 • Homozygous 인 검출합니다 . D, C, c, E, e, Fy a , Fy b , Jk a , Jk b 를 각각 • R 1 R 1 와 R 2 R 2 profiles 이 있습니다 .

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•

Unexpected Antibody Test

용 진단제품

IdentiSera

-

비예기항체 동정용 cells • 비예기항체 동정을 위한 적혈구 시약으로 있습니다 .

11 vial 로 구성되어 • • 정확히 동정되지 않는 특수케이스를 위해 추가동정을 할 수 있는 4 vial 이 별도로 준비되어 있습니다 . (IdentiSera Extend) 세 개의 Kell-positive cells 을 함유하고 있습니다 .

• Homozygous 항체 Fy a , Fy b , Jk a , Jk b - M, N, S, s, C, c, E, e, D, Lu 모두 동정 가능합니다 .

b , k, Kp b , Js b ,

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•

Unexpected Antibody Test

용 진단제품

IdentiSera

-

비예기항체 동정용 cells • Minimum two homozygous cells for: Fy a , Fy b , Jk a , Jk b , S, s, M, N • • Presence of: P 1 한 판넬에 Fy, MNS cells negative, Le(a-b-), 2 cells Le(a 3 R 1 R 1 이 모두 있습니다 .

+ cells, 4 rr homozygous cells, ) 그리고 보조적 Jk, • With presence of anti-D, anti-C or anti-E, a 2 nd Duffy, MNS or Kell can be detected antibody of Kidd, • 4 possible complementary combinations for Fy and Jk

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Identisera + Identisera Extended Total 15 cells –

비예기항체 동정이 완벽합니다

.

Control 1 patient = 2 cards = 15 cells + 1 Control Marketing GI Immunohematology

DG Gel

®

+ Identisera + Identisera Extended

최대수의 homozygous 항원과 보조적 적혈구항원의 조합으로 비예기항체의 검출과 동정을 완벽히 수행 할 수 있습니다 . Maximum efficiency and information per card and test Optimization of the unique 8 column format

Grifols Clear Card Technology

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Diana System Equipment

Diana Incubator

•

24 Gel cards 동시장착가능 (24 cards X 4 Pt’s/card = 96 Pts) • Timer 내장 • Incubation 시간 : 37 도 15 분간 • 열전도에 의한 가온법 • 실험 튜브도 가온 가능

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Diana System Equipment DianaFuge

• 동시 24 Gel cards 원심분리 가능 • 1100 rpm 에서 9 분간 원침

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• • •

Unexpected Antibody Test

용 진단제품

우수한 성능 – 기존 제품과의 평가에서 가장 우수한 성적을 보였습니다 . (2005 AABB poster 및 2006 논문 full text 참조 )

(J. Cid et al.

Comparison of three microtube column agglutination systems for antibody screening: DG Gel, DiaMed-ID and Ortho BioVue

Transfusion Medicine 2006, 16. 131-136

) 간편한

2 step

법 ① 혈청 ② 선별용 혈구  25uL 분주 50uL 분주 (LISS 사용 Incubation + Centrifuge step 없음 ) – – 경제성 명확한 결과 및 정확한 검사 Gel card 한 개당 4 환자 검사

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Advantages of DG Gel Card

• • • • • 8-column format 분명한 결과 간편한 사용법 쉬운 판독 사전 수혈검사시 높은 신뢰성

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Comparison of three microtube column agglutination systems for antibody screening: DG Gel, DiaMed-ID and Ortho BioVue Joan Cid, Núria Nogués, Rosa Montero, Marta Hurtado, Albert Briega and Rafael Parra.

Blood and Tissue Bank. Barcelona, Spain.

INTRODUCTION

Antibody screening is performed to detect clinically relevant red blood cell (RBC) antibodies in transfusion recipients and blood donors. The classical low-ionic-strength-solution (LISS) tube indirect antiglobulin test (IAT) for detecting clinically significant antibodies has been replaced in many countries by microtube column agglutination systems, described for first time in 1988 and published in 1990 (1). The first microtube system was manufactured and distributed by DiaMed-ID (Switzerland), followed by Otho BioVue (Otho Diagnostic GmBH, Germany). Recently, a new microtube column agglutination system has been launched on the market by Diagnostic Grifols, S.A. (Barcelona, Spain) with the name of DG Gel.

The aim of the present study was to evaluate the diagnostic accuracy of this new system, to analyze the antibody reactivity and to compare the data with the two cited well-established methods.

METHODS

Samples. We collected 3,024 specimens: 2,408 (79.6%) were plasma samples obtained from EDTA tubes and 616 (20.4%) were serum samples. We obtained 2,422 (80.1%) samples from blood donors in our blood bank; 382 (12.6%) samples from transfusion recipients and 220 (7.3%) from pregnant women in Hospital Vall d’Hebron (Barcelona, Spain). We stored all these samples in the refrigerator at 2-8 ºC and we processed them before 72 hours from extraction. We additionally obtained 100 samples containing antibodies of known specificity from the immunohematology laboratory of our blood bank. These samples had been stored in a frozen state at -30 ºC and were thawed immediately before processing.

Test RBCs. The RBCs used for the antibody screening and for the panel identification were obtained from our selected blood donors. These RBCs were suspended to final concentration 0.8 % in the corresponding modified LISS provided by the manufacturers according to their instructions as follows. We calculated the conductivity of these solutions.

DG Gel: 50 µL of RBCs in diluent provided by Diagnostic Grifols (Cond.: 5.09 mS/cm) + 25 µL of sample. Incubation: 15 min, 37 ºC. Centrifugation: 9 min.

