Download Lab 4

Human Anatomy and Physiology. Frolich, Page 1
Blood Lab Instructions and Wish List
Blood Lab
MY NAME: ______________
MY BLOOD SAMPLE #:_____
(from microscope slide—use same tube slot in centrifuge)
Goals for this activity:
1. Safely acquire and analyze a sample of a few drops of blood
2. Use a centrifuge to separate out the components of blood, including hematocrit. Understand
why that works and what you are seeing in the capillary tube with your blood in it.
3. Type your own blood and appreciate what blood type means in regard to the antigens on your
blood cells and what types of donors can be used for transfusions to what types of recipieints.
4. View your own blood under the microscope and identify its various cellular components.
Getting the Blood—Finger Prick
You will need three drops of blood for three purposes
 Hematocrit—to see volume of cells and plasma in your own blood
 Blood smear—to view your own blood under microscope
 Blood typing—to see antigen reactions for typing your own blood (A,B,O,Rh)
Be sure you have the following ready before you prick your finger
 Sterile lance
 Alcohol pad
 Hematocrit or capillary tube
 Two microscope slides (for smear)
 Blood typing card
 Helper wearing gloves
Read the entire lab, get everything ready and then do these steps quickly in this order:
1. FINGER PRICK: Wipe with alcohol, let dry, prick with lance—use side of fourth finger.
2. DROP ON MICROSCOPE SLIDE. Apply blood to microscope slide and smear with other slide as
shown below—best to watch video on course website first.
3. HEMATOCRIT TUBE. Touch hematocrit tube to drop on finger so that capillary action sucks it into
tube. Then, holding pricked finger over end of tube, press other end into “play-dough” capping material
to cap tube. Give tube to helper—only helper should wear gloves.
4. DROPS ON TYPING CARD: Apply three blood drops to blood typing card. Have helper add Anti-A,
Anti-B and Anti-D (Rh) reagents and mix right away. Use a separate toothpick to mix each reagent
with the blood on the card. Read results in two minutes.
5. STOP BLEEDING. Wipe again with alcohol swab and then cover with cotton gauze or band-aid if
needed.
Human Anatomy and Physiology. Frolich, Page 2
Blood Lab Instructions and Wish List
Part I: Blood Typing
For two minutes after mixing, watch for agglutination or clumping of your blood/reagent mixture and record
your results:
Anti-A Serum
Anti-B Serum
Anti-D (Rh) Serum
Then compare to determine what type of blood you have. Blood type: ___________
Please be sure you understand, based on what kinds of antigens are made by each kind of blood,
why you get the agglutination reactions for each blood type. Appreciate why Type O are universal
donors and Type AB are universal recipients.
Human Anatomy and Physiology. Frolich, Page 3
Blood Lab Instructions and Wish List
ABO Blood typing is clinically important because if an antigen and an antibody of the
same letter are mixed an agglutination reaction will occur. For example, if a patient
with A antigen on the surface of their RBCs receives a type B blood transfusion
(containing anti-A antibodies) the donor cells will clump together (agglutinate). This
clumping is called a transfusion reaction and the clumped cells can clog small blood
vessels cutting off blood supply to different regions of the body. As some of the RBCs
begin to break down due to the action of phagocytes, hemoglobin is released and may
block kidney tubules leading to kidney failure. Furthermore, ruptured RBCs can no
longer carry O2. If the reaction is severe enough, the patient receiving this mismatched
blood transfusion can die.
Rh blood typing is also important as once an individual receives mismatched blood (e.g.
an Rh- patient receives Rh+ blood), the patient will develop anti-D antibodies which will
lead to a transfusion reaction if this same mistake is ever repeated. The most common
concern, however, is erythroblastosis fetalis. In this case, an Rh- mother carrying an
Rh+ baby may produce anti-D antibodies if some of the fetal blood crosses the placenta
and enters her circulation. Upon a second pregnancy with an Rh- fetus, the anti-D
antibodies can cross the placenta and destroy the fetal RBCs leading to fetal death. To
prevent this, all Rh- mothers are given RhoGAM, a serum derived product, which can
block the mother’s immune system from generating anti-D antibodies by agglutinating
the Rh factor that enters her blood stream. It is therefore given to pregnant mothers who
experience bleeding, at about 28 weeks of pregnancy, and within 72 hours after a birth,
miscarriage, abortion, or amniocentesis.
