Sistem Penggolongan Darah

                                                                                                           

             

   Efrida, dr., SpPK., MKes4 Desember 2014

4

Tujuan Pembelajaran

Mampu menjelaskan:1.Dasar pembagian golongan darah (terutama

sistem ABO dan Rhesus)2.Prinsip pemeriksaan golongan darah ABO dan

Rhesus3.Kepentingan menentukan golongan darah

Dasar Pembagian Golongan DarahDasar Pembagian Golongan Darah

Konsep dasar Imunohematologi:Konsep dasar Imunohematologi:

1.1.Seroimunologi, genetik, biokimia Seroimunologi, genetik, biokimia

2.2.Studi molekular antigen terkait dengan Studi molekular antigen terkait dengan struktur membran pada komponen selular struktur membran pada komponen selular darah darah

3.3.sifat-sifat serta reaksi imunologi seluruh sifat-sifat serta reaksi imunologi seluruh komponen darah dan konstituennyakomponen darah dan konstituennya

Susunan Darah

                      

Volume darah dewasa: 4–6 literTerdiri atas plasma dan sel-sel darah (eritrosit, leukosit, trombosit)

Eritrosit/the red blood cells (RBCs): mengandung hemoglobin, suatu protein yang mengikat oksigen. RBCs mentranspor oksigen ke jaringan dan mengangkut karbondioksida dari jaringan.

Leukosit/the white blood cells: respons imun selular terhadap infeksi.

Trombosit/the platelets: peran pada hemostasis primer (membentuk sumbat trombosit pada luka pembuluh darah)

Plasma: mengandung berbagai substansi terlarut (protein, lipid, elektrolit) 

Eritrosit• Struktur

– Bikonkaf, anukleat

• Komponen– Hemoglobin– Lipid, ATP, karbonik

anhidrase

• Fungsi– Transpor oksigen dari

paru ke jaringan dan karbondioksida dari jaringan ke paru-paru

19-5

ABO antigen BiochemistryABO antigen Biochemistry

• CarbohydrateCarbohydrate• ABH antigens ABH antigens

expressed on expressed on RBC RBC glycoproteins & glycoproteins & glycosphingolipiglycosphingolipid (type 2,3,4 d (type 2,3,4 chain) chain) RBC RBC origin.origin.

Type 1 chain are synthesized by gastrointestinal mucosa Type 1 chain are synthesized by gastrointestinal mucosa secreted into plasma secreted into plasma passively adsorbed onto RBC membranepassively adsorbed onto RBC membrane

Glikoprotein dan glikolipid pada permukaan eritrosit:- substansi selular bersifat spesifik- diatur secara genetik (Mendelian)- bersifat imunogenik- mampu merangsang pembentukan aloantibodi spesifik bila masuk ke dalam tubuh seseorang yg tidak memiliki substansi tsb

Blood Grouping

• Determined by antigens (agglutinogens) on surface of RBCs

• Antibodies (agglutinins) can bind to RBC antigens, resulting in agglutination (clumping) or hemolysis (rupture) of RBCs

19-10

Erythrocyte Antigens & Erythrocyte Antigens & AntibodiesAntibodies

• > 700 antigens > 700 antigens organized into 29 blood organized into 29 blood group systems by the International Society group systems by the International Society of Blood Transfusion (ISBT). of Blood Transfusion (ISBT).

• Sistem penggolongan terpenting:1. ABO2. Rhesus

Sistem lain diperiksa jika antigennya menimbulkan reaksi transfusi yg tidak disebabkan gol ABO atau Rhesus, misal:- Sistem Kell- Sistem Duffy dan Kidd- Sistem MN, P, Lewis, Lutheran (reaksi transfusi jarang/ringan)

Dasar Pembagian Golongan Darah Sistem ABO

• 1901: Landsteiner menemukan 2 jenis antigen pada permukaan eritrosit- antigen (aglutinogen) A- antigen (aglutinogen) B

Pengamatannya berdasarkan:Reaksi aglutinasi eritrosit dan adanya antibodi (aglutinin) pada serum seseorang yang bereaksi langsung terhadap antigen eritrosit

ABO AntigensABO Antigens

• Also express in many tissues, body fluids, platelet Also express in many tissues, body fluids, platelet and endotheland endothel

• Most important blood group system in Most important blood group system in transfusion and organ transplantation.transfusion and organ transplantation.

