Immunohema Lab Activities

IMMHMA1: Immunohematology
Name: Rating:
Laboratory Schedule: Group Number:
Date Performed: Date Submitted:
Experiment Number 1
BLOOD BANKING LABORATORY INSTRUMENTATION
OBJECTIVES:
At the end of this experiment, the students will be able to:
▪ Recognize the different glasswares, apparatus and equipment used for blood banking procedures
▪ Understand the functions and describe the proper maintenance of the instruments used in blood bank
MATERIALS NEEDED:
1. Glasswares:
a. Plain test tubes (15 x 100 mm, 13 x 100 mm, 10 x 75 mm, and 8 x 75mm)
b. Serological pipettes (0.1 mL, 0.2 mL, 1.0 mL, 2.0 mL, 5.0 mL, and 10.0 mL)
c. Evacuated tubes (red, pink and lavender top)
d. Graduated centrifuge tube
e. Blood typing glass slide
f. Glass slide (ordinary)
g. Petri dish cover
h. Pasteur pipette
i. Beaker (50 mL)
j. Capillary tube
2. Blood Sampling Set:

a. Tourniquet c. Hypodermic syringe e. 2-way needle and
b. Hypodermic Needle d. Lancet and autolancet holder/adapter
3. Blood Collection Set:

a. Blood bag (single, triple and quadruple blood bag) and hemostat
b. Stethoscope and sphygmomanometer (aneroid BP apparatus)
4. Electrical Equipment:
a. Compound microscope d. Clinical sterilizer f. Rh viewbox
b. Mechanical rotator e. Hot air oven g. Water bath
c. Clinical centrifuge
5. Others:
a. Applicator Stick b. Wash bottle
PROCEDURE: The instructor shall present and discuss the important functions and features of the instruments
DRAWING: Illustrate and label accordingly all the above glasswares, apparatus and equipment (include descriptions,
functions and/or uses in blood banking)
QUESTIONS FOR RESEARCH:

1. Describe the functions of each of the instruments according to use in blood banking laboratory
2. Enumerate several points to consider in the quality control of the instruments used in blood banking
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Name: Rating:
Experiment Number 2
BLOOD SAMPLING BY ROUTINE VENIPUNCTURE AND CAPILLARY METHOD
***The procedure shall be presented by the instructor thru a video conference/recorded video or via virtual lab
presentation in which the students can watch and observe
Phlebotomy – the drawing of blood – is an integral part of medical laboratory practice. It is good to remember that no
laboratory procedure will be any better than the quality of the specimen that is being tested. Each step in the process of
phlebotomy affects the quality of the specimen and is this important for preventing laboratory error and patient injury.
OBJECTIVES:
▪ Demonstrate necessary skills and perform proper venipuncture and capillary puncture procedures
▪ Appraise the importance of proper specimen collection for blood banking tests
MATERIALS NEEDED:
2-way needle and holder/adapter Evacuated tubes
Hypodermic syringe and needle Plain test tubes
Plaster tape (Microscope) Capillary tube
Lancet and autolancet Cotton balls
70% isopropyl alcohol Tourniquet
PROCEDURE:
Venous puncture technique (Venipuncture)
1. Verify requisition and check patient identification. Ask the patient for his or her full name, address and/or date of birth.
2. If a fasting specimen or a dietary restriction is required, confirm patient has fasted or eliminated foods from diet as
ordered by physician.
3. Position the patient properly. Assemble equipment and supplies.
4. Apply a tourniquet and ask the patient to make a fist without vigorous hand pumping. Select a suitable vein for
puncture.
5. Put on gloves with consideration of latex allergy for the patient.
6. Cleanse the venipuncture site with 70% isopropyl alcohol. Allow the area to dry.
7. Anchor the vein firmly.
8. Enter the skin with the needle at approximately a 15 – 30° angle or less to the arm, with the bevel of the needle up:
a. Follow the geography of the vein with the needle.
b. Insert the needle smoothly and fairly rapidly to minimize patient discomfort.
c. If using a syringe, pull back on the barrel with a slow, even tension as blood flows into the syringe. Do not pull
back too quickly to avoid hemolysis or collapsing the vein.
d. If using an evacuated system, as soon as the needle is in the vein, ease the tube forward in the holder far as it will
go, firmly securing the needle holder in place. When the tube is filled, remove it by grasping the end of the tube
and pulling gently to withdraw, and gently invert tubes containing additives.
9. Release the tourniquet when blood begins to flow. Never withdraw the needle without removing the tourniquet.
10. Withdraw the needle, and then apply pressure to the site. Apply adhesive bandage strip over a cotton ball or gauze
to adequately stop bleeding and to avoid a hematoma.
11. Mix and invert tubes with anticoagulant; do not shake the tubes. Check condition of the patient. Dispose of
contaminated material in designated containers (sharps container) using Universal Precautions.
12. Label the tubes before leaving patient’s side with:
a. Patient’s first and last name
b. Identification number
c. Date of collection
d. Time of collection
e. Identification of person collecting specimen
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13. Deliver tubes of blood for testing to appropriate laboratory section or central receiving and processing area.
Skin Puncture Technique (Capillary Puncture)

1. Verify requisition form and check patient identification. Ask the patient for his or her full name, address and/or date of
birth.
2. Position the patient properly. Assemble equipment and supplies.
3. Select and appropriate puncture site.
a. For infants younger than 12 months old, this is most usually the lateral or medial plantar heel surface.
b. For infants older than 12 months, children, and adults, the palmar surface of the last digit of the second, third, or
fourth finger may be used.
c. The thumb and fifth finger must not be used, and the site of puncture must not be edematous or a previous
puncture site because of accumulated tissue fluid.
4. Warm the puncture site with a warm, moist towel no hotter than 42°C; this increases the blood flow through arterioles
and capillaries and results in arterial-enriched blood.
5. Cleanse the puncture site with 70% aqueous isopropanol solution. Allow the area to dry. Do not touch the swabbed
area with any nonsterile object.
6. Make the puncture with a sterile lancet or other skin-puncturing device, using a single deliberate motion nearly
perpendicular to the skin surface. For a heel puncture, hold the heel with the forefinger at the arch and the thumb
proximal to the puncture site at the ankle. If using a lancet, the blade should not be longer than 2 mm to avoid injury to
the calcaneus (heel bone).
7. Discard the first drop of blood by wiping it away with a sterile pad. Regulate further blood flow by gentle thumb
pressure. Do not milk the site, as this may cause hemolysis and introduce excess tissue fluid.
8. Collect the specimen in a suitable container by capillary action.
a. Closed systems are available for collection of non-anticoagulated blood and with additives for whole blood
analysis.
b. Open-ended, narrow-bore disposable glass micropipettes are most often used up to volumes of 200 mL.
c. Both heparinized and non-heparinized micropipettes are available.
d. Use the appropriate anticoagulant for the test ordered. Mix the specimen as necessary.
9. Apply pressure and dispose off the puncture device.
10. Check condition of the patient. Dispose off contaminated materials in designated containers (sharps container) using
Universal Precautions.
11. Label the specimen container with date and time of collection and patient demographics.
12. Indicate in the report that test results are from skin puncture.
DRAWINGS:
1. Illustrate and label accordingly the important steps of venipuncture.
2. Illustrate and label accordingly the important steps of capillary puncture.

1. What types of evacuated tubes are used for blood banking procedures? Give brief description according to
composition and use
2. Enumerate preferred venous access sites and factors to consider during site selection
3. Differentiate the feel of vein, tendon, and artery
4. Factors to be considered to avoid hemolysis during blood sampling
5. Enumerate the advantages and disadvantages of (a) venipuncture and (b) capillary puncture
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Name: Rating:
Experiment Number 3
SAMPLE PREPARATION FOR BLOOD BANKING PROCEDURES
Many procedures done in the laboratory require demonstration of in vitro antigen and antibody reactions. Thus, preparing
plasma and serum as antibody source and red cell suspension as antigen and/or indicator cell source is needed. Red cell
suspension (RCS) is prepared using an anticoagulated blood and 2 – 5% concentration is universally employed for
serological and blood banking procedures.
OBJECTIVES:
▪ Identify the steps of preparing samples used for blood banking procedures
▪ Describe the characteristics of samples that are accepted for blood banking tests
▪ Properly manipulate the equipment used for preparation of samples used for blood banking
▪ Display proper pipetting techniques for preparing red cell suspensions
MATERIALS NEEDED:
Centrifuge Test tube and rack
Aspirator bulb Graduated centrifuge tube
Pasteur pipette Evacuated tubes (red and lavender top)
Venipuncture set Serological pipette (0.1 mL, 2.0 mL, 5.0 mL, 10.0 mL)
REAGENT:
Normal Saline Solution (0.85 – 0.90% NaCl)
PROCEDURE:
1. Whole Blood preparation
a. Draw a sufficient amount of blood corresponding to the volume of the additive/anticoagulant
b. To achieve an optimum ratio of blood to additive/anticoagulant, the volume of blood should
(1) Fill the tube to the line indicated on the evacuated tube label, or
(2) Be according to the appropriate ratio of the sample to the additive/anticoagulant
c. Mix evacuated tube immediately after collection, before clotting can occur, gently by inversion according to the
type of additive/anticoagulant
d. Label the specimen container with patient information including the date and time of collection
2. Plasma Preparation
a. Draw a sufficient amount of blood into an evacuated tube with the required anticoagulant to yield the plasma
volume required by the test.
b. Gently mix immediately after collection by appropriate number of inversions according to the type of anticoagulant
used. Avoid hemolysis of the specimen during collection and mixing.
c. Place the specimen in a rack at room temperature and centrifuge within 2 hours of collection. Do not refrigerate
the sample until the plasma is separated from the cells.
d. Samples should undergo centrifugation immediately. Centrifuge anticoagulated blood for 5 – 10 minutes at 2,500
rpm or refer to speed and duration recommended by manufacturer of the evacuated tubes. Do not use brake to
stop centrifuge.
e. After centrifugation, three layers are formed: (from top to bottom) plasma, buffy coat, red blood cells. The plasma
should appear clear and no pink to red tinge is manifested.
f. Carefully aspirate or collect the supernatant (plasma) with a Pasteur pipette and transfer to a test tube.
g. Label the sample container.
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3. Serum Preparation
a. Draw a sufficient amount of blood to yield the serum volume required by the test.
b. Allow the tube containing venous blood to stand in an upright position at room temperature for 20 – 30 minutes
(no longer than 60 min) to allow complete clotting to occur. Centrifuging specimens before coagulation is
complete causes fibrin clots to form in the serum.
(1) If using a clot activator tube, invert carefully 5 – 6 times to mix clot activator and blood.
(2) If using a serum separator tube (SST), mix tubes well by carefully inverting the collection tube 8 – 10 times for
clot formation to occur.
Avoid hemolysis of the specimen during collection and mixing.
c. After 20 – 30 minutes, check for complete clotting by inclining the tube gently and when blood does not ooze from
the clot, coagulation time is reached.
d. Rim or ring the sides of the clot by thrusting the applicator stick to a depth of 1 cm and perform full turn at the
sides of the clot gently. Do this once.
Note: If the clot clings to the stick, dislodge the clot from the stick carefully and include the clot for centrifugation.
Do not discard the clot.
e. Centrifuge clotted blood for 5 – 10 minutes at 2,500 rpm or refer to speed and duration recommended by
manufacturer of the evacuated tubes. Do not use brake to stop centrifuge.
Note: Centrifuge within 2 hours of collection. Do not refrigerate the sample until the serum is separated from the
cells.
f. After centrifugation, check the supernatant (serum) and carefully aspirate it with a Pasteur pipette and transfer to
a test tube. Take care not to disrupt the packed red blood cell layer or transfer any cells.
g. Inspect serum for turbidity. Turbid samples should be re-centrifuged to remove remaining insoluble matter. Serum
should appear clear and no pink to red tinge is manifested.
h. Label the sample container.
4. Red Cell Suspension Preparation (2 – 5%)

Method A:
a. Transfer carefully ≤1.0 mL of the packed red blood cells from plasma preparation into a graduated centrifuge
tube.
b. Washing phase:
(1) Add normal saline solution (NSS) up to 10 mL mark of the centrifuge tube
(2) Cover the mouth of centrifuge tube using Parafilm then mix by gentle inversion, 2 – 3 times
(3) Centrifuge the tube for 5 – 10 minutes at 2,500 rpm then discard the supernatant fluid using a Pasteur pipette,
leaving the red blood cells.
(4) Repeat steps 1 → 2 → 3 two or three times until the supernatant is clear.
c. After the final washing, note the volume of the washed packed red blood cells before the supernatant is
completely removed by aspiration.
d. Compute for the total volume (TV) of the RCS preparation and the volume of the diluent (NSS) to be added to the
washed red blood cells to come up with the desired concentration (in %) of the RCS using the following formulas:
✓ Desired % red cell suspension (RCS) = Washed packed red blood cell (WPRBC) volume x 100
Total volume (TV) of the RCS preparation
✓ Total volume (TV) of RCS = WPRBC volume + volume of diluent (NSS) added to prepare desired %RCS
e. Add the appropriate volume of NSS to the WPRBC to prepare the desired concentration (in %) of the RCS
f. The suspension is mixed thoroughly to produce a homogeneous solution. Check for the presence of clots in the
suspension.
g. Label the RCS according to the concentration and the blood (antigen) type.
Method B:
a. Preparation of washed packed red blood cell
(1) Using a Pasteur pipette, transfer 2 mL of blood into a plain test tube
(2) Fill the tube half-full with NSS
(3) Cove the tube with Parafilm and mix by gentle inversion
(4) Centrifuge for 1 minute at 3,400 rpm
(5) Remove the parafilm then aspirate the supernatant using Pasteur pipette this can also be done by discarding
the supernatant as quickly as possible in order not to disturb the cell button.
(6) Repeat steps 2 → 3 → 4 → 5 three times (washing phase)
(7) After the last washing, completely aspirate the supernatant using a Pasteur pipette.
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b. Preparation of tube with fixed volume of fluid