Part II: Hematocrit
This is the normal % of erythrocytes in a whole blood sample. RBCs normally make up
~45% of the total volume of a blood sample. Leukocytes make up less than 1% of the
total sample. Plasma makes up the rest of the volume (~55%). These percentages can be
determined by centrifuging a sample of whole blood which will cause the red blood cells
to settle to the bottom.
1. Put your tube in the same slot as the number on your microscope slide. All samples from the whole
class can be spun at once. Spin the tubes in the centrifuge for about five minutes.
2. Estimate the percentage of blood volume that is RBC’s or erythrocytes—this is your hematocrit. Can
you see a coat of leukocytes or “white blood cells?”
The Standard method
for taking a
hematocritreading
involves lancing a
finger and then pulling
a blood sample into a
capillary tube. The tube
is then spun in a
centrifuge. The amount
of red blood cells per unit volume is defined
as the hematocrit level.
Human Anatomy and Physiology. Frolich, Page 4
Blood Lab Instructions and Wish List
Hematocrit tube after centrifugation from a
cow with acute lymphoblastic leukemia.
The sample had a white blood cell count of
290,000/µl. The buffy coat is easily
recognized in this tube; it constitutes
nearly half of the cellular mass.
Part III: Blood Smear under Microscope
1. Stain your blood smear with a drop or two of Wright’s stain.
2. Observe the slide under the microscope highest power. You can even use the oil lens to get great
views of the cells. Ask for help if you have not done this before.
3. Be sure you identify all the cell types:
a. Erythrocytes or RBC’s—most abundant cells
b. Platelets: are these really cells?
c. Leukocytes or white blood cells—granulocytes (have distinct granules) and agranulocytes (with
indistinct granules)
DISTINCT GRANULES
Neutrophils: Most numerous WBC’s. Multi-lobed nucleus, pale cytoplasm, fine granules. (considered neutral
because they take up both acidic and basic stains). Immunologically, these are active phagocytes.
Eosinophils: Less numerous and hard to find…let others know if you find one. Bi-lobed nucleus,
reddish/orange granules in the nucleus, stains red because absorbs acidic stain eosin. These phagocyte-type
cells are involved mostly in immune signaling and will burst releasing signal proteins. Also involved in allergic
and inflammatory responses.
Basophils: Least numerous and harder to find…let others know if you find one. Nucleus large and U-shaped,
absorbe basic-staining dye and have large bluish-black granules with histamine. Also contain heparin.
WITHOUT DISTINCT GRANULES
Lymphocytes: Most numerous of agranulocytes and second-most common WBC’s (after neutrophils).
Nucleus is large and round and fills cell. Most lymphocytes are outside blood in lymph tissues. These are the
B-cells and T-cells of the specific immune response.
Monocytes: Largest WBC’s. Kidney-shaped nucleus and light gray-blue cytoplasm. Wandering phagocytes
that can leave blood and move into tissues. Can become fixed macrophages in tissues (e.g. Langerhans cells
in skin or microglial cells in CNS)
Human Anatomy and Physiology. Frolich, Page 5
Blood Lab Instructions and Wish List
Human Anatomy and Physiology. Frolich, Page 6
Blood Lab Instructions and Wish List
Wish List for Blood Lab
FROM BLOOD TYPING
 Antigens: A, B, D (Rh)
 Antibodies: Anti-A, Anti-B, Anti-D (Rh)
 Blood Types: A, B, O, Rh+/ Agglutination reactions
 What blood types are compatible?
 What happens when someone is given blood from a non-compatible donor?
 When is Rh factor (+/-) most important and why?
FROM HEMATOCRIT
 Percentage or fraction
 Hematocrit—normal range
 Plasma, WBC’s or “buff coat”
 What might affect hematocrit?
 How does centrifuge separate out components of blood?
FROM BLOOD SMEAR
 Erythrocytes or RBC’s
 Platelets
 Leukocytes or WBC’s
o Neutrophils
o Eosinophils
o Basophils
o Monocytes
o Lymphocytes
 Know each WBC function in immune response