• 3 antigens: A, B, H (biosynthetic precursor of A & 3 antigens: A, B, H (biosynthetic precursor of A & B antigens).B antigens).

• A & B: autosomal codominant antigens A & B: autosomal codominant antigens expressed on group A, B & AB RBCexpressed on group A, B & AB RBC

ABO AntibodiesABO Antibodies

• Weak or absent in newborn Weak or absent in newborn 3-6 mo 3-6 mo• 5-10 yo 5-10 yo adult level adult level• Advancing age Advancing age slight decrease slight decrease• Detected at room temperature, with optimal reactivity at Detected at room temperature, with optimal reactivity at

4400C.C.• Merupakan suatu IgM atau IgG atau IgAMerupakan suatu IgM atau IgG atau IgA• IgG (reactive at 37IgG (reactive at 3700C) can occur after transfusion/pregnancy; C) can occur after transfusion/pregnancy;

higher titerhigher titer• Can fix complement Can fix complement hemolysis in vivo/vitro hemolysis in vivo/vitro • Can cause: hemolytic transfusion reaction & hemolytic Can cause: hemolytic transfusion reaction & hemolytic

disease of the new born.disease of the new born.

Less common ABO antibodiesLess common ABO antibodies

• Anti-AAnti-A11

• Anti-HAnti-H

Cont….

• Berdasarkan antigen A,B pada permukaan eritrosit dan antibodi ABO pada plasma Sistem ABO (golongan darah ABO)

• Von Decastello dan Sturli menemukan golongan darah ke-4: AB

• Penemuan selanjutnya: 2 jenis antigen A (A1 dan A2)6 fenotipe golongan darah

Cont…

Golongan darah ABO1. A1 (genotipe A1/A1, A1/A2, A1/O)2. A2 (genotipe A2/A2, A2/O)3. A1B (genotipe A1/B)4. A2B (genotipe A2/B)5. B (genotipe B/B, B/O)6. O (genotipe O/O)

              

Blood group AIf you belong to the blood group A, you have A antigens on the surface of your RBCs and B antibodies in your blood plasma.

 

               Blood group BIf you belong to the blood group B, you have B antigens on the surface of your RBCs and A antibodies in your blood plasma.

AB0 blood grouping system

                

              

              

Blood group ABIf you belong to the blood group AB, you have both A and B antigens on the surface of your RBCs and no A or B antibodies at all in your blood plasma.

 

              

Blood group OIf you belong to the blood group O (null), you have neither A or B antigens on the surface of your RBCs but you have both A and B antibodies in your blood plasma.

• The ABO gene is autosomal (the gene is not on either sex chromosomes)

• The ABO gene locus is located on the chromosome 9.

• Each person has two copies of genes coding for their ABO blood group (one maternal and one paternal in origin)

• A and B blood groups are dominant over the O blood group

• A and B group genes are co-dominant

ABO inheritance and genetics

AUTOSOMAL CHROMOSOME

father

mother

one alleles from father and one from mother

The alleles for Blood group are in the same

place on the chromosome 9.

However the genes have a different code giving the different

blood group

A B

Parent

Allele

A B O

A

B

O

Possible Blood group Genotypes

Parent

Allele

A B O

A AA AB AO

B AB BB BO

O AO BO OO

Possible Blood group Genotypes

The ABO blood groups

• The table shows the four ABO phenotypes ("blood groups") present in the human population and the genotypes that give rise to them.

Blood Group

Antigens on RBCs

Antibodies in Serum Genotypes

A A Anti-B AA or AO

B B Anti-A BB or BO

AB A and B Neither AB

O Neither Anti-A and anti-B OO

Golongan darah sistem Rhesus (Rh)

Cont…

• Golongan darah sistem Rhesus:1.Rh + (DD, Dd)2.Rh – (dd)

Golongan Darah Rhesus

• Ciri-ciri antigennya mirip dengan sistem ABO• Antibodi imun (tidak alamiah), terbentuk kr

reaksi imunologik (transfusi atau kehamilan/transfusi fetomaternal)

• Dapat menyebabkan reaksi ketidakcocokan pada transfusi atau kehamilan

• Fisher dan Race: sistem rhesuscampuran antigen yg kompleks, diatur oleh very closely linked genes pada kromosom 1