(1) Using a 0.1 mL serological pipette, aspirate NSS up to the highest mark then calibrate the volume at 0.1
mark.
(2) Deliver exactly 0.1 mL into a plain test tube (use plain test tubes with identical size for the whole procedure)
(3) Mark the level. The volume in this tube shall be used for comparing drops administered that will be equivalent
to 0.1 mL.
c. Preparation of RCS for various concentration

(1) Label the 4 plain test tubes (with identical size) as 2% RCS, 3% RCS, 4% RCS, 5% RCS
(2) Using only the Pasteur pipette, deliver drops of the washed packed red blood cell to the labeled tubes until it
reaches a volume equal to 0.1 mL by comparing it to the volume of the tube prepared in step b (preparation of
tube with fixed volume of fluid)
(3) Using a 2.0 mL or 5.0 mL serological pipette, deliver exactly the following required volume of NSS to the
labeled tubes containing 0.1 mL if washed packed red blood cells:
▪ 2% RCS → add 4.9 mL NSS
(4) Cover the tubes with Parafilm then mix gently. These are the red cell suspensions at different concentrations.
Note: A proper red cell suspension has a tomato red color
DRAWINGS:
1. Illustrate and label accordingly the important steps in whole blood preparation.
2. Illustrate and label accordingly the important steps in plasma separation.
3. Illustrate and label accordingly the important steps in serum preparation.
4. Illustrate and label accordingly the important steps in red cell suspension (RCS) preparation using method A (include
the computation) and method B.

1. Why are hemolyzed, lipemic and icteric samples not preferred for testing in a blood bank?
2. When are icteric samples used for blood banking procedures? Discuss the rationale
3. Why is serum the preferred sample for immunohematologic testing?
4. Why is red cell suspensions prepared in 2 – 5% concentration?
5. Why is NSS chemically? Why should you use it in washing red cells?
6. Why is it necessary to wash the red cells with NSS for three times prior to their use?
7. Why is a Pasteur pipette used in delivering 0.1 mL of washed packed red blood cells instead of a serological
pipette?
8. What is the importance of preparing red cell suspension in the laboratory?
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Name: Rating:
Experiment Number 4
DEMONSTRATION OF COMMON IMMUNOHEMATOLOGIC REACTIONS
Hemagglutination and hemolysis are common and clinically significant serologic reactions observed in blood banking
procedures and are considered as positive results demonstrating the presence of antigen – antibody interactions.
Hemagglutination is the clumping together of red blood cells resulting from interaction of red blood cell antigen and its
corresponding antibody. On the other hand, hemolysis is the disruption or destruction of the red blood cell membrane and
the subsequent release of hemoglobin into the suspending medium.
OBJECTIVES:
▪ Demonstrate agglutination reaction macroscopically and microscopically
▪ Interpret the different grades of hemagglutination reaction
▪ Describe the different types of hemolysis through examination of the cell button and supernatant fluid from the
reaction tube
▪ Differentiate and characterize hemagglutination and hemolysis
▪ Explain the mechanisms and relate the importance of hemagglutination and hemolysis in blood banking
MATERIALS NEEDED:
Compound microscope Pasteur pipette
Test tubes and rack Glass slides
Applicator sticks Centrifuge
REAGENT:
Distilled water
Normal saline solution (0.85 – 0.90% NaCl)
Typing sera (anti – D) undiluted and diluted 1:10 (1 mL anti – D + 9mL NSS)
SAMPLE:
5% red cell suspension (any ABO Rh positive blood)
PROCEDURE:
1. Demonstration of hemagglutination reactions: Slide Method

a. Using a marker, divide one glass slide into two sides.
b. Label one side as U (unknown) and the other as NC (negative control)
c. Place a drop of anti – D in U portion and a drop of NSS in NC portion
d. Place a drop of blood sample in each side of the glass slide.
e. Mix the reagent and blood sample using applicator stick. Use separate applicator sticks for each portion
f. Observe the result macroscopically and microscopically using LPO.
g. Interpret the result within 2 minutes as positive (+) or negative (- or 0). There is no need to grade the reaction
2. Demonstration of hemagglutination reactions: Tube Method

a. Prepare two plain test tubes and label them as U (undiluted) and D (diluted)
b. Place two drops of undiluted anti – D in tube labeled as U.
c. Place two drops of 1:10 diluted anti – D in tube labeled as D.
d. Place one drop of 5% red cell suspension of an Rh (+) blood in each test tube
e. Mix and cover each test tube with Parafilm
f. Centrifuge for 15 seconds at 3,400 rpm
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g. Gently dislodge the cell button and interpret the results

h. Compare the two tubes and take note of the difference
i. Grade the reactions accordingly by comparing the reactions to hemagglutination charts
Guide in the interpretation of Results:

MACROSCOPIC EVALUATION: Red Cell Antigen – Antibody Reactions Serologic Grading
4+ One solid aggregate, clear background
3+ Several large aggregates, clear background
2+ Medium – sized agglutinates, clear background
1+ Small agglutinates, turbid background
W+ Tiny agglutinates, turbid background
0 No agglutination or hemolysis
MICROSCOPIC EVALUATION
Microscopic examination is generally performed:
▪ to differentiate pseudo agglutination (rouleaux) from true agglutination
▪ to detect mixed-field reactions or mf (type of agglutination pattern in which numerous small clumps of cells exist
amid a sea of free cells), and
▪ to confirm a negative reaction.
3. Demonstration of hemolysis
a. Prepare two plain test tubes and label them as P (expected positive) and N (expected negative)
b. Place 2 drops of 5% red cell suspension of an Rh (+) blood in each tube
c. Place 2 drops of distilled water in tube labeled as P
d. Place 2 drops of NSS in tube labeled as N
e. Mix and cover each test tube with Parafilm
f. Centrifuge for 15 seconds at 3,400 rpm. Do not dislodge the cell button.
g. Interpret the results
h. Compare the two tubes and take note of the difference
Interpretation:
▪ No hemolysis: intact cell button with clear supernatant
▪ Partial hemolysis (PH): presence of cell button with pink supernatant
▪ Complete hemolysis (H): absence of cell button with red supernatant
DRAWINGS:
1. Illustrate and label accordingly:
a. Macroscopic Evaluation: Red cell antigen – antibody reactions serologic grading
b. Microscopic Evaluation: True agglutination and Pseudoagglutination
c. Demonstration of hemolysis: No hemolysis, partial hemolysis and complete hemolysis

1. Drawing: Illustrate and label the following:
a. Zeta potential of red blood cells
b. Red cell membrane showing blood group antigens
c. Structure of blood group soluble substances
d. Structure of the common antibodies associated in immunohematologic reactions
2. Describe the zeta potential and charge of the red blood cells
3. Differentiate agglutination from hemolysis
4. Why is it necessary to observe agglutination reactions microscopically?
5. Differentiate and enumerate causes of
a. True agglutination
b. Pseudoagglutination
6. What is rouleaux formation? What causes it? How do we resolve this problem?
7. Describe the mechanisms and clinical significance of hemolysis in immunohematology/blood banking
Note: Review antigen – antibody interactions
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D. ASSESSMENT:
Offline Learners: Answer the given activity and send it to your instructors’ email. Quiz will be given through text/email.
Weak connectivity: Answer the given activity and send it to your instructors’ email. Quiz will be given through email.
Online Learners: Answer the given activity. All activities which include Quizzes, Seat works, Assignments etc. will be
posted in Canvas Instructure.
LECTURE ASSESSMENT:
I. For Items #1-6. Identify who discovered or introduced the following

___________________________1. Syringe-valve apparatus
___________________________2. Sodium citrate
___________________________3. Sodium phosphate
___________________________4. Vein-to-vein transfusion
___________________________5. ACD
___________________________6. CPD
___________________________7. What is the extracellular to intracellular ratio for Na+ and K+ respectively?
___________________________8.
___________________________9. Give 3 additive solutions licensed in the US and write their other names beside it
___________________________10.
___________________________11.
___________________________12. AS-3 contains what component/s which protects the blood from/against storage
related hemolysis?
___________________________13. What is the shelf life of additive solutions?
___________________________14. Transcribe SAGM
___________________________15. The loss of RBC membrane is exemplified by the formation of what cells?
___________________________
II. Explain and briefly describe the changes observed in the RBC storage lesion.
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LABORATORY ASSESSMENT (EXPERIMENT nos. 1-4)
A. Match the Stopper Color in Column A with the corresponding Anticoagulant/Additive in Column B.
Answer Stopper Color Anticoagulant/Additive

1. Red (Glass) A. Sodium citrate
2. Red (Plastic/Hemogard) B. Clot activator separation gel
3. Lavender (glass) C. Sodium fluoride/potassium oxalate
4. Lavender (plastic) D. Clot Activator
5. Pink E. Sodium heparin/Lithium heparin
6. White F. K3EDTA in liquid form
7. Light Blue G. K2EDTA
8. Black H. None
9. Light green/black I. K2EDTA (spray dried)
10. Green J. Thrombin
11. Royal blue K. EDTA and gel
12. Gray L. Sodium heparin, K2EDTA
13. Yellow M. Sodium heparin
14. Tan (glass) N. Acid citrate dextrose
15. Tan (plastic) O. Lithium heparin and gel
16. Yellow/gray and orange P. Sterile containing sodium polyanetholsulfonate
17. Red/gray and gold Q. Thrombin and soybean trypsin inhibitor
B. Order of Draw for Venipuncture

Arrange the following according to the correct Order of Draw. Use letters A-F.
_________18. Heparin tubes with or without gel (green stopper)

_________19. Blood Culture tubes (yellow)
_________20. Coagulation sodium citrate tube (light blue)
_________21. Glycolytic inhibitor tubes (gray stopper)
_________22. Sodium heparin (Tan/glass)
_________23. Ethylenediaminetetraacetic acid tubes (lavender stopper)
C. Enumerate the anticoagulant preservative solutions and write the corresponding shelf-life on the blank
provided
______________24.
______________25.
______________26.
______________27.
D. Enumerate the additive solutions and write the corresponding shelf life on the blank provided
______________28.
______________29.
______________30.
E. REFERENCES
1. Harmening, D. (2012). Modern blood banking and transfusion practices (6th ed.) Philadelphia: F. A. Davis Company
2. American Association of Blood Banks (2011). Technical manual (17th ed.). USA: American Association of Blood
Banks
3. Blaney K.D. & Howard, P.R. (2008). Concepts of immunohematology (2nd ed.). USA: Mosby
4. Harmening, D. (2019). Modern Blood Banking and Transfusion Practices (7 th ed.) F. A. Davis Company
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Name: Rating:
Experiment Number 5
ABO FORWARD GROUPING
Routine testing for determining the ABO blood group of an individual consists of testing the red blood cells with anti – A
and anti – B antisera (cell, antigen, forward or direct type). In this procedure, the medical technologist/laboratory scientist
uses slide and tube methods to detect unknown antigen/s patient red blood cells using commercially-prepared antisera of
known specificity thereby identifying the ABO blood group of the patient.
OBJECTIVES:
▪ Manifests the necessary skills needed ABO forward typing using slide and tube methods
▪ Determine the ABO blood group of an individual by proper discrimination of the presence or absence and grading
reactions of hemagglutination indicating the presence of red cell ABO antigens
▪ Describe and characterize the two methods of ABO forward grouping
MATERIALS NEEDED:
Graduated centrifuge tube Venipuncture set
Blood typing glass slide Applicator stick
Glass slide (ordinary) Pasteur pipette
Test tubes and rack Lancet
REAGENTS:
Anti – A antiserum 22% Bovine serum albumin (BSA)
Anti – B antiserum Normal saline solution (0.85 – 0.90% NaCl)
Anti – AB antiserum
SAMPLES:
Slide Method: blood sample obtained through skin puncture or anticoagulated blood with EDTA obtained through
venipuncture
Tube Method: 2 – 5% red cell suspension of sample
PROCEDURE:
1. Slide Method
a. Prepare 2 blood typing slides or ordinary glass slides
b. Divide each slide into 2 sides by using a marking pen.
c. Label the two sides of the 1st slide as A and B, 2nd slide as AB and NC (negative control).
d. Deliver one drop of anti – A onto the A portion, anti – B onto the B portion, anti – AB onto the AB portion and
22% bovine serum albumin onto the NC portion.
e. Perform skin (capillary) puncture technique to obtain sample. Discard the first drop of blood.
f. Deliver 1 drop of blood to each portion of the slide
g. Mix the reagents and red cells thoroughly, using a clean applicator stick for each reagent. Spread the mixture
over an area approximately 20 mm x 40 mm.
h. Gently rotate or tilt the slide back and forth within 2 minutes. Observe agglutination. Do not read results after
2 minutes.
i. Report as “+” for agglutination and “0” for no agglutination
j. Interpret the results by identifying the antigen/s present and the ABO blood group of the sample
2. Tube Method
a. Prepare 2 – 5% red cell suspension using the unknown blood sample.
b. Label four test tubes as A, B, AB and NC (negative control).
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c. Proceed as follows:
Content Tube A Tube B Tube AB NC
Anti – A 1 drop - - -
Anti – B - 1 drop - -
Anti – AB - - 1 drop -
22% BSA - - - 1 drop
d. Deliver exactly 1 drop of well mixed 2 – 5% unknown red cell suspension in all tubes.
e. Cover each tube with parafilm and mix contents by gentle inversion.
f. Centrifuge all tubes for 15 seconds at 3,400 rpm.
g. Gently dislodge the cell button and examine for agglutination or hemolysis.
Note: It is important to dislodge the tubes gently because vigorous dislodging may cause agglutinated cells to
break and result might be interpreted as false negative
h. Grade each reaction and record results.
i. Interpret the results by identifying the antigen/s present and the ABO blood group of the sample
Guide in the interpretation of Results:

ABO Blood Group:
Anti – A Anti – a Anti – AB Negative Control
Cell (Antigen)
A + 0 + 0
B 0 + + 0
AB + + + 0
O 0 0 0 0
+ = with agglutination; 0 = no agglutination
Type A: indicates that the RBC of the patient has A antigen only
Type B: indicates that the RBC of the patient has B antigen only
Type AB: indicates that the RBC of the patient has both A and B antigens
Type O: indicates that the RBC of the patient has neither A nor B antigens
DRAWINGS:
1. Illustrate and label accordingly the procedure and standard results for ABO forward grouping slide method
2. Illustrate and label accordingly the procedure and standard results for ABO forward grouping tube method

1. What is antiserum?
2. What are the potency requirements in an antiserum?
3. Describe the formation of antigens of the ABO blood group
4. Describe the importance of doing forward grouping
5. What kind of antigen will anti-A detect? anti-B?
6. Describe the importance of determining the ABO subgroups
7. Differentiate slide method from tube method in ABO typing (include their advantages and disadvantages over the
other)
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Name: Rating:
Experiment Number 6
ABO REVERSE GROUPING
Aside from testing the red blood cell for antigens, testing the serum or plasma with A and B red cell suspensions (serum,
antibody, backward, indirect or reverse type) can also be performed to determine the ABO blood type. In this procedure,
the medical technologist/laboratory scientist detects the unknown antibodies present in the patient’s serum by using red
cell suspensions of known antigenic specificity. Both red cell and serum testing are required for routine ABO tests on
donors and patients because each serve as a check on the other.
OBJECTIVES:
▪ Manifests the necessary skills needed for ABO reverse typing
▪ Determine the ABO blood group of an individual by proper discrimination of the presence or absence and grading
reactions of hemagglutination indicating the presence of plasma ABO antibodies.
MATERIALS NEEDED:
Graduated centrifuge tube
Test tubes and rack
Venipuncture set
Pasteur pipette
REAGENTS:
2 – 5% of known A, B, AB and O red cell suspensions
SAMPLE:
Serum (should be free from contamination, hemolysis and lipemia)
PROCEDURE:
1. Prepare 2 – 5% of red cell suspension of A, B, AB and O cells. Label the centrifuge tubes of the reagent RCS
accordingly.
2. Label 4 tubes as A, B, AB, and O respectively.
3. Add serum sample and reagent to the tubes according to the table below:
Content Tube A Tube B Tube AB Tube O
Unknown Serum 2 drops 2 drops 2 drops 2 drops
A – cell 1 drop - - -
B – cell - 1 drop - -
AB – cell - - 1 drop -
O – cell - - - 1 drop
4. Cover the tubes with Parafilm.
5. Mix gently and centrifuge for 15 seconds at 3,400 rpm.
6. Gently dislodge the cell button and examine for hemolysis or agglutination.
7. Grade reactions and record results.
49 | P a g e
Guide in the Interpretation of Results:

ABO Blood Group: Known AB –
Known A – cells Known B – cells Known O – cells
Serum (Antibody) cells
A 0 + + 0
B + 0 + 0
AB 0 0 0 0
O + + + 0
Type A: presence of anti – B that shows agglutination in B and AB cells

Type B: presence of anti – A that shows agglutination in A and AB cells
Type AB: absence of both anti – A and anti – B that shows no agglutination in A, B and AB cells
Type O: presence of anti – A and anti – B that shows agglutination in A, B and AB cells
DRAWING:
Illustrate and label accordingly the procedure and standard results for ABO reverse grouping

1. Describe the importance of doing serum typing
2. Can plasma be used in reverse grouping? Justify. What is/are its disadvantage/s?
3. Describe the type and formation of the antibody/ies found in (a) group “A”, (b) group “B”, (c) group “AB”, and (d)
group “O”
4. Enumerate causes of false-positive and false-negative ABO typing results and give corresponding corrective
actions
50 | P a g e
Name: Rating:
Experiment Number 7
SECRETOR STATUS DETERMINATION USING SALIVA
Certain blood group substances occur in soluble form in secretions such as saliva in a large proportion of individuals
termed as “secretors”. These water-soluble blood group substances are readily detected in very minute quantities
because they have the property of reacting with their corresponding antibodies and thereby neutralizing or inhibiting the
capacity of the antibody to agglutinate erythrocytes possessing the corresponding antigen. The reaction is termed
hemagglutination inhibition and provides a means of measuring the relative activity or strength of these water-soluble
group substances in secretions.
OBJECTIVES:
▪ Manifest the necessary skills needed for testing the secretor status of an individual
▪ Determine the secretor status of an individual through saliva testing
▪ Describe the principle of hemagglutination inhibition for determination of secretor status
MATERIALS NEEDED:
Test tubes and rack
Centrifuge tube
Pasteur pipette
Beaker
REAGENTS:
5% red cell suspension of known A,B and O cells
Anti – A serum (1:4 dilution) 1 mL anti – A + 3 mL NSS
Anti – B serum (1:4 dilution) 1 mL anti – B + 3 mL NSS
Anti – H lectin (1:4 dilution) 1 mL anti – H +3 mL NSS
SAMPLE:
Saliva
PROCEDURE:
1. Collection and Preparation of Saliva

a. Collect approximately 5 to 10 mL of saliva in a small beaker or wide-mouthed test tube. Most people can
accumulate this amount in several minutes. To simulate salivation, the subject may be asked to chew wax,
paraffin, or a clean rubber band, but not gum or anything else that contains sugar or protein.
b. Centrifuge saliva at 3,400 rpm for 10 minutes.
c. Transfer supernatant to a clean test tube, and place stoppered tube in a boiling water bath for 10 minutes. This
inactivates enzymes that might otherwise destroy blood group substances.
d. Recentrifuge at 3,400 rpm for 10 minutes.
e. Transfer the clear or slightly opalescent supernatant into a clean test tube. Discard any opaque or semi-solid
material. If the supernatant is not yet clear, it suggests incomplete heating and must be reheated for another 5
minutes in the boiling water bath.
f. Make a 1:2 dilution of the collected supernatant with NSS (1 mL clear supernatant + 1 mL NSS)
2. Test for Secretor Status

a. Prepare 1:4 dilution of anti – A, anti – B and anti – H antisera
51 | P a g e
b. Prepare 5% red cell suspension of A, B and O cells

c. Prepare 3 test tubes and label as A, B and H
d. Add sample and reagent to the tubes according to the following table:
CONTENT Tube A Tube B Tube H
Diluted saliva 1 drop 1 drop 1 drop
Diluted anti – A 1 drop - -
Diluted anti – B - 1 drop -
Diluted anti – H - - 1 drop
e. Mix and cover Parafilm. Incubate all tubes at room temperature for 10 minutes.
f. Add 5% known red cell suspension according to the following table:
CONTENT Tube A Tube B Tube H
5% known A – cells 1 drop - -
5% known B – cells - 1 drop -
5% known O – cells - - 1 drop
g. Mix and over with Parafilm. Incubate at room temperature for 30 to 60 minutes or for 15 minutes in a 37°C water
bath.
h. Centrifuge tubes for 15 seconds at 3,400 rpm.
i. Gently dislodge the cell button and examine for agglutination.
j. Grade each reaction and record the results
Guide in the Interpretation of Result:

Tube A Tube B Tube H
Blood Group Soluble Substance
Saliva + Anti – A Saliva + Anti – B Saliva + Anti – H
(Secretor) Detected
+ A cells + B cells + O cells
A 0 + 0 A, H
B + 0 0 B, H
AB 0 0 0 A, B, H
O + + 0 H
Nonsecretor
+ + + None
(for ABH antigens)
DRAWING:
Illustrate and label accordingly the procedure and standard results for the determination of secretor status using saliva

1. Describe the clinical significance of determining the secretor status of a patient
2. Why must saliva be subjected into increased temperature using a water bath prior to its use?
3. Describe the principle involved in this test
4. What is the purpose of diluting both the saliva and the antisera in this test?
5. Why is saliva preferred as specimen for determination of blood group soluble substances?
6. Enumerate other sources of the blood group soluble substances
52 | P a g e
Name: Rating:
Experiment Number 8
SCREENING TEST FOR LOW TITER GROUP “O” BLOOD
Type O individuals are considered to be the universal red blood cell donors due to the absence of A and B antigens on
their red blood cells. If their blood is given to any non-O and non-Bombay recipients, no major transfusion reaction is
expected to happen. This is with the assumption that they also belong to the same Rh blood group and no other atypical
antibodies are found in the serum of both the recipient and the donor. However, the serum/plasma of type O individuals
contains naturally occurring anti-A and anti-B that can react to patient’s red cells having the corresponding antigens. If
non-O recipients transfused with type O whole blood, incompatibilities can be detected in the minor crossmatch if the titer
of the antibodies is high enough to cause a reaction. In this case, only packed red cells should be given to the recipient to
avoid reactions brought about by the antibodies coming from the donor. Type O donor is considered only as a universal
donor if the antibody titer is less than 1:50.
OBJECTIVES:
▪ Display performance of the screening test for type O universal donors
▪ Describe the concept of group O universality in transfusion
MATERIALS NEEDED:
Beaker or Erlenmeyer flask Venipuncture set
Test tubes and rack Centrifuge tube
Serological pipette Pasteur pipette
REAGENTS:
2 – 5% A red cell suspension (freshly prepared known A – cells)
2 – 5% B red cell suspension (freshly prepared known B – cells)
SAMPLE:
Group O serum/plasma (1:50 dilution)
PROCEDURE:
1. Prepare 1:50 dilution of group “O” serum/plasma in an Erlenmeyer flask or beaker
(0.1 mL of serum/plasma + 4.9 mL of NSS)
2. Label 4 test tubes. Mark 2 tubes as “UA” and “UB” for the set of undiluted samples. Mark the other 2 tubes as “DA”
and “DB” for the set of undiluted samples.
3. Proceed as follows:
CONTENT UA UB DA DB
Undiluted
0.1 mL 0.1 mL -- --
serum/plasma
Diluted
-- -- 0.1 mL 0.1 mL
serum/plasma
2-5% known A cells 0.1 mL -- 0.1 mL --
2-5% known B cells -- 0.1 mL -- 0.1 mL
Undiluted serum/ Undiluted serum/ Diluted serum/ Diluted serum/ plasma with B
Legend
plasma with A cells plasma with B cells plasma with A cells cells
4. Mix the contents of the tubes and cover all tubes with Parafilm
5. Centrifuge for 15 seconds at 3,400 rpm
53 | P a g e
6. Gently dislodge the cell button and examine for agglutination or hemolysis
7. Interpret and grade the agglutination reaction
Guide in the Interpretation of Result in Screening Low Titer O:

REACTION TUBES
INTERPRETATION
UA UB DA DB
Low Titer O: Antibody titer is less than 1:50.
Concentration of anti-A and anti-B is sufficiently low to cause reaction.
+ + 0 0
✓ Whole blood can be safely transfused irrespective of the ABO blood type
of the recipient
High Titer O: Antibody titer is more than 1:50.
Concentration of anti-A and anti-B is sufficiently high to cause reaction.
+ + + +
✓ Packed RBC can be safely transfused irrespective of the ABO blood
type of the recipient
Inconclusive: Absence of agglutination reaction suggests doubtful results
Further investigation is suggested
0 0 0 0
✓ This includes taking note of the age and medical history of the type O
individual or considering technical factors
DRAWING:
Illustrate and label accordingly the important steps and standard results of screening test for low titer group “O” blood

