Human Anatomy and Physiology Lab 1 Blood

Background: I. Blood is a connective tissue composed of formed elements (cells and cell fragments) and intercellular material (plasma)

Blood Sample Centrifuged adult males: 5-6 liters adult females: 4-5 liters

Hematocrit: the portion (% by volume) of the blood that is composed of packed red blood cells adult males: 47% + 5% adult females: 42% + 5%

Objective 1 Plasma Characteristics Plasma is the extracellular (intercellular) material of the connective tissue, blood: Characteristics: 90% H2O 10% solutes: plasma proteins (albumin, antibodies, clotting proteins) nutrients (eg, proteins, carbohydrates, lipids, vitamins) hormones wastes (eg, urea, uric acid, creatinine) dissolved gases (CO2, O2)

Lab Objective 1: In this exercise, you will obtain a sample of sheep plasma and determine its: color transparency pH consistency

Formed Elements: cells and cell fragments Leukocyte Erythrocyte Platelets Erythrocyte/Red Blood Cells (RBCs): anucleate cells that carry oxygen & carbon dioxide 4-6 X 106 cells/mm3 blood Leukocyte/White Blood Cells (WBCs): nucleated cells that provide immunity 4800 - 10,800 cells/mm3 blood Platelets (thrombocyte): cell fragments used for hemostasis 150,000 – 400,000 cells/mm3 blood

Red Blood Cells (erythrocytes): - lack nuclei and organelles Contain hemoglobin (Hb) which binds to O2 (oxyhemoglobin ) & CO2 (carbaminoglobin) - have a “biconcave” shape - diameter averages 7.5 m.

Never Let Monkeys Eat Bananas There are five types of leukocytes organized into two classes: Never Let Monkeys Eat Bananas

Platelets (Thrombocytes): - fragments derived from bone marrow cells (megakaryocytes) contain granules but no nuclei or organelles prevent blood loss (hemostasis)

Objective 2 You will identify each of the formed elements on a prepared blood smear that has been stained with Wright’s stain Wright’s Stain is a mixture of two dyes: 1. Methylene Blue: a basic dye that stains acidic components deep blue/purple 2. Eosin: an acidic dye that stains basic components red/deep pink/orange

Methylene blue (blue dye) Different stains can be applied – one popular stain is a differential stain called Wright’s Stain. Wright’s stain is a mixture of eosin and methylene blue. Methylene blue (blue dye) has a positive charge and stains negatively charged substances (acids found in some granules, and DNA and RNA) structures that combine with methylene blue are called basophilic Eosin (red dye) has a negative charge and stains positively charged substances (bases found in some granules, and hemoglobin) structures that combine with eosin blue are called acidophilic Eosin/Methylene Blue Complex stains neutral substances (such as components of some granules) lilac

When blood is smeared onto a slide, dried and stained with Wright’s stain, the individual formed elements (including types of WBC’s) can be distinguished:

Granulocytes: Neutrophil: 50 – 70% of the circulating WBCs - 9-16 m in diameter - 2-5 nuclear lobes - pale staining, lavender granules

- 2-4% of the total circulating WBCs Eosinophils: - 2-4% of the total circulating WBCs - 10-14 m in diameter - have a bilobed nucleus - bright reddish/orange/pink cytoplasmic granules

- 0 – 1% of the total circulating WBCs Basophils: - 0 – 1% of the total circulating WBCs - 8-10 m in diameter - unsegmented or bilobed nucleus - deep blue/purple cytoplasmic granules

Agranulocytes: Lymphocytes: - 25 - 45 % of the total WBC count - size ranges from 5 m (small) to 17 m (large); small lymphocytes predominate - nucleus is round or slightly oval; it may be indented - clear blue cytoplasm that may be seen only as a ring around the nucleus Small lymphocyte Large lymphocyte

Monocyte - 3-8 % of the total WBC count - 14-24 m in diameter - nucleus is horseshoe shaped or kidney shaped - abundant blue gray cytoplasm that may contain vacuoles

Platelets appear as small, granular cell fragments(2-3m) that may occur singly or in clumps

Objective 3 A differential white blood cell count is performed to determine the relative percentage of each type of WBC It is used to detect diseases, such as acute infection, chronic infection, allergy, parasitic diseases, anemia, HIV infection, and others

In this activity, you will systematically scan a prepared slide and observe a TOTAL of 100 white blood cells - identify each one and record your data to determine the relative percentages of each type of WBC # observed % = X 100 # counted

Each of the leukocytes exists within a range of normal values in peripheral blood. Type of Leukocyte Normal % #/100 Cells If Elevated? Neutrophil 50-70% 50-70 acute infection phagocytic Eosinophil 2-4% 2-4 allergic reaction, parasitic infection asthma Basophil 0.5-1% 0-1 histamine & heparine Lymphocyte 25-45% 25-45 Immunity Monocyte 3-8% 3-8 chronic infection

Activity 4 Blood Typing Pretested blood samples will be tested for the presence of A, B and Rh agglutinogens Type A Type B Type AB Type O Clinically Important Blood Groups Include the ABO System

Rh System – which includes the D agglutinogen Rh+ Rh-

Blood Type is determined by which agglutinogens are present on RBC surfaces: Type A+ Type AB+ Type B- Type AB- Type O-

Agglutinins are antibodies that combine with agglutinogens and cause a clumping reaction called agglutination + Type B Cells Anti-B agglutinin Agglutination

To determine the blood type of a sample, mix separate drops of a blood sample with anti A agglutinin, anti B agglutinin and anti-D agglutinin: Blood Typing Slide With 3 Drops Of Blood To Be Tested, One Drop In Each Well

Add Anti-A serum to this well and mix Add anti-D serum to this well and mix Make sure that you use a separate toothpick for mixing in each well……………. Add Anti-B serum to this well and mix If the agglutinin in the antiserum binds to agglutinogen you will soon observe agglutination

For example: Type B+ Type B- Type O+ Anti A Anti D Serum Serum Anti B Serum Type B+ Type B- Type O+

This is what agglutination will look like !!!!!!!!!!!!!!!!!!!!!!!!

Safety Precautions for Dealing With Blood Samples: Wear gloves Wear goggles Do not test your own blood – use our samples Place any materials that contact blood in bleach solution immediately after use 5. Avoid spilling the samples – do not carry the slides or samples from place to place. If there is a spill, cover with bleach solution immediately and notify your lab instructor

Practice Blood Typing http://nobelprize.org/medicine/educational/landsteiner/index.html