Cont…• 3 kelompok gen: C, D, E• Tiap kelompok bersamaan dg beberapa alel

- C dg c dan cw- D dg d dan Du- E dg e

Ag D: terkuat (sangat imunogenik)Ag Du: subgrup lemah Ag D (deteksi dg

antiglobulin tdk langsung, setelah eritrosit diinkubasi dengan anti-D)

The Rh(D) Antigen

• RH is the most complex system, with over 45 antigens

• Discovered in 1940 after work on Rhesus monkeys

• Subsequently discovered to be unrelated to monkeys

• RH gene located on short arm of chromosome 1

ABO & Rh(D) 32

Simple Genetics of Rh(D)

• 86% of caucasians are Rh(D) pos• The antithetical antigen d has not been found• The d gene is recessive:

– Dd, dD, DD, persons are Rh(D) pos– Only dd persons are Rh(D) neg

ABO & Rh(D) 33

Distribution of Rh(D) Types

Population Rh(D) pos Rh(D) neg

Caucasian 86% 14%

African-American

95% 5%

Oriental >99% <1%

ABO & Rh(D) 34

Significance of Rh(D)

• 80% of Rh(D) neg persons exposed to Rh(D) pos blood will develop anti-D

• Anti-D can also be stimulated by pregnancy with an Rh(D) positive baby– Sensitisation can be prevented by the use of anti-D

immunoglobulin, antenatally and post natally

• Rh(D) neg females of childbearing potential should never be given Rh(D) positive blood products

ABO & Rh(D) 35

Inheritance

• ABO & RH genes are not linked• ABO & Rh(D) type are inherited independentlyFor example:

An A Rh(D) positive mother and a B Rh(D) positive father could have an O Rh(D) negative child

ABO & Rh(D) 36

Inheritance of ABO and Rh(D)

ABO & Rh(D) 37

Mother

Group A AO

Rh(D) pos Dd

Father

Group B BO

Rh(D) pos Dd

Group A AO

Rh(D) pos Dd

Group B BO

Rh(D) pos Dd

Group O OO

Rh(D) neg dd

BloodType

GenotypeAllelesProduced

Rh positiveDD D

Dd D or d

Rh negative dd d

Rh Blood Group and Rh Incompatibility

A person with Rh- blood does not have Rh antibodies naturally in the blood plasma

According to above blood grouping systems, you can belong to either of following 8 blood groups:

Do you know which blood group you belong to?

                                                                           

• A person with Rh- blood can develop Rh antibodies in the blood plasma if he or she receives blood from a person with Rh+ blood, whose Rh antigens can trigger the production of Rh antibodies.

•A person with Rh+ blood can receive blood from a person with Rh- blood without any problems.

Aplikasi Klinis Sistem Penggolongan Darah

History of Blood Groups and Blood Transfusions

                     

•Experiments with blood transfusions have been carried out for hundreds of years. Many patients have died and it was not until 1901, when the Austrian Karl Landsteiner discovered human blood groups, that blood transfusions became safer.

• He found that mixing blood from two individuals can lead to blood clumping. The clumped RBCs can crack and cause toxic reactions. This can be fatal.

 http://nobelprize.org/medicine/educational/landsteiner/readmore.html

• Karl Landsteiner discovered that blood clumping was an immunological reaction which occurs when the receiver of a blood transfusion has antibodies against the donor blood cells.

•Karl Landsteiner's work made it possible to determine blood types and thus paved the way for blood transfusions to be carried out safely. For this discovery he was awarded the Nobel Prize in Physiology or Medicine in 1930.

History of Blood Groups and Blood Transfusions (Cont.)

Kepentingan Klinis

• Transfusi darah dan transplantasi organ• Identifikasi dan pencegahan terhadap

aloimunisasi ibu hamil oleh antigen Rhesus• Menentukan diagnosis, prognosis, dan terapi

HDN (hemolytic disease of the newborn) akibat aloantibodi

• Diagnosis dan pemeriksaan destruksi eritrosit yg disebabkan autoantibodi atau aloantibodi

• Kedokteran kehakiman• antropologi

Prinsip Pemeriksaan Golongan Darah Sistem ABO dan Rhesus

Prinsip: Imunoserologi (Reaksi Antigen-Antibodi)

• Membran Eritrosit: protein dan karbohidrat

Antigen Eritrosit

Antibodi spesifik

Penampilan Golongan Darah ABO

• Uji golongan darah harus dilakukan terhadap :– Eritrosit– Serum

• Pada keadaan meragukan harus diulang

Several methods for testing the ABO group of an individual exist. The most common method is:

Serology: This is a direct detection of the ABO antigens. It is the main method used in blood transfusion centres and hospital blood banks.