1. What is the importance of determining the antibody titer of type “O” blood?
2. Why is the test done among type “O” blood?
3. Enumerate the reasons why type O serum does not result to a positive reaction if mixed with A and B cells?
E. ASSESSMENT:
Offline learners: Answer the given activity and send it to your instructors’ email. Quiz will be given through text/email.
Online Learners: Answer the given activity. All activities which include Quizzes, Seat works, Assignments etc. will be
posted in Canvas Instructure.
LECTURE ASSESSMENT:
1. Enumerate and briefly explain the Landsteiner Laws and give example of each.
2. What should be the first thing that a medical technologist can do when there is an ABO discrepancy?
54 | P a g e
LABORATORY ASSESSMENT (EXPERIMENT nos. 5-8)
______________________1. This term refers to an individual with regards to secretion of A, B, and H soluble antigens in
body fluids
______________________2. What is the corresponding result when the patient is a secretor?
______________________3. What is the corresponding result when the patient is a non-secretor?
______________________4. Demonstration of A, B, and H substances in saliva is evidence for the
______________________5. Inheritance of these genes.
______________________6.
______________________7.
______________________8. True or False. Secretor studies may be helpful in identifying a subgroup of A or B antigens.
______________________9. Enumerate at least 3 fluids in which A,B and H substances can be
______________________10. detected in secretors except for saliva.
______________________11.
For Items # 12-15. Identify the ABH substances present in the saliva of secretors given the following blood types.
_____________12. Blood type A

_____________13. Blood type AB
_____________14. Blood type B
_____________15. Blood type O
For Items 16-20. (2 points each). Given the genes inherited. Write down the equation and result of the secretor
status determination process.
Follow this template: Saliva + antisera + RCS= agglutination or no agglutination= Secretor or nonsecretor
16. AO Hh Sese
17. AB hh SeSe
18. OO Hh sese
19. BO HH Sese
20. B hh sese
F. REFERENCES
Banks
4. Harmening, D. (2019). Modern Blood Banking and Transfusion Practices (7th ed.) F. A. Davis Company
5. Hubbard, J. D. (2010). A concise review of Clinical Laboratory Science. (2nd ed.). Philadelphia, PA: Lippincott
Williams & Wilkins.
55 | P a g e
Name: Rating:
Experiment Number 9
Rh GROUPING
The terms “Rh positive” and “Rh negative” refer to the presence or absence of the red cell antigen D. after the A and B
antigens, D is the most important red cell antigen in transfusion practice for it has greater immunogenicity than other red
cell antigens. In contrast to A and B, however, persons whose red cells lack the D antigen do not regularly have anti-D.
formation of anti-D results from exposure, through transfusion or pregnancy, to red cells possessing the D antigen.
In clinical practice, five blood typing reagents are readily available: anti-D, -C, -E, -c, and –e. routine pretransfusion
studies include only tests for D. other reagents are used principally in the resolution of antibody problems or in family
studies. The assortment of antigens detected on a person’s red cells constitutes that person’s Rh phenotype.
OBJECTIVES:
▪ Manifest the necessary skills needed for identifying the blood group
▪ Determine the Rh blood group of an individual
▪ Describe and characterize the methods used for Rh grouping
MATERIALS NEEDED:
Rh viewbox (surface temperature 45 - 50°C)
Test tubes and rack
Venipuncture set
Evacuated tubes
Pasteur pipette
Glass slides
REAGENTS:
Anti – D antiserum (also anti –C, anti – c, anti – E, and anti – e antisera if available)
22% Bovine serum albumin (BSA)
Rh/Hr control cells (if available)
SAMPLES:
Slide Method: blood sample obtained through skin puncture
Tube Method: 2 – 5% red cell suspension of sample
PROCEDURE:
1. Slide Method
a. Prepare 2 glass slides.
b. Divide each slide into 3 portions by using a marking pen.
c. Label 3 portions of the first slide as D, C, and c. label 3 portions of the second slide as E, e, and CTRL.
d. Place the glass slides on Rh viewbox (surface temperature: 45 – 50°C) for 5 minutes.
e. Remove the pre-warmed slides and deliver 1 drop of anti – D, anti – C, anti – c, anti – E, anti – e, and Rh/Hr
control on the approximately marked position of the glass slide.
f. Perform skin puncture by prick method and add 1 drop of whole blood to each portion.
g. Mix well with separate applicator sticks.
h. Tilt the slides back and forth and observe for agglutination within 2 minutes.
66 | P a g e
Note: Reading of the results should be completed within 2 minutes; otherwise, drying could be falsely interpreted
as a positive reaction. Results that show no agglutination within 2 minutes are considered to be negative and
therefore suggest absence of that specific antigen.
i. Interpret and report results according to the antigen/s detected and the Rh blood group

✓ Agglutination with anti-D and a smooth suspension on the control slide constitute a positive test result and
indicate that the cells being tested are D (+) → Rh positive
✓ No agglutination with either anti-D or the Rh control suggests that the cells are D (-) → perform tube method
and/or antiglobulin test
Note: Testing by the antiglobulin procedure will show weak expression of D on cells that are not agglutinated on
slide testing
2. Tube Method
a. Prepare a 2 – 5% red cell suspension of the unknown blood sample.
b. Label 2 tubes as U (unknown) and NC (negative control) respectively.
c. Add the sample and reagents to the labeled tubes by following the table below:
Content Unknown Negative Control
Anti – D 1 drop -
22% Bovine Serum Albumin - 1 drop
2 – 5% Red Cell Suspension 1 drop 1 drop
d. Mix gently. Cover each tube with Parafilm.

e. Centrifuge all tubes for 15 seconds at 3,400 rpm.
f. Gently dislodge each cell button and examine for agglutination.

✓ Agglutination in the anti-D tube, combined with a smooth suspension in the control tube, indicates that the red
cells under investigation are D (+) → Rh positive
✓ A smooth suspension of red cells in both the anti-D and the control tubes is a negative test result.
✓ Donor blood must be further tested for the presence of weak D antigen → perform antihuman globulin test
(indirect) to differentiate weak D expression from Rh negative phenotype
DRAWING:
1. Illustrate and label accordingly the procedure and standard results for Rh grouping slide method
2. Illustrate and label accordingly the procedure and standard results for Rh grouping tube method

1. Why is it necessary to pre-warm the slide prior to its use?
2. How is Rh typing serum prepared?
3. What is the importance of control in Rh typing?
4. What is meant by the terms (a) Rh positive and (b) Rh negative?
5. Enumerate causes of false-positive and false-negative Rh typing results and give corresponding corrective
actions
6. In cases of hemolytic diseases of the fetus and the newborn, what is the effect of primary administration of Rh
antibodies in response to the offending antigen?
67 | P a g e
Name: Rating:
Experiment Number 10
TEST FOR WEAK D ANTIGEN (Du variant)
In Rh phenotyping, the absence of agglutination reaction in slide and tube methods is not immediately reported as Rh
negative. Some red blood cells express the D antigen so weakly that most anti-D reagents do not directly agglutinate
them. Thus, test for weak expression of the D antigen must be performed before reporting Rh negative results. Weak D
expression, which is reported as Rh positive, can be recognized most reliably by an indirect antihuman globulin test (IAT),
discriminating it from Rh negative phenotype.
OBJECTIVES:
▪ Manifests the skills necessary for the performance of an indirect antiglobulin test for weak D testing
▪ Determine the presence or absence of weak D antigen
▪ Describe and discriminate Rh positive from Rh negative phenotype
MATERIALS NEEDED:
Graduated centrifuge tube Evacuated tubes
Test tubes and rack Pasteur pipette
Venipuncture set
REAGENTS:
Anti – D antiserum
Antihuman globulin antiserum
SAMPLE:
2 – 5% red cell suspension of sample
PROCEDURE:
1. Prepare a 2 – 5% red cell suspension of the unknown blood sample.
2. Label 2 tubes as U (unknown) and NC (negative) respectively.
3. Add sample and reagents according to the table below:
Content Unknown Negative Control
Anti – D 1 drop -
22% Bovine Serum Albumin - 1 drop
2 – 5% Red Cell Suspension 1 drop 1 drop
4. Mix gently. Cover each tube with parafilm. Incubate both tubes for 15 minutes at 37°C water bath. Centrifuge all tubes
for 15 seconds. Gently dislodge each cell button and examine for agglutination.
Note: Unknown tube with agglutination is regarded as Rh positive. Unknown tube without agglutination goes to the
next procedure.
5. Using the unknown tube without agglutination, wash the cells (steps a → b → c below) for 3 times using normal saline
solution
a. Decant or discard the saline completely
b. Add equal volume of saline
c. Centrifuge cells at 15 seconds for 3,400 rpm
6. Decant or discard the saline completely after the final washing.
7. Add 2 drops of antihuman globulin reagent to all tubes and mix gently. Cover with parafilm.
8. Centrifuge for 15 seconds at 3,400 rpm.
68 | P a g e
9. Gently dislodge each cell button and examine for agglutination.

10. Interpret and record results.
Note: Absence of agglutination confirms the blood group to be Rh negative. Presence of agglutination means
presence of weakly reacting D.
11. For each negative tube, add 1 drop of well mixed check cells.
Note: a positive result is expected after the addition of check cells. This implies that the test has been properly
performed. Negative results with these cells indicate an improperly performed test and the test should be repeated.

Anti – D
Manner of Reporting Additional Comment
(Slide or Tube Method)
Positive reaction indicates presence
+ Rh Positive
of D antigen
Negative reaction should be further
tested for the presence of weak D
0 Rh Negative (initial)
with the use of antihuman globulin
test
Indirect Antihuman Globulin Test for Weak D Antigen Expression Interpretation of Results:
Anti – D
Manner of Reporting Additional Comment
(Slide or Tube Method)
Rh (-) Du (+) should be given
proper classification:
+ Du positive
a. If patient: Rh negative
b. If donor: Rh positive
Rh (-) Du (-): Negative reaction
u
0 D negative Confirms that the person is Rh
negative (patient or donor)
Note:
✓ If the reaction is positive for the test of Du, the person has to be given proper designation because the person
might have weakened D reaction due to missing part in the D antigen (D mosaic). Thus, if given accidentally to an
ABO compatible Rh-positive blood with complete D antigen, the person might develop an antibody against that
missing part which should be avoided.
✓ Remember: Du positive individuals are classified as Rh positive when they donate blood and Rh negative when
they receive blood.
DRAWING:
Illustrate and label accordingly the procedure and standard results for weak D testing using indirect antihuman
globulin test.