DiaMed-ID: 50 µL of RBCs in ID-CellStab (Cond.: 5.89 mS/cm) + 25 µL of sample. Incubation: 15 min, 37 ºC. Centrifugation: 10 min.

Ortho BioVue: 50 µL of RBCs in Ortho BLISS (Cond.: 3.03 mS/cm) + 40 µL of sample. Incubation: 10 min, 37 ºC. Centrifugation: 5 min.

RESULTS Antibody screening

We performed the antibody screening of 3,024 fresh samples with the three microtube systems and we obtained concordant results in 3,019 (99.83%) and discrepant results in 5 (0.17%). Table 1 shows the specificities of the antibody after the identification steps using 11-cell panel and additional techniques, e.g. papain-treated reagent RBCs and IAT with polyethylenglycol (PEG) tube technique (2).

Antibody titration

We titrated 100 frozen samples containing antibodies with known specificities. The score mean and standard desviation of the antibody titration for each microtube system were the 34.31±26.7 for DG Gel, 30.3±27.76 for Diamed-ID, and 37.38±28.68 for Ortho BioVue (see Table 2).

The antibody titration score was higher with the DG Gel when compared with DiaMed-ID in samples containing anti-K (46.23 vs.

44.38; p<0.05) and in samples containing anti-M (11.88 vs. 5.38; p<0.05). No significant differences were seen in other antibodies specificities.

Specificity

Anti-D Anti-C Anti-c Anti-E Anti-e Anti-C w Anti-Ch Anti-Jk a Anti-Jk b Anti-Fy a Anti-Fy b Anti-K Anti-Kp a Anti-Kp b Anti-M Anti-N Anti-s Anti-S Anti-Le a Anti-Le b Anti-H/HI/I Anti-PP 1 P K Anti-Ge:2 Anti-Vel Anti-Xg a

Total n

3 2 7 2 5 7 4 9 10 2 4 13 2 1 8 1 3 5 2 2 3 2 1 1 1

100 Mean DG Gel

74.14

± 24.23

63 ± 16.45

23.67

 19.44

37.40

 23.14

20  0 15.33

 9.6

38.50

 19.09

20.57

 14.50

19  4.24

34  24.87

50.25

 14.68

46.23

 36.73

17.50

 6.36

12 11.88

 4.73

39 24  17.08

34  17.01

8  0 11.50

 2.12

13.33

 13.79

48.50

 34.64

17 46 86

34.31

±

SD score of antibody titration DiaMed-ID

74.43

± 23.86

60.75

± 15.54

23  19.55

33.30

 28.98

13  0 14.67

 9.50

24  8.48

15.86

 11.65

8  0 24.40

 24.70

45.45

 11.14

44.38

 36.72

16.50

 12.02

12 5.38

 2.82

25 24.67

 27.79

30.40

 20.73

4  1.41

5.50

 3.53

12.33

 10.69

41.50

 37.47

17 33 74

30.3

Ortho BioVue

82.29

± 29.35

70 ± 19.71

28.22

 24.80

34.30

 29.33

16.50

 12.02

16  7.21

36  18.38

22.43

 16.20

21  0 35.80

 26.01

52  17.37

50.31

 37.60

26  14.14

19 16.88

 7.69

46 29.67

 18.14

41  21.15

8  7.07

16  1.41

20.67

 14.18

48  29.69

24 43 74

37.38

Rh

Blood Group System

Kell Duffy Kidd MNS Lewis Mixed Nonspecific False positive Negative False negative Total

Antibody specificity

Anti-D Anti-c Anti-E Anti-K Anti-Fy a Anti-Jk a Anti-M Anti-S Anti-Le a Anti-C/-D Anti-E/c/panaglutinin Anti-Jk b /-K Panaglutinin Sensitivity Specificity

CONCLUSION DG Gel

100% (97.07-100) 100% (99.88-100)

DG Gel

6 4 3 1 1 1 1 1 1 2 1 1 0 0 3000 0 3024 Number of Samples Detected

DiaMed-ID Ortho BioVue DiaMed-ID

97.58% (93.09-99.50) 100% (99.88-100) 6 4 1 1 1 0 1 1 1 2 1 1 0 0 3003 3 3024 Table 1. Antibody detected by the three systems using 2 screening cells.

Ortho BioVue

100% (97.07-100) 99.93% (99.76-99.99) PPV( PNV( a b ) ) Efficiency 100% (97.07-100) 100% (99.88-100) 100% (99.88-100) 100% (97-100) 99.9% (99.71-99.98) 99.9% (99.72-99.98) 98.41% (94.38-99.81) 100% (99.88-100) 99.94% (99.77-99.99) Table 3. Estimated diagnostic accuracy. In parenthesis the 95 % CI. (a) PPV: predictive positive value, (b) PNV: predictive negative value.

6 4 3 1 1 1 1 1 1 2 1 1 2 2 2998 0 3024

Sensitivity and Specificity

In order to obtain a comparative value between the new test and the other two we calculated the estimated diagnostic accuracy for each system (see Table 3). For calculating estimates of these statistical measures we include the 100 positive samples and we reported the confidence intervals (CI) for a sample size of 3,124. All three microtube column agglutination systems work well showing a high estimated sensitivity and specificity.

Table 2. Samples containing antibodies of known specificities used for antibody titration.

REFERENCES

1. Lapierre, Y. Rigal, D., Adam, J. Josef, D., Meyer, F., Greber, S. and Drot, C. (1990) Transfusion, 30, 109-113.

2. AABB (2002) Technical Manual, 14 th edition, Beshesda.

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