This form of testing involves two components:

a) Antibodies that are specific at detecting a particular ABO antigen on RBCs (forward method)         b) Cells that are of a known ABO group that are agglutinated by the naturally occurring antibodies in the person's serum (reverse methods)

Pemeriksaan Golongan Darah ABO

1. Forward Grouping - Menentukan gol darah ABO berdasarkan antigen pada eritrosit - Aglutinasi antigen bereaksi dengan antisera (IgM anti-A atau anti-B). Anti-A antigen A Anti-B antigen B

When RBCs carrying one or both antigens are exposed to the corresponding antibodies, they agglutinate; that is, clump together. People usually have antibodies against those red cell antigens that they lack.

Human RBC before (left) and after (right) adding serum containing anti-A antibodies. The agglutination reaction reveals the presence of the A antigen on the surface of the cells.

                                    

    

http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/B/BloodGroups.html

                                   

         

Anti AB dan Anti A

• Anti-AB dianjurkan diuji pada penggolongan darah donor

• Anti-A mempunyai gabungan 2 antibodi– Anti-A yang mengaglutinasi sel-sel A1, A2, A1B

dan A2B– Anti-A1 yang hanya mengaglutinasi sel-sel A1 dan

A1B

Prosedur kerja: Dibuat suspensi eri 2-4% dg NaCl 0.9%Disediakan 3 tabung & diberi label: A, B, & ABTabung A di(+) 1 tetes antiserum anti-ATabung B di(+) 1 tetes antiserum anti-BTabung AB di(+) 1 tetes antiserum anti-ABMasing2X tabung di(+) 1 tetes suspensi eri Ketiga tabung disentrifus 15” dg 3400 rpmResuspensi & dilihat ada aglutinasi/ tidak

Interpretasi hasil:

Antisera Interpretasi Gol Darah

Anti-A Anti-B Anti-AB

0 0 0 O

+ 0 + A

0 + + B

+ + + AB

0 0 + Subgolongan A lemah

Keterangan:

+ = Aglutinasi

0 = Tidak ada aglutinasi

2. Reverse Grouping

- Menentukan gol darah ABO berdasarkan ada/tidak adanya antibodi - Aglutinasi antibodi dalam serum

bereaksi dengan antigen A/B pd eritrosit

Prosedur kerja:Dua buah tabung diberi label: A, dan BMasing2x tabung di(+) 2 tetes serum/ plasma yg akan diperiksaKe dalam tabung A di(+) 1 tetes reagen eri A1Ke dalam tabung B di(+) 1 tetes reagen eri BKedua tabung disentrifus 15 ” dg 3400 rpmResuspensi & dilihat ada aglutinasi/ tidak

Interpretasi hasil:

A1 B Antibodi Golongan Darah

+ + Anti-A & Anti-B O

0 + Anti-B A

+ 0 Anti-A B

0 0 Tidak ada AB

Keterangan:

+ = Aglutinasi

0 = Tidak ada aglutinasi

• Illustration of the forward and reverse grouping reaction patterns of the ABO groups using a blood group tile

http://www.bh.rmit.edu.au/mls/subjects/abo/resources/genetics1.htm

Metode Pemeriksaan

Metode SlideProsedur kerja:Pemeriksaan forward grouping: Pada kaca objek ditaruh 1 tetes reagen anti-A, anti-B, & anti-AB Darah diteteskan pada reagen itu & dicampur Kaca objek digoyang perlahan-lahan Dilihat apakah ada aglutinasi

Pemeriksaan reverse grouping:Pada kaca objek ditaruh 1 tts reagen eri A1&BSerum/ plasma diteteskan pd reagen itu & dicampur dengan ujung lidiKaca objek digoyang perlahan-lahan

Dilihat apakah ada aglutinasi

Interpretasi Hasil: Forward Grouping

Antisera Golongan Darah

Anti-A Anti-B Anti-AB

0 0 0 O

+ 0 + A

0 + + B

+ + + AB

Keterangan:

+ = Aglutinasi

0 = Tidak ada aglutinasi

Reverse Grouping

A B Golongan Darah

+ + O

0 + A

+ 0 B

0 0 AB

Keterangan:

+ = Aglutinasi

0 = Tidak ada aglutinasi

Keuntungan: Cepat, mudah, sederhana, memerlukan sedikit sampel.