1. What is Du?
2. Why do we test for Du when weak or no reaction is obtained in Rh typing?
3. What is required to demonstrate the presence of cells carrying a weak D antigen?
4. Explain the mechanism/s behind the existence of Du
5. Describe the importance of testing weak D in blood typing, compatibility testing and transfusion
69 | P a g e
LABORATORY ASSESSMENT (EXPERIMENT no. 8 and 9)
I. Identification
____________________________1. What is the surface temperature of the Rh viewbox.
____________________________2. Time needed to prewarm the slides.
____________________________3. Time needed for the completion of reading results.
____________________________4. What test is performed to differentiate weak D expression from Rh negative
phenotype?
____________________________5. Incubation requirements for the weak D testing (tube method).
____________________________6. Negative results with these cells indicate an improperly performed test and the test
should be repeated.
For items 7-10. Identify the Rh type given the following results:
____________________________7. Rh(-) Du (+) Patient.
____________________________8. Rh (-)Du (-) Donor.
____________________________9. Rh(-) Du (+) Donor.
____________________________10. Rh (-)Du (-) Patient.
____________________________11. ISBT number of Rh c antigen.
____________________________12. Performed to confirm whether the father possess one or two copies of the RHD
gene.
____________________________13. Codes for the presence or absence of the RhD protein.
____________________________14. The coexpressor gene resides on what chromosome.
____________________________15. It has over 110,000-202,000 antigen sites.
____________________________16. It possess the largest number of D antigen sites among the commonly
encountered Rh genotypes.
____________________________17. What is the category of the weak D expression characterized by having D
appearing to be complete but fewer in number?
____________________________18. Part of an antigen molecule to which an antibody attaches itself.
____________________________19. Category of weak D occurring in individuals whose RBCs possess an extremely
low number of D antigen sites.
____________________________20. Immunogenicity of common Rh antigens from greatest to least.
F. REFERENCES
Banks
4. Harmening, D. (2019). Modern Blood Banking and Transfusion Practices (7 th ed.) F. A. Davis Company
5. Hubbard, J. D. (2010). A concise review of Clinical Laboratory Science. (2nd ed.). Philadelphia, PA: Lippincott
Williams & Wilkins.
71 | P a g e
Name: Rating:
DIRECT ANTIHUMAN GLOBULIN TEST
Coombs, Mourant and Race describe procedures for detecting attachment of antibodies that did not produce
agglutination. This test uses antibody to human globulins and is known as the antihuman globulin (AHG) test, antiglobulin
test (AGT), or Coombs test. In immunohematology, antiglobulin testing generates visible agglutination of sensitized red
cells.
The direct antiglobulin test (DAT) is used to demonstrate in-vivo sensitization or coating of red blood cells with antibody or
complement components, in particular IgG and C3d.
OBJECTIVES:
▪ Manifests the skills necessary for the performance of direct antiglobulin test
▪ Interpret and describe the results of direct antiglobulin test
▪ Describe clinically significant results and conditions identified by direct antiglobulin test
MATERIALS NEEDED:
Evacuated tube (EDTA)
Test tubes and rack
Venipuncture set
Pasteur pipette
REAGENT:
Coombs control cells (to be prepared)
SAMPLE:
2 – 5% red cell suspension of EDTA anticoagulated sample
PROCEDURE:
1. Preparation of Coombs Control Cells
a. Dilute Anti-D (IgG) / Anti D (polyclonal) reagent 1:50 in isotonic saline
b. Prepare a 5% suspension of group “O” Rho (D) positive cells in isotonic saline
c. Mix equal volumes of diluted Anti-D reagent and 5% suspension of “O” Rho (D) positive cells and incubate at 37°C
for 15 minutes.
d. Decant and wash thoroughly with isotonic saline at least thrice.
e. Resuspend in isotonic saline to make a 5% suspension of Coombs control cells.
2. Performance of Direct Antiglobulin Test
a. Prepare 2 tubes, one labeled as U (unknown) and the other as C (control)
b. Dispense 2 drops of a 2% to 5% suspension of red cells into each test tube
c. Wash each tube (steps 1→ 2→ 3 below) for three or four times with saline
(1) Add equal volume of saline
(2) Centrifuge cells at 15 seconds for 3,400 rpm
(3) Decant or discard the saline
d. Completely decant the final wash
92 | P a g e
e. Proceed as follows:
CONTENT U (unknown) C (control)
Antihuman globulin serum
2 drops -
(add immediately)
Normal saline solution - 2 drops
f. Mix and centrifuge for 15 seconds at 3,400 rpm
g. Gently dislodge the cell button and examine for hemolysis or agglutination
h. Grade each reaction and record the results
Note: if using polyspecific AHG or anti-C3d, incubate nonreactive tests at room temperature for 5 minutes, then
centrifuge, and read again.
i. Confirm the validity of negative tests by adding 1 drop of IgG-coated red cells (Coombs control cells) to tests
containing anti-IgG. Centrifuge at 15 seconds at 3,400 rpm. Examine the cells for agglutination and record the
reaction.
Note: Coombs check cells are also added to confirm if AHG was previously added. If AHG was not added, no
agglutination will be observed after the addition of check cells. It is suggested that the procedure should be
repeated.

✓ Presence of agglutination indicates positive test result and absence indicates negative test result.
Control must show absence of agglutination
Result Manner of Reporting Interpretation
Red blood cells were sensitized in
Presence of agglutination DAT Positive
vivo
Red blood cells were not sensitized
No agglutination DAT Negative
in vivo
INVESTIGATION OF POSITIVE DIRECT ANTIGLOBULIN TEST:

1. Elution
▪ A process whereby cells that are coated with antibody are treated in such a manner as to disrupt the bonds
between the antigen and antibody
▪ The objective of ail elution techniques is to interfere with the noncovalent binding forces that hold antibody-
antigen complexes together on the red cell surface. The cell membrane can be physically disrupted by heat,
ultrasound, freezing and thawing, detergents, or other solvents. The binding forces of antigen-antibody complexes
can be interrupted by alterations in pH or salt concentration.
2. Adsorption
▪ Providing an antibody with its corresponding antigen under optimal conditions so that the antibody will attach to the
antigen, thereby removing the antibody from the serum
✓ Dissociation of antibodies from red cells enables the identification of autoantibodies or alloantibodies.
✓ Elution methods used in conjunction with adsorption techniques are also useful in detecting weak antigen
expression on the adsorping cells and in separating mixtures of antibodies against red cell antigens.
DRAWING:
Illustrate and label accordingly the procedure and standard results for direct antiglobulin test

1. What is a Coombs’ reagent? How is it prepared?
2. Describe the principle of direct antihuman globulin test (DAT)
3. Describe and discuss the significance of DAT in the following:
a. Hemolytic transfusion reactions (HTR)
b. Hemolytic disease of the fetus and the newborn (HDFN)
c. Warm autoimmune hemolytic anemia (WAIHA)
d. Paroxysmal called hemoglobinuria (PCH)
e. Cold agglutinin disease (CAD)
f. Graft-versus-host disease (GVHD)
g. Transfusion-associated graft-versus-host disease (TAGVHD)
4. Describe the purpose of (a) elution and (b) adsorption procedures in relation to DAT
5. Enumerate and briefly describe methods based on (a) elution and (b) adsorption
93 | P a g e
Name: Rating:
INDIRECT ANTIHUMAN GLOBULIN TEST
If direct antiglobulin tests is used to detect in-vivo red blood cell sensitization by antibodies and complement components,
indirect antiglobulin test (IAT) on the other hand is used to demonstrate in-vitro reactions between red blood cells and
unexpected antibodies that sensitize, but do not agglutinate, cells that express the corresponding antigen.
In indirect antiglobulin procedures, serum (or plasma) is incubated with red cells, which are then washed to remove
unbound globulins. Agglutination that occurs when antihuman globulin (AHG) serum is added indicates that antibody has
bound to a specific antigen present on the red cells. This procedure is used to determine whether any sort of antigen-
antibody interaction has occurred in-vitro. It uses type O red blood cells suspension because it primarily detects
antibodies other than the naturally occurring anti-A and anti-B, IAT is useful in red blood cell phenotyping, detecting weak
D antigen expression, antibody screening and identification, and crossmatching.
OBJECTIVES:
▪ Manifests the skills necessary for the performance of indirect antiglobulin test
▪ Interpret and describe the results of indirect antiglobulin test
▪ Describe clinically significant results and conditions identified by indirect antiglobulin test
▪ Discriminate the principles and applications of direct and indirect antiglobulin test
MATERIALS NEEDED:
Graduated centrifuge tube Venipuncture set
Evacuated tube (EDTA) Pasteur pipette
Test tubes and rack
REAGENTS:
Coombs control cells (to be prepared)
SAMPLES:
Serum (should be free from contamination, hemolysis and lipemia)
2 – 5% “O” red cell suspension
PROCEDURE:
1. Prepare two tubes. Label tubes as U (unknown) and NC (negative control) respectively
2. Proceed as follows:
Contents Unknown Negative Control
Unknown serum 2 drops -
2 – 5% “O” red cell suspension 2 drops 2 drops
22% Bovine Serum Albumin - 2 drops
3. Mix gently and cover with Parafilm. Incubate at 37°C water bath for 15 minutes.
4. At the end of incubation, centrifuge for 15 seconds at 3,400 rpm. Examine for agglutination.
5. If no agglutination is observed, wash the unknown tube (steps a → b → c below) for three times with normal saline
solution
a. Add equal volume of saline
b. Centrifuge cells at 15 seconds for 3,400 rpm
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c. Decant or discard the saline

6. Decant completely after the last washing.
7. Add 2 drops of antihuman globulin serum and mix well.
8. Centrifuge for 15 seconds at 3,400 rpm.
9. Gently dislodge cell button and observe for agglutination.
10. If the test does not show agglutination, add 1 drop of Coombs check cells to the unknown tube and centrifuge for 15
seconds at 3,400 rpm.
11. Gently dislodge the cell button. Observe for agglutination or hemolysis.
12. Grade each reaction and record the results.
Guide in Interpretation of Results:

Result Manner of Reporting Interpretation
Presence of atypical antibodies in
With agglutination or hemolysis IAT positive the patient’s serum coating red
blood cells in vitro
Absence of atypical antibodies in
No agglutination or hemolysis IAT negative the patient’s serum coating red
blood cells in vitro
DRAWING:
Illustrate and label accordingly the procedure and standard results for indirect antiglobulin test

1. Describe the principle of indirect antihuman globulin test
2. Describe the uses of indirect antihuman globulin test
3. Enumerate the sources of error in antiglobulin testing and the corresponding corrective action
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LABORATORY ASSESSMENT (EXPERIMENT nos. 11 and 12)
A. Matching Type: Match the following task to its specific purpose

TASKS PURPOSE
______1. Centrifuge a. Interprets test as positive or negative
______2. Examine for Agglutination b. Accelerates agglutination
______3. Perform a minimum of 3 saline washes c. Determines the strength of reaction
______4. Grade agglutination reactions d. Checks for neutralization of antisera by free globulin
molecules
______5. Add antiglobulin reagent e. Allows time for antibody molecule attachment to RBC
______6. Add antibody coated RBCs to negative f. Removes free globulin molecules
reactions
______7. Incubate RBCs with antisera g. Forms RBC agglutinates (RBC Ag+ Ab+ anti-IgG)
B. Headings: A. False-Positive Result

B. False-Negative Result
_________8. Polyagglutinable cells

_________9. Cell suspension either too weak or too heavy
_________10. Low pH of saline
_________11. Improper specimen (refrigerated, clotted)
_________12. Autoagglutinable cells
_________13. Saline contaminated by heavy metals or colloidal silica
_________14. Preservative-dependent antibody in LISS reagents (IAT)
_________15. Inadequate or improper washing of cells
C. Enumeration. Enumerate the different modified and automated antiglobulin test techniques employed in blood
banking.
16.
17.
18.
19.
20.
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Name: Rating:
COMPATIBILITY TESTING: CROSSMATCH
Compatibility testing is a series of procedures designed to ensure the safety of blood transfusion. Included in the
compatibility test is crossmatching which is performed prior to transfusion of blood components containing red blood cells.
It is the testing of the patient’s serum with the donor red blood cells, including an antiglobulin phase or simply an
immediate spin phase to confirm ABO compatibility. A crossmatch procedure is divided into major crossmatch, consisting
of mixing recipient (patient) serum with donor red cells and minor crossmatch, consisting of mixing donor serum with
recipient (patient) red cells.
OBJECTIVES:
▪ Display performance of the different methods of major and minor crossmatch accurately
▪ Describe the significance and phases of broad-spectrum crossmatch
▪ Interpret the results of a major and minor crossmatch
▪ Differentiate compatible from incompatible recipient and donor
MATERIALS NEEDED:
Test tubes and rack Applicator stick
Venipuncture set Glass slides
Centrifuge tube Water bath
Pasteur pipettes Centrifuge
REAGENTS:
Low ionic strength solution (LISS)
Antihuman globulin (AHG) sera
Check cells (to be prepared)
SAMPLES:
Donor serum (should be free from contamination, hemolysis and lipemia)
Recipient (patient) serum (should be free from contamination, hemolysis and lipemia)
Slide (Emergency) Crossmatch: anticoagulated (whole) blood
Broad Spectrum Crossmatch:
2 – 5% suspension of recipient (patient) red cells
2 – 5% suspension of donor red cells
PROCEDURE:
Slide (Emergency) Crossmatch

1. Prepare a clean, dry slide and divide it into 2 portions using a marker
2. Mark the one portion as MAJOR (for major crossmatch) and the other as MINOR (for minor crossmatch)
3. To MAJOR glass slide portion, add 1 drop of recipient (patient) serum using a Pasteur pipette. To MINOR glass slide
portion, add 1 drop of donor serum using another Pasteur pipette.
4. Add 1 drop of donor red blood cells to MAJOR glass slide portion 1 drop of recipient (patient) red blood cells to
MINOR glass slide portion using separate Pasteur pipettes.
5. Mix each portion using separate applicator sticks.
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6. Gently rotate or tilt the slide back and forth to mix the samples then incubate at room temperature for 3 minutes
covered with a petri dish cover.
7. After incubation, gently rotate or tilt the slide back and forth. Observe for agglutination macroscopically and/or
microscopically (to confirm agglutination). Report major and minor crossmatch results.
Guide in the Interpretation of Result: With agglutination = INCOMPATIBLE; No agglutination = COMPATIBLE
Broad Spectrum Crossmatch:
1. Immediate Saline Spin Phase:

a. Label 4 test tubes: Ma – S, Ma – P, Mi – S, Mi – P
b. Proceed as follows:
TUBE
CONTENT
Ma – S Ma – P Mi – S Mi – P
Patient’s serum 2 drops 2 drops -- --
Donor’s serum -- -- 2 drops 2 drops
2 – 5% suspension of donor’s cells 1 drop 1 drop -- --
2 – 5% suspension of patient’s cells -- -- 1 drop 1 drop
22% Bovine Serum Albumin (BSA) -- 2 drops -- 2 drops
Legend:
▪ Ma – S: Major crossmatch – Saline [Recipient/patient’s serum + Donor’s red cell]
▪ Ma – P: Major crossmatch – Protein [recipient/patient’s serum + Donor’s red cell + Albumin/protein]
▪ Mi – S: Minor crossmatch – Saline [Recipient/patient’s red cell + donor’s serum]
▪ Mi – P: Minor crossmatch – Protein [Donor’s serum + Recipient/patient’s red cell +Albumin/protein]
c. Mix the content and cover with Parafilm.
d. Centrifuge for 15 seconds at 3,400 rpm.
e. Gently dislodge the cell button and observe for agglutination or hemolysis.
f. If no agglutination or hemolysis is observed, only Ma – P and Mi – P will proceed to the THERMO PHASE
Note: LISS may be used instead of 22% Bovine Serum Albumin if the tubes proceeding to the next step will be
incubated for 10 minutes
2. Thermo/Incubation Phase (37°C)

a. Incubate Ma – P and Mi – P tubes showing no agglutination for 30 minutes at 37°C water bath.
b. Centrifuge for 15 seconds at 3,400 rpm.
c. Gently dislodge cell button and examine for agglutination or hemolysis.
d. If no agglutination or hemolysis is observed in tubes Ma – P and Mi – P, proceed to ANTIHUMAN GLOBULIN
PHASE
3. Antihuman Globulin (AHG) Phase/Coombs’ Phase

a. Wash cells of tubes Ma – P and Mi – P with NSS for 3 times.
b. Decant NSS completely after the last washing.
c. Add 2 drops of antihuman globulin sera.
d. Centrifuge for 15 seconds at 3,400 rpm.
e. Dislodge cell button and examine for hemolysis or agglutination.
f. Add 1 drop of Coombs check cells to tubes showing no agglutination.
Interpretation:
▪ COMPATIBLE = NO agglutination or hemolysis in all tubes
▪ INCOMPATIBLE = PRESENCE of agglutination or hemolysis in any tubes
Guide in the Interpretation of Crossmatching Test Results:

CROSSMATCH
CASE PROCEDURAL PHASE INTERPRETATION
MAJOR MINOR
Immediate Saline
0 0
Spin/Protein Phase
1
Blood is compatible both in major and minor
Thermo Phase 0 0
crossmatch. Blood is safe for transfusion.
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AHG Phase 0 0
Immediate Saline
+ 0
Spin/Protein Phase
Blood is incompatible in major crossmatch but
2 Thermo Phase 0 compatible with minor crossmatch. Blood is NOT
safe for transfusion.
AHG Phase 0
Immediate Saline
+ +
Spin/Protein Phase
Blood is incompatible both in major and minor
3 Thermo Phase
crossmatch. Blood is NOT safe for transfusion.
AHG Phase
Immediate Saline
0 +
Spin/Protein Phase
Blood is compatible in major crossmatch but not
4 Thermo Phase 0 incompatible in minor crossmatch. Blood can be
transfused but with caution.
AHG Phase 0
DRAWINGS:
1. Illustrate and label accordingly the important steps and standard results of slide (emergency) crossmatch
2. Illustrate and label accordingly the important steps and standard results of broad-spectrum crossmatch

1. Discuss the significance of the components of compatibility test in transfusion medicine
2. What is (a) a major crossmatch and (b) a minor crossmatch? Discuss the purpose of performing crossmatch
3. What incompatibilities are detected in the (a) protein phase, (b) thermo phase, and (c) antihuman globulin phase
of broad-spectrum crossmatch?
4. Why is antibody screen and identification preferred over minor crossmatch
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Name: Rating:
ANTIBODY SCREEN AND IDENTIFICATION
***The procedure shall be presented by the instructor thru a reference video/recorded video or via virtual lab
Antibody screen and identification is used to detect unexpected antibodies. These antibodies may be classified as
immune (the result of RBC stimulation in the patient), passive (transferred to the patient through blood products or
derivatives), or naturally occurring (the result of environmental factors). Antibodies may also be classified as
alloantibodies, directed at foreign antigens, or autoantibodies, directed at one’s own antigens.
It applies the principle of antiglobulin test. The procedure uses blood group O screen cells and panels cells to detect
antibodies other than the naturally occurring anti – A and anti – B.
OBJECTIVES:
▪ Describe the phases and uses of antibody screen and identification
▪ Apply the exclusion method for antibody identification
▪ Differentiate antibody screen from panel testing
MATERIALS:
Test tubes and rack
Venipuncture set
Pasteur pipette
REAGENTS:
Screening Cells 1 and 2 (SC – 1 and SC – 2)
Panel Cells 1 – 11 (PC – 1 to PC – 11)
Antihuman globulin reagent
SAMPLE:
Unknown serum (should be free from contamination, hemolysis and lipemia)
PROCEDURE:
1. Antibody Screening
a. Prepare two tubes and follow the table below
Content Cell – 1 Cell – 2
Unknown serum 2 drops 2 drops
Screening Cell – 1 (SC – 1) 2 drops -
Screening Cell – 2 (SC – 2) - 2 drops
b. Immediate Spin (IS) phase: Mix the contents. Cover with Parafilm.
c. centrifuge for 15 seconds at 3,400 rpm
d. dislodge the cell button gently and observe for agglutination. Record your result from the given antibody screening
antigram.
e. If no agglutination is observed, proceed to the next step
f. Incubation phase (37°C): incubate the tube showing no agglutination at 37°C water bath for 15 minutes.
g. Centrifuge for 15 seconds at 3,400 rpm. Dislodge the cell button gently and observe for agglutination.
Record your result from the given antibody screening antigram.
h. If no agglutination is observed, proceed to the next phase.
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i. Antihuman Globulin (AHG) phase: wash the unknown tube showing no agglutination three times with NSS.
Decant completely after the last washing.
j. Add 2 drops of antihuman globulin serum and mix well. Cover with Parafilm.
k. Centrifuge for 15 seconds at 3,400 rpm.
l. Dislodge the cell button gently and observe for agglutination. Record your result from the given antibody
screening antigram.
Note: Agglutination indicates the presence of atypical antibodies in the patient’s serum.
No agglutination indicates the absence of atypical antibodies in the patient’s serum
m. If the rest does not show agglutination, add 1 drop of Coombs check cells (CC) to the unknown tube and
centrifuge for 15 seconds at 3,400 rpm.
n. Dislodge the cell button gently and observe for agglutination
ANTIBODY SCREENING ANTIGRAM

Rh Kell Duffy Kidd Lewis P MNSs Lutheran Reaction
Cell
a b a Jk a b a b 37°
No. D C E c e K k Fy Fy Jk b Le Le P1 M N S s Lu Lu IS AHG CC
C
SC1 + 0 + + + + + 0 + + + + 0 + + + + 0 0 +
SC2 0 + 0 + + + 0 + 0 0 + 0 + 0 + 0 0 + + 0
List all possible antibodies present in the unknown serum:
2. Antibody Identification
a. Prepare 11 tubes and label them as 1 to 11 and follow the table below:
Tube Labels
Content
1 2 3 4 5 6 7 8 9 10 11
Unknown 2 2 2 2 2 2 2 2 2 2 2
Serum drops drops drops drops drops drops drops drops drops drops drops
2
PC – 1 -- -- -- -- -- -- -- -- -- --
drops
2
PC – 2 -- -- -- -- -- -- -- -- -- --
drops
2
PC – 3 -- -- -- -- -- -- -- -- -- --
drops
2
PC – 4 -- -- -- -- -- -- -- -- -- --
drops
2
PC – 5 -- -- -- -- -- -- -- -- -- --
drops
2
PC – 6 -- -- -- -- -- -- -- -- -- --
drops
2
PC – 7 -- -- -- -- -- -- -- -- -- --
drops
2
PC – 8 -- -- -- -- -- -- -- -- -- --
drops
2
PC – 9 -- -- -- -- -- -- -- -- -- --
drops
2
PC – 10 -- -- -- -- -- -- -- -- -- --
drops
2
PC – 11 -- -- -- -- -- -- -- -- -- --
drops
Note: Prepare autocontrol tube (tube 12) and test together with the other tubes. Label tube as “AUTO” then add 2
drops of patient’s (unknown) serum and 2 drops of patient’s red blood cells
b. Immediate Spin (IS) phase: Mix the contents. Cover with Parafilm and centrifuge for 15 seconds at 3,400 rpm
c. Dislodge the cell button gentle and observe for agglutination. Record your result from the given antibody
identification antigram.
d. If no agglutination is observed, proceed to the next step.
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e. Incubation phase (37°C): incubate the tube showing no agglutination at 37°C water bath for 15 minutes.
f. Centrifuge for 15 seconds at 3,400 rpm. Dislodge the cell button gently and observe for agglutination.
Record your result from the given antibody identification antigram.
g. If no agglutination is observed, proceed to the next phase.
h. Antihuman Globulin (AHG) phase: Wash the unknown tube showing no agglutination three times with NSS.
Decant completely after the last washing.
i. Add 2 drops of antihuman globulin serum and mix well. Cover with Parafilm.
j. Centrifuge for 15 seconds at 3,400 rpm
k. Dislodge the cell button gently and observe for agglutination. Record your result from the given antibody
identification antigram.
Note: Agglutination indicates the presence of atypical antibodies in the patient’s serum.
No agglutination indicates the absence of atypical antibodies in the patient’s serum.
l. If the test does not show agglutination, add 1 drop of Coombs check cells (CC) to the unknown tube and
centrifuge for 15 seconds at 3,400 rpm.
m. Dislodge the cell button gently and observe for agglutination
ANTIBODY SCREENING ANTIGRAM

Rh Kell Duffy Kidd Lewis P MNSs Lutheran Reaction
Cell
Jk P 37°
No. D C E c e K k Fya Fyb Jka b Lea Leb M N S s Lua Lub IS AHG CC
1 C
1 + + + 0 + + 0 0 + + + 0 + + 0 + 0 0 + 0
2 + 0 0 + + 0 + + + + 0 + 0 + 0 0 + + 0 +
3 0 0 + + + + + + + 0 + 0 + + + 0 + + 0 +
4 + + 0 0 + + 0 + 0 0 + + 0 + 0 + + + 0 +
5 0 + + + 0 0 0 + + + 0 0 + 0 + + 0 + 0 +
6 0 + 0 + + + 0 + + 0 0 + 0 + + 0 + 0 + 0
7 + 0 0 + + + + + 0 + + + 0 0 + + + + 0 +
8 + + + 0 + + + 0 + + 0 + + 0 + + 0 + + +
9 0 + 0 + + 0 0 + + 0 0 + 0 + + 0 + 0 + 0
10 0 + + + 0 + + 0 + 0 + + + 0 + + 0 + + 0
11 + 0 + + + + 0 0 + + 0 + + + 0 0 + 0 + +
Auto
Interpretation of Result:
Unknown antibody: _______________________________________________________________________________
Type of antibody: ________________________________________________________________________________
Clinical significance: ______________________________________________________________________________
DRAWING:
1. Illustrate and label accordingly the important steps antibody screen
2. Illustrate and label accordingly the important steps antibody identification

1. Describe (a) antibody screen and (b) antibody identification
2. Discuss the purpose of antibody screen and identification in pre-transfusion testing
3. Enumerate and briefly describe the factors to be considered in antibody screen and identification
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D. ASSESSMENT:
Offline learners: Answer the given activity and send it to your instructors’ email. Quiz will be given through text/email.
Online Learners: Answer the given activity.
LECTURE ASSESSMENT:
Answer whether it is Compatible or Incompatible given the blood types of the donor and recipient.
MAJOR Crossmatching
Donor
Blood Type O B AB A
B
Recipient
AB
MINOR Crossmatching
Recipient
Blood Type AB B O A
AB
B
Donor
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LABORATORY ASSESSMENT (EXPERIMENT nos. 13 and 14)
I. Identification
_________________________1. It is one of the principal tools for investigating potential HTR’s and immune hemolytic
anemias.
__________________________2. These antibodies are produced in response to RBC stimulation through transfusion,
transplantation, or pregnancy.
__________________________3. These are antibodies produced in one individual and then transmitted to another via
plasma containing blood components or derivatives such as IVIG.
__________________________4. What is the ABO blood type of antibody screening cells?
__________________________5. TRUE or FALSE. A positive antibody screen occurs as a result of prior exposure to red
cell antigens from pregnancy or transfusions.
__________________________6. Term used to refer to a list of antigens present in the reagent red cell suspension.
__________________________7. Other term for enhancement reagents.
__________________________8. Polyspecific AHG reagent is also known as __________
__________________________9. In the gel method, screen cells are suspended in LISS to a concentration of __
__________________________10. In the gel method, how are you going to interpret a result that is characterized by
“cells trapped in gel”?
__________________________11. An antibody identification panel is a collection of how many blood type specific
RBC’s?
__________________________12. It determines whether alloantibody or autoantibody specificity exists.
__________________________13. In the gel method, how are you going to interpret a result that is characterized by
“cells in pellet at bottom of microtube”?
__________________________14. Give at least 2 potentiators used in the laboratory
__________________________15.
B. Antibody screen. Identify the antibody causing the incompatibility
E. REFERENCES
2. Harmening, D. (2019). Modern Blood Banking and Transfusion Practices (7th ed.) F. A. Davis Company
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Name: Rating:
BLOOD DONATION PROCESS
Blood centers and transfusion services depend on volunteer donors to provide the provide the blood necessary to meet
the needs of the patient they serve. To attract volunteer donors and encourage their continued participation, it is essential
that conditions surrounding blood donation be as pleasant, safe, and convenient as possible. To protect donors and
recipients, donors are questioned about their medical history and are given a mini physical examination to help blood
center staff determine whether they are eligible donors. The phlebotomy is conducted carefully to minimize any potential
donor reactions or bacterial contamination of the unit (AABB Technical Manual, 15 th edition)
OBJECTIVES:
▪ Describe the important steps of the blood donation process
▪ Understand the rationale behind the procedures and tests for identifying appropriate blood donors
MATERIALS:
Capillary tube (with parafilm wax and clay seal) Sphygmomanometer
Hemofuge and microhematocrit reader Weighing scale
Blood pressure (BP) apparatus Beaker (50 mL)
Blood bag and hemostat Stopwatch
Thermometer (digital) Lancet
REAGENTS:
Copper sulfate solution (specific gravity 1.053 – 1.055)
Anti – A, anti – B and anti – D antisera
PROCEDURE: Blood donation process includes:
A. Donor Registration
The donor must fill up the necessary information and details on the Blood Donor Interview Data Sheet
B. Interview and Physical Examination
The donor recruitment personnel of the blood bank shall have an interview with the donor to discuss the important
details of the procedure and to respond to concerns that might be raised by the donor.
Furthermore, it also includes physical examination by the blood bank physician after obtaining details of the donor’s
weight, blood pressure, hemoglobin, hematocrit and blood type.
Weight determination:
1. Ask the donor to remove shoes, heavy clothing or any apparel that may add to the standard weight requirement
for a donor
2. Ask the donor to stand straight on the weighing scale
3. Determine the weight
4. Record the weight in pounds and kilograms on the blood donor interview data sheet
Remember: 1 kilogram = 2.2 pounds
Note:
a. The weight tells you the maximum amount of blood that can be withdrawn from a donor
b. The maximum allowable blood is suspended in a standard 63 mL of anticoagulant present in the primary
blood bag
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c. If the weight is <110 pounds or <50 kilograms, the blood bank may collect lesser amounts of blood provided
that the standard amount of anticoagulant will be adjusted and the blood collected will be used within 24
hours. Removal of anticoagulant means that the procedure is already an open system
d. Formula to be used:
Maximum volume of blood to donate
= donor’s weight in pounds/110 pounds x 450 mL ; or
= donor’s weight in kilograms/50 kilograms x 450 mL
Volume of anticoagulant needed to suspend collected blood

