University of Bahrain College of Science Department of Biology Second Semester- 2011/2012

Lab 4 Lab 4 Agglluttiinattiion Reacttiions: Bllood Typiing Agg u na on Reac ons: B ood Typ ng

Name: Bisma Bashir Mohammad ID # 20094704 Date: March 18, 2012

AIM AIM
To study different types of agglutination reactions e.g. Blood typing, confirmatory test for STDs, and the testing for pregnancy.

INTRODUCTION INTRODUCTION
Blood can be categorized according to the type and Rh factor. There are 4 major blood groups: A, B, O and AB. These can be further divided on the bases of Rh factor as A +, A-, B+, B-, O+, O-, AB+ and AB-. The 4 basic blood groups are explained in the below mentioned table:
Table 1: ABO Blood Group System ABO Blood System A Antigen A Yes [1] If someone belongs to A blood groups, he/she would have A antigen on the surface of RBC and B antibody in blood plasma. B No Yes Yes No B, AB O, B Antigen B No Antibody anti-A No Antibody Anti-B Yes Can give blood to A, AB Can receive blood from O, A

[1] If someone belongs to B blood groups, he/she would have B antigen on the surface of RBC and A antibody in blood plasma. O No No Yes Yes O, A, B, AB O

[1] If someone belongs to O blood groups, he/she would have no antigen on the surface of RBC but would have both A and B antibodies in blood plasma [1]. AB Yes [1] If someone belongs to AB blood groups, he/she would have both A and B antigens on the surface of RBC but no antibodies. Thats why this blood group is considered as universal acceptor [1]. Yes No No AB O, A, B, AB

People with blood group O Rh - are called "universal donors" and people with blood group AB Rh+ are called "universal recivers. Rh+ blood can never be given to someone with Rh -blood, but the other way around works. For example, O Rh+ bloods cannot be given to someone with the blood
Figure 1: Rh System [1]

type AB Rh - . These blood groups are inherited from parents and are located on chromosome 9. An individuals blood type can be determined by the inheritance of 1 of the 3 alleles i.e. A, B and O form each of the parents. Possible results are shown in the table [2].

Table 3: Genotypes of blood groups [2].

Parent Alleles A B O A AA (A) AB (AB) AO (A) B AB (AB) BB (B) BO (B) O AO (A) BO (B) OO (O)

Agglutination reaction is a type of precipitation reaction done to check the type of blood group an individual has. For a blood transfusion to be successful, both the ABO system and Rh system must be compatible otherwise the RBCs from the donor would clump causing them to clog in the blood vessels and stop the circulation of the blood. They can even break open to release their components containing hemoglobin which can be toxic outside the cell. Thus, results in the death of recipient [1]. There are several major applications of agglutination reactions widely used in clinics. Few are mentioned below: 1. To Confirm Pregnancy: Pregnancy can be detected by one of the agglutination reaction called latex agglutination test. In here, urine is checked for the presence of hCG which is Human chorionic gonodrotropin. This hormone is secreted by the placental tissues during pregnancy, and excreted in urine after 20 days of menstrual cycle. It has 2 subunits, namely alpha and beta. For agglutination to occur, the urine is mixed with antibodies which are directed against the beta subunit of hCG and then mixed with latex coupled with hCG and allowed to react. For non-pregnant women, as hCG is unavailable to clump with anti-beta hCG so no agglutination will be observed whereas for pregnant women, agglutination occurs due to the presence of hCG in high concentration [3]. 2. To Check Blood Group: One of a rapid ways to detect the blood group of an individual is by rapid slide agglutination test. Here, a drop of patients blood is placed on a slide next to a drop of stained antigen against which serum is to be tested. These 2 drops are then mixed using a glass rod and observed in an oblique light for clumping. This test is more specific and gives fewer doubtful results [4]. 3. To Detect STD: Diseases in certain individuals can be diagnosed by mixing the patients serum with known antigen. The rheumatoid arthritis test makes use of a form of polystyrene latex and human gamma globulin. When this mixture is mixed with the serum from a victim of rheumatoid arthritis, agglutination will be observed as the patients serum containing antibodies for rheumatoid arthritis would clump with the latex mixture, and positive diagnose the disease can be interpreted [5].