Kerugian: 1) tidak dpt menentukan aglutinasi lemah 2) tidak ada kontrol (+), kontrol (-), & autokontrol

Metode Tube Pemeriksaan eri & serum scr bersamaan. Pemeriksaan atas eritrosit: Dipakai 12 tbg kecil, msg2x diisi 1 tts NaCl 0.9% Di(+) 1 tetes susp 2% eri X, eri A, eri B, & eri O pada masing-masing baris Di(+) 1 tetes anti-A, anti-B, & anti-AB pd masing masing kolom, dicampur Hasil dibaca setelah 2 jam

Pemeriksaan atas serum:

Dipakai 4 tabung reaksi kecilTiap tabung diisi 1 tetes NaCl 0.9%Tabung ke-1 ditambah suspensi 2% eri ATabung ke-2 ditambah suspensi 2% eri BTabung ke-3 ditambah suspensi 2% eri X Tabung ke-4 ditambah suspensi 2% eri OEritrosit & zat anti dicampur Hasil dibaca setelah 2 jam

Interpretasi hasil: Golongan Darah A

Atas Eri Atas Serum

+ O + Eri X O Eri A

+ O + Eri A + Eri B

O + + Eri BAuto

kontrolO Eri X

O O O Eri O O Eri O

Anti A Anti B Anti AB

Interpretasi hasil: Golongan Darah B

Atas Eri Atas Serum

O + + Eri X + Eri A

+ O + Eri A O Eri B

O + + Eri BAuto

kontrolO Eri X

O O O Eri O O Eri O

Anti A Anti B Anti AB

Interpretasi hasil: Golongan Darah AB

Atas Eri Atas Serum

+ + + Eri X O Eri A

+ O + Eri A O Eri B

O + + Eri BAuto

kontrolO Eri X

O O O Eri O O Eri O

Anti A Anti B Anti AB

Interpretasi hasil: Golongan Darah O

Atas Eri Atas Serum

O O O Eri X + Eri A

+ O + Eri A + Eri B

O + + Eri BAuto

kontrolO Eri X

O O O Eri O O Eri O

Anti A Anti B Anti AB

Keuntungan:1) Pemeriksaan eritrosit dan serum dilakukan secara bersamaan2) Ada kontrol (+), kontrol (-), dan autokontrol3) Dapat menentukan aglutinasi lemah4) Dapat dilakukan pemeriksaan imun antibodi

Kerugian:Sukar & memerlukan waktu lama

Metode Gel Test

Didasarkan reaksi antigen & antibodiReaksi antigen & antibodi terjadi di dalamtabung kecil yang berisi gelPartikel aglutinasi dipisahkan dari partikelnon-aglutinasi oleh gel.

Prosedur kerja:

Dibuat suspensi sel (tanpa dicuci) dg ID Diluent (Modified Low Ionic Strength Solution), 500 L diluent:5 L darah, dicampur

Suspensi sel dipipet ke dlm tabung berisi gel yang mengandung antibodi spesifik: 50 L

ID Card diinkubasi 37C, 15 menit ID Card disentrifus selama 10 menitHasil Reaksi dibaca secara makroskopis

Interpretasi hasil:

+4 : Menggumpal & menyatu shg cairan tampak jernih+3 : Menggumpal tp tdk menyatu. Jd tdd beberapa gumpalan kasar, sekitarnya tampak cairan jernih+2 : Gumpalan agak kasar tp tdk semua sel darah beraglutinasi, tampak cairan agak keruh+1 : Gumpalan halus, tampak cairan keruh

Keuntungan: Sederhana, mudah Hasil reaksi stabil Tdk ada fase pencucian Hanya memerlukan sedikit sampel Pembacaan reaksi secara makroskopis Masa kadaluarsa reagen panjang Mengurangi limbah Meningkatkan keamanan laboratorium

Kerugian:Memerlukan peralatan yang mahal Memerlukan waktu > lama dr M. slide