= maximum volume of blood to donate/100 x 14
Amount of anticoagulant to be removed from the primary bag

= 63 mL – volume of anticoagulant needed to suspends collected blood
Blood pressure (BP) determination

1. Ask the donor to sit comfortably on an arm chair
2. Place the inflatable cuff on the donor’s arm
3. Locate the brachial artery and place the chest piece of the stethoscope to hear the beating sounds
4. Squeeze the BP apparatus bulb until such time the pressure is enough for you not to hear any beating sounds
(approximately 200 mmHg)
5. Slowly release the pressure and wait for the first loud sound to reappear. This is the donor’s systolic pressure.
6. Continue releasing the pressure until such time the sound ceases. This represents the donor’s diastolic pressure.
7. Record the blood pressure (systolic/diastolic) in mmHg on the Blood Donor Interview Sheet.
a. The blood pressure of the potential donor should be within the allowable limits:
✓ Systolic blood pressure: ≤ 180 mmHg
✓ Diastolic blood pressure: ≤ 100 mmHg
b. If the initial blood pressure is high and the donor is not known to be hypertensive, tell the donor to rest for
approximately 15 – 30 minutes to repeat the BP determination. If the BP is still high, defer the donor and refer
to the blood bank physician
Pulse rate determination

1. Ask the donor to sit comfortably on an arm chair
2. Locate the radial artery
3. Count the number of regular beats in exactly 1 minute by using your timer.
4. Record the pulse rate in beats per minute on the Blood Donor Interview Data Sheet
Note:
a. The acceptable pulse rate for a potential donor is 50 – 100 beats per minute
b. If the pulse rate is higher, allow the donor to rest for 15 minutes to take the pulse rate again. If the pulse rate
is still high, defer the donor and refer to the blood bank physician
Temperature determination
1. Ask the donor to sit comfortably on a chair
2. Place the digital thermometer in between his or her armpit for about five minutes
3. Record the temperature in °C and °F on the Blood Donor Interview Sheet
Remember: °F = 1.8 °C + 32; °C = (°F – 32)/1.8
Note:
a. Standard mandates that the donor temperature must be ≤ 37.5 °C or ≤ 99.5 °F
b. Higher temperature indicates active or current infection. Donor must be further evaluated by a blood bank
physician
c. Donors are advised not to have any coffee or hot beverages intake while waiting
Hemoglobin determination
1. Place 30 mL of CuSO4 solution (specific gravity: 1.053 – 1.055) in a small beaker
2. Ask the donor to sit comfortable on a chair
3. Perform a skin puncture on the ring finger
4. Wipe the first drop of blood
5. Collect an amount of blood with capillary tube enough to produce a drop
6. Hold the capillary tube with blood in vertical position over the CuSO 4 solution.
7. Let the drop fall freely on to the CuSO4 solution.
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Note: the distance of the blood drop to the CuSO 4 must be __________.
8. Take note of the following:
a. If the drop of blood sinks within 15 seconds, the hemoglobin level is estimated to be > 12.5 g/dL
b. If the drop of blood floats or simply suspends, the hemoglobin level is estimated to be < 12.5 g/dL
9. Record the hemoglobin level as > 12.5 g/dL or < 12.5 g/dL on the Blood Donor Interview Data Sheet
Hematocrit determination
1. Ask the donor to sit comfortably on a chair
2. Perform a skin puncture on the ring finger
3. Wipe the first drop of blood
4. Squeeze the finger until such time a solid drop is formed from the finger
5. Collect the blood using a heparinized capillary tube until ¾ filled by using the same puncture site from hemoglobin
determination
6. Seal one end with clay and permanently secure the content with paraffin wax. Label the capillary tube correctly.
7. Spin the capillary tube in a microhematocrit centrifuge for about 15 minutes.
8. Read the packed cell volume using the hematocrit reader.
9. Record the hematocrit as percentage on the Blood Donor Interview Data Sheet.
Note: the hemoglobin level of the donor should be ≥ 12.5 g/dL and the hematocrit level ≥ 38% for allogeneic
donation. If the level is less than the allowable value, it may result to a deferral.
Blood type determination
1. Divide a glass slide using a marker and label the portions as A, B and Rh respectively
2. Using the same puncture site from hemoglobin and hematocrit determination, place 3 separate drops of blood on
the glass slide which were previously labeled as A, B and Rh
3. Deliver anti – A on the glass slide portion labeled as A, anti – B on the glass slide portion labeled as B and anti –
D on the glass slide portion labeled as Rh
4. Mix the blood sample and antiserum using different applicator sticks
5. Interpret the reaction and determine the ABO and Rh type
6. Record the ABO and Rh type on the Blood Donor Interview Data Sheet
Note: Screening of blood donors will not proceed to the next series of tests for blood transmissible diseases if the
donors failed in the preliminary test procedures. This is to save on the blood bank screening reagents
Tests for the blood transmissible diseases:
1. Malarial parasite determination: The gold standard for malarial parasite diagnosis is the thick and thin smear.
However, for the purposes of blood donor screening, a thin smear is prepared from the collected blood. Blood
may be from the skin puncture done for hemoglobin and hematocrit determination or from EDTA blood collected
through venipuncture.
a. Place a drop of blood on one portion of a glass slide.
b. Using another glass slide, perform the standard procedure in smear preparation
c. Allow the smear to dry. Proceed to proper staining using Giemsa.
d. Examine the stained slide using oil immersion objective (OIO)
e. Report as POSITIVE or NEGATIVE for the presence or absence respectively of Plasmodium species
f. Record the result on the Blood Donor Interview Data Sheet
Note:
Immunochromatography (rapid cassette-based) test can be done for serologic detection of malarial parasite
using whole (anticoagulated) blood
2. Serological tests of other blood transmissible disease
a. Perform a venipuncture and place the blood sample in a red top tube
b. Allow the blood to clot, rim, centrifuge then separate the serum
c. Test serum for HIV, HCV, HBsAg and Treponema pallidum infection
d. Report either REACTIVE or NONREACTIVE
e. Record the result on the Blood Donor Interview Data Sheet
C. Blood Collection from Donor

Proper Bleeding Procedure
1. Ask donor to confirm his or her identification.
2. Ensure that all labeling on blood container, processing tubes, retention segment, and donor records is correct.
3. Prepare donor’s arm
a. Apply tourniquet or blood pressure cuff (40 – 60 mmHg); identify venipuncture site, then release tourniquet or
cuff.
151 | P a g e
b. Scrub area at least 4 cm (1.5 inches) in all directions from the intended site of venipuncture (i.e., 8cm or 3
inches in diameter) for a minimum of 30 seconds with 0.7% aqueous solution of iodophor compound. Excess
foam may be removed, but the arm need not be dry before the next step.
c. Starting at the intended site of venipuncture and moving outward in a concentric spiral, apply “prep” solution;
let stand for 30 seconds or as indicated by manufacturer.
d. Cover the area with dry, sterile gauze until the time of venipuncture. After the skin has been prepared, it must
not be touched again. Do not repalpate the vein at the intended venipuncture site.
4. Inspect bag for any defects and discoloration. The anticoagulant and additive solutions should be inspected for
particulate contaminants.
5. Position bag below the level of the donor’s arm.
a. If a balance system is used, be sure the counterbalance is level and adjusted for the amount of blood to be
drawn. Unless metal clips and a hand sealer are used, make a very loose overhand knot in the tubing. Hang
the bag and route the tubing through the pinch clamp. A hemostat should be applied to the tubing before the
needle is uncapped to prevent air from entering the line.
b. If a balance system is not used, be sure to monitor the volume of blood drawn.
6. Reapply tourniquet or inflate blood pressure cuff. Ask the donor to open and close hand until previously selected
vein is again prominent.
7. Uncover sterile needle and perform the venipuncture immediately. A clean, skillful venipuncture is essential for
collection of a full, clot-free unit. Once the bevel has penetrated the skin, palpation of the skin above the needle
stem may be performed with a gloved finger, provided the needle is not touched. When the needle position is
acceptable, tape the tubing to the donor’s arm to hold the needle in place and cover the site with sterile gauze.
8. Release the hemostat. Open the temporary closure between the interior of the bag and the tubing.
9. Ask the donor to open and close hand slowly every 10 to 12 seconds during collection.
10. Keep the donor under observation throughout the donation process. The donor should never be left unattended
during or immediately after donation.
11. Mix blood and anticoagulant gently and periodically (approximately every 45 seconds) during collection. Mixing
may be done by hand or by continuous mechanical mixing.
12. Be sure blood flow remains fairly brisk, so that coagulation activity is not triggered. If there is continuous,
adequate blood flow and constant agitation, rigid time limits are not necessary. However, units requiring more
than 15 minutes to draw may not be suitable for preparation of platelets, fresh frozen plasma (FFP), or
cryoprecipitated antihemophilic factor (AHF). The time required for collection can be monitored by indicating the
time of phlebotomy or the maximal allowable time (start time plus 15 minutes) on the donor record.
13. Monitor volume of blood being drawn. If a balance is used, the device will interrupt blood flow after the proper
amount has been collected. One mL of blood weighs at least 1.053 g, indicated by the minimum allowable specific
gravity for donor. A convenient figure to use is 1.06 g/mL; a unit containing 405 to 550 mL should weigh 429 to
583 g plus the weight of the container and anticoagulant. For a 500-mL bag, this is 565 to 671 g.
14. Clamp the tubing near the venipuncture using a hemostat, metal clip, or other temporary clamp.
Release the blood pressure cuff/tourniquet to 20 mm Hg or less and fill the tube(s) for blood processing sample(s)
by a method that prevents contamination of the contents of the bag.
15. Deflate the cuff and remove the tourniquet. Remove the needle from the donor’s arm, if not already removed.
Apply pressure over the gauze and ask the donor to raise his or her arm (elbow straight) and hold the gauze
firmly over the phlebotomy site with the other hand.
16. Discard the needle assembly into a biohazard container designed to prevent accidental injury to, and
contamination of, personnel.
17. Strip donor tubing as possible into the bag, starting at the seal. Work quickly, to prevent the blood from clotting in
the tubing. Invert the bag several times to mix the contents thoroughly; then allow the tubing to refill with
anticoagulated blood from the bag. Repeat this procedure a second time.
Note: Collect blood samples for laboratory testing.
18. Seal the tubing attached to the collection bag into segments, leaving a segment number clearly and completely
readable. Attach a unit identification number to one segment to be stored as a retention segment. Knots, metal
clips, or a dielectric sealer may be used to make segments suitable for compatibility testing. It must be possible to
separate segments from the unit without breaking sterility of the bag. If a dielectric sealer is used, the knot or clip
should be removed from the distal end of the tubing after a hermetic seal.
19. Reinspect the container for defects.
20. Recheck numbers on the container, processing tubes, donation record, and retention segment.
21. Place blood at appropriate temperature. Unless platelets are to be removed, whole blood should be placed at 1 to
6°C immediately after collection. If platelets are to be prepared, blood should not be chilled but should be stored
in a manner intended to reach a temperature of 20 to 24°C until platelets are separated. Platelets must be
separated within 8 hours after collection of the unit of Whole Blood.
152 | P a g e
Adverse Events in Blood Donation