MATERIALS & METHOD MATERIALS & METHOD

Human RBC blood. Human serum. Isotonic solution saline. Different types of anti-bodies.

2 different methods were adopted in this lab to study blood typing. 1 type involved the use of blood (serum + cells) whereas other type used only serum. Method 1: Human RBCs were diluted to 1:5 in an isotonic solution. From that diluted blood sample, 2, 2 drops were taken in 3 different tubes named A, B and D. Their respective antibodies were added in each of the 3 tubes. Latter they were centrifuged at 1000rpm for 2min. and then shaked to observe results. Method 2: from previous identified bloods, type A and Type B were again diluted to 1:5 in an isotonic solution. 2, 2 drops of diluents was again taken into another tubes named A and B and 1 drop of blood serum to identify, was mixed in each tubes and centrifuged at 1000rpm for 2 min. The tubes were again shaked to observe results.

RESULTS RESULTS
In this lab we check the blood groups of few samples and our own blood group by the above mentioned procedures. The results of those tests are given below: From method 1: After centrifugation, following results were obtained. A
Agglutination

B
No Agglutination

D
Agglutination

Blood Group = A+

My blood group obtained from this method was: A

No Agglutination

B
Agglutination

D
Agglutination

Blood Group = B+

From method 2: After centrifugation following results was obtained. Here the known blood group was from type B. A
No Agglutination

B
Agglutination

Blood Group = A

Table 2. Agglutination Reactions of ABO Blood-Typing Sera Reaction Anti-A serum Agglutination No Agglutination Agglutination No Agglutination Anti-B serum No Agglutination Agglutination Agglutination No Agglutination Blood type Type A Type B Type AB Type O

This table shows a detail explanation of each type of blood group obtained when serum treated with a known blood group, as done in part b.

DISCUSSION DISCUSSION
This lab deals with the agglutination reactions for checking blood type of an individual. 2 methods were adopted which were based on ABO cell grouping and ABO serum grouping. In ABO cell grouping, the blood specimen containing RBC were checked for the type of glycoprotein present on them. Here known anti-bodies were mixed with a blood sample. Agglutination with anti-A indicates that A antigen (glycoprotein) is present on RBC and blood is of type A, whereas agglutination with anti-B indicates antigen B is present on the RBC and blood is of type B. This type of ABO grouping is also called direct or forward grouping. 2nd method which deals with ABO serum grouping also called indirect or reverse grouping is one in which the blood samples were of known group and were mixed with unknown serum. Here, agglutination with type A indicated that anti-A is present in the recipients serum and no agglutination means anti-A is not present in the serum.
Figure 2: ABO Blood Grouping [3]

CONCLUSION CONCLUSION
Hence we can conclude that, in case of blood transfusion, it is important to carefully match the donor and recipient blood types. If the donor blood cells have antigens that are different from those of the recipient, antibodies in the recipient's blood recognize the donor blood as foreign. This triggers an immune response resulting in blood clotting. If the donor blood cells have similar antigens as those of the recipient, the recipient's body will not see them as foreign and will not mount an immune response. Also, antigens are specific to each individual.

REFERENCES REFERENCES
1. Blood Groups, Blood Typing and Blood Transfusions (23 Mar 2012): Nobel Media AB Accessed: March 23, 2012. Available: http://www.nobelprize.org/educational/medicine/landsteiner/readmore.html 2. Dennis O'Neil (2011): ABO Blood Types. Accessed date: April 5, 2012. Available: http://anthro.palomar.edu/blood/ABO_system.htm 3. Dr. M. D. Mazumdar (Nk): Pregnancy tests. Accessed: April 3, 2012. Available: http://www.gynaeonline.com/pregnancy_test.htm 4. B. Z. Werbin and A. Kasher (Nk): The rapid slide agglutination test in the diagnosis of typhoid fevers and typhus. Published by: Group.bmj.com 5. Laboratory Tests Interpretation (Nk): Accessed: April 3, 2012. Available: http://www.nurseslearning.com/courses/nrp/labtest/course/section7/index.htm 6. Dr. Mohammed H Saiem Aldahr (Nk): ABO Blood Grouping. Faculty of Applied Medical Sciences. Blood Bank, Medical Technology, Kaau