1. Hematoma
a. Causes:
• Poor or failed venipuncture
• Skin pierced at too great an angle and exiting vein
• Needle puncturing the vein twice during the donation
• Inadequate pressure after the donation
b. Management:
• Apply pressure and a firm bandage
• Advise donor to move arm freely but to avoid heavy lifting
• Apologize, and reassure the donor
2. Vasovagal reaction or faint, due to a hypothalamic response resulting in bradycardia, vomiting, sweating, arterial
dilatation and a low blood pressure
a. Causes:
• Anxiety
• Lowered blood volume and other associated causes: hypoglycemia, lack of fluids, poor sleep
• Atmosphere in donation room (hot or humid)
Signs and symptoms
• Staring Drop in blood pressure
• Sighing Vomiting
• Pallor or sweating Loss of consciousness (occasionally)
• Slow pulse Convulsions (rare)
b. Management:
Mild vasovagal reaction
• Discontinue donation Reassure donor
• Recline chair Give fluids to the donor to drink (recovery is
• Loosen clothes usually rapid)
• Monitor blood pressure and pulse
Severe vasovagal reaction
• Call physician
• If the donor becomes unconscious, put the person in recovery position (i.e. head to the side and chin
up) and ensure that airways are clear
3. Delayed faint (syncope)
a. Causes
• Physical stress
• Inadequate fluid intake
• Cause unknown
Occurs 1 – 4 hours after donation, usually outside the blood bank
b. Management
• Hot drinks or water before donating blood; sitting in a supine position, audio or visual distraction; and
minimal pain and stress during blood donation
4. Arterial puncture
a. Causes:
• Brachial artery sometimes lies anatomically very close to the vein
• Detected by observing that the blood collected is bright red and has a rapid flow
b. Management:
• Discontinue donation or continue if identified towards the completion of the donation
• Call the donor care physician
• Apply firm pressure (by the nurse or medical staff) for at least 15 minutes
• Apply pressure bandage and check the radial pulse
• Inform and reassure donor, and explain that the puncture is unlikely to have serious consequences, but
that bad bruising may occur, and healing takes about 10-14 days
5. Nerve damage
a. Causes:
• Nerve endings brushed during venipuncture; pressure from hematoma
Symptoms and signs:
153 | P a g e
• Pain or paresthesia
• Motor or sensory loss
b. Management:
• Recovery is usually spontaneous and rapid within 24 hours (in rare cases, up to 6 months)
• Refer the donor to the physician to explain and reassure the donor, and refer the donor to a neurologist if
the damage is severe
After blood donation

Donor Care:
After blood has been collected:
1. Ask the donor to remain in the bed or chair and relax for a few minutes (around 10 – 12 minutes under close
pressure);
2. Inspect the venipuncture site; if it is not bleeding, apply a bandage to the site; if it is bleeding, apply further
pressure;
3. Ask the donor to sit up slowly and ask how the person is feeling;
4. Offer the donor some refreshments
5. Give the donor instructions about post phlebotomy care
a. Eat and drink refreshments which were provided before leaving the blood donation facility
b. Do not leave until permitted by the phlebotomist or blood bank physician
c. Drink more fluids than usual in the next 4 hours
d. The donor is advised to refrain from consuming alcoholic beverages until a complete meal has been taken in
e. Smoking is prohibited 30 minutes from the time of blood donation
f. If there is bleeding from the phlebotomy site, raise the arm above heart level and apply pressure
g. If feeling faint or dizzy, lie down or sit down lowering the head between the knees, or return to the blood bank
or see a physician
h. May resume all normal activities after about half an hour if feeling well. Recommend not to perform strenuous
activities or engage in critical work for 24 hours from the time of donation where maximum abilities are
required
6. Before the donor leaves the donation room, ensure that the person can stand up without dizziness and without a
drop in blood pressure
Blood Unit and Samples

1. Transfer the blood unit to a proper storage container according to the blood centre requirements and the product
2. Ensure that collected blood samples are stored and delivered to the laboratory with completed documentation, at
the recommended temperature, and in a leak-proof, closed container
154 | P a g e
Name: Rating:
Experiment Number 15 Output

BLOOD DONATION PROCESS
After identifying your prospective donor, accomplish the following:
BLOOD DONOR INTERVIEW DATA SHEET

I. PERSONAL DATA (to be filled up by the donor)
NAME:
Surname First Name Middle Name
Birthdate Age Civil Status Sex
Nationality Religion Educational Attainment Occupation
PERMANENT ADDRESS
No. Street Barangay Town/Municipality Province/City Zip Code
Office Address
CONTACT
INFORMATION
Telephone No. Cell Phone No. E – Mail Address
IDENTIFICATION
No.
School Company PRC Driver’s SSS/GSIS/BIR Others
II. MEDICAL HISTORY (Please read carefully and check √ the appropriate answer)
YES NO REMARKS
1. Do you feel well and healthy today?
Have any flu or cold?
2. Have you ever been refused as a blood donor or told not to donate blood?
3. Have you within the last 12 hours had an alcohol intake?
4. Do you intend to drive a heavy transport vehicle or operate a heavy machine
within the next 12 hours?
5. Do you intend to ride/pilot an airplane within 24 hours?
6. In the past 72 hours have you had a tooth extraction?
7. In the last 3 days have you taken aspirin?
8. In the past 4 weeks have you taken any medication? Vaccinations?
9. In the past 3 months have you:
Had any chicken pox and/or cold sores?
155 | P a g e
Travelled or lived outside of your place of residence?

Donated whole blood, platelet or plasma?
10. For the past 6 months have you been under the doctor’s care for a certain
disease/illness
11. Have you ever had the following:
Cancer, blood disease or bleeding disorder (haemophilia)?
Heart disease/ surgery. Rheumatic fever or chest pains?
Lung disease, tuberculosis or asthma?
Kidney disease, thyroid disease, diabetes, epilepsy?
Any other chromatic medical conditions or surgical operations?
12. Have you ever had malaria in the past?
13. Have you had jaundice/hepatitis/personal contact with person who had
hepatitis?
14. From 1980 to present, did you spend time in the United Kingdom or Europe?
(CJD or madcow disease)
15. In the past year:
Have you traveled or lived-in other countries?
Have you been incarcerated, jailed or imprisoned?
Have you taken prohibited drugs (orally, by nose, or by injection)?
16. Have you received blood or blood products and/or had tissue/organ
transplant or graft?
Have you had a tattoo applied, ear piercing, acupuncture, accidental
needle stick injury or come in contact with someone else’s blood?
17. Have you ever had any sexually transmitted disease like syphilis, gonorrhea,
HIV/AIDS, etc?
Have you engaged in unprotected or unsafe sex?
18. Are you giving blood only because you want to be tested for HIV or the AIDS
virus, or hepatitis virus?
19. Are you aware that an HIV infected person can still transmit the virus despite
a negative AIDS test?
FOR FEMALE DONORS ONLY
20. Are you currently pregnant?
21. Have you ever been pregnant?
22. When was your last delivery? Date:
23. Did you have an abortion in the 1 year?
24. When was your last menstrual period? Date:
25. Are you currently breastfeeding?
from Philippine Red Cross-National Blood Services (PRC-NBS) Form 312-E Revised 2012
III. DONOR SCREENING (to be filled up by interviewer or donor recruitment officer/personnel)

Type of Donation:
IN – HOUSE
Walk – in/ Voluntary Replacement
Patient directed Patient Name: Hospital:
MOBILE BLOOD DONATION Place: Organizer:
Has history of previous donation?

Number or times: ________________
Date of last donation: _____________ Place of last donation: ___________________
____________________________________
Interviewer (signature over printed name)
IV. INITIAL SCREENING

PHYSICAL EXAMINATION
Weight: Pulse Rate: Hemoglobin:
Blood Pressure: Temperature: Hematocrit:
156 | P a g e
BLOOD SCREENING
Blood Type: HBsAg: RPR:
HIV: HCV: Malaria:
BLOOD BANK PHYSICIAN REMARKS: (to be accomplished by Blood Bank Physician)

General Appearance:
Skin: HEENT: Heart/Lungs:
Decision: Accepted Temporarily deferred Permanently deferred
Reason:
Volume to Donate: 250 mL 450 mL
________________________________________
Blood Bank Physician (signature over printed name)
V. DONOR’S CONSENT
I certify that I am the person referred to above that all the entries read and well understood by me and to the best of
my knowledge, truthfully answered all the questions in this Blood Donor Interview Data Sheet. I understand that all
questions are pertinent for my safety and for the benefit of the patient who will undergo blood transfusion. I am
voluntarily donating my blood through _____________________(state the name of the blood bank). I am aware that
my blood will be tested and screened for blood group, hematocrit, hemoglobin, syphilis, malaria, HIV, hepatitis B and
hepatitis C. I also certify that I am fully aware of the possible risks and consequences of the procedure which were
clearly explained to me by the donor recruitment personnel.
For ages 16 – 17
_______________________
______________________
Signature of Parent or Guardian Relationship to
Donor
___________________________________
Signature over printed name
VI. BLOOD COLLECTION

BLOOD COLLECTION DETAILS (to be accomplished by the phlebotomist)
Blood Bag Used: UNIT SERIAL NUMBER:
Amount of Blood Taken: Start Time: End Time:
Successful: Yes No
Donor Reaction: _______________________________________
Management Done: Phlebotomist (signature over printed name)
DRAWING:
1. Illustrate and label accordingly the important steps and standard results of hemoglobin determination using
copper sulfate method.
2. Illustrate and label accordingly the proper bleeding procedure

1. Describe the importance of screening potential blood donors prior to blood collection
2. Describe the law in the Philippines that has provision about voluntary blood donation
3. Enumerate factors to be considered during blood collection from the donor? Such as the equipments, site of
collection, donor care, etc.
4. Enumerate and briefly describe the different types of blood donation
5. Describe and discuss the significance of performing component preparation
157 | P a g e

Immunohema Lab Activities

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Immunohema Lab Activities

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IMMHMA1: Immunohematology

Laboratory Schedule: Group Number:

Date Performed: Date Submitted:

2. Blood Sampling Set:

3. Blood Collection Set:

QUESTIONS FOR RESEARCH:

Laboratory Schedule: Group Number:

Date Performed: Date Submitted:

Skin Puncture Technique (Capillary Puncture)

QUESTIONS FOR RESEARCH:

Laboratory Schedule: Group Number:

Date Performed: Date Submitted:

4. Red Cell Suspension Preparation (2 – 5%)

b. Preparation of tube with fixed volume of fluid

c. Preparation of RCS for various concentration

QUESTIONS FOR RESEARCH:

Laboratory Schedule: Group Number:

Date Performed: Date Submitted:

1. Demonstration of hemagglutination reactions: Slide Method

2. Demonstration of hemagglutination reactions: Tube Method

g. Gently dislodge the cell button and interpret the results

Guide in the interpretation of Results:

QUESTIONS FOR RESEARCH:

I. For Items #1-6. Identify who discovered or introduced the following

LABORATORY ASSESSMENT (EXPERIMENT nos. 1-4)

Answer Stopper Color Anticoagulant/Additive

B. Order of Draw for Venipuncture

_________18. Heparin tubes with or without gel (green stopper)

Laboratory Schedule: Group Number:

Date Performed: Date Submitted:

Guide in the interpretation of Results:

QUESTIONS FOR RESEARCH:

Laboratory Schedule: Group Number:

Date Performed: Date Submitted:

Guide in the Interpretation of Results:

Type A: presence of anti – B that shows agglutination in B and AB cells

QUESTIONS FOR RESEARCH:

Laboratory Schedule: Group Number:

Date Performed: Date Submitted:

1. Collection and Preparation of Saliva

2. Test for Secretor Status

b. Prepare 5% red cell suspension of A, B and O cells

Guide in the Interpretation of Result:

QUESTIONS FOR RESEARCH:

Laboratory Schedule: Group Number:

Date Performed: Date Submitted:

Guide in the Interpretation of Result in Screening Low Titer O:

+ = with agglutination; 0 = no agglutination

QUESTIONS FOR RESEARCH:

LABORATORY ASSESSMENT (EXPERIMENT nos. 5-8)

_____________12. Blood type A

Laboratory Schedule: Group Number:

Date Performed: Date Submitted:

Guide in the Interpretation of Results:

d. Mix gently. Cover each tube with Parafilm.

Guide in the Interpretation of Results:

QUESTIONS FOR RESEARCH:

Laboratory Schedule: Group Number:

Date Performed: Date Submitted:

9. Gently dislodge each cell button and examine for agglutination.

Guide in the Interpretation of Results:

QUESTIONS FOR RESEARCH:

LABORATORY ASSESSMENT (EXPERIMENT no. 8 and 9)

Laboratory Schedule: Group Number: