BSC2094 Lab Unit 1-6

**Mnemonic to Remember Which Cells Are Granulocytes and Agranulocytes

"Ben Loves Money": • Granulocytes: Basophil, Eosinophil, Neutrophil • Agranulocytes: Lymphocytes, Monocytes

The clumping of red blood cells that occurs when incompatible blood types are mixed

Agglutination

Antibodies that react to specific surface antigens on red blood cells:

Agglutinins

Agranulocytes

Agranulocytes are noted for the absence of granules in the cytoplasm. As noted previously, the nucleus does NOT undergo segmentation as the granulocytes do.

HEART LOCATION

Apex: -Left fifth intercostal space, slightly medical to the midclavicular line Base: -Formed by the left atrium and by part of the posterior right atrium Right Border: -Superior vena cava, Right atrium, and inferior vena cava Left Border: Left Ventricle

A woman comes to the emergency department bleeding profusely. A quick blood test shows agglutination using the anti-B antibody and the anti-Rh antibody. There was no reaction with the anti-A antibody. What is her blood type?

B+

You have determined that the woman has type B+ blood. The blood bank is running low on blood, which of the following is the best option for your patient?

B-

A patient's blood test revealed an elevated level of neutrophils, a condition called neutrophilia. Which of the following is a possible cause for the neutrophilia?

Bacterial infection Submit

Basophils

Basophils are characterized by large dark granules in the cytoplasm. The nucleus usually is just indented or partially lobed. The granules may cover the entire cell or may be found in a few clusters. Sometimes, the granules may be missing because the granules are soluble in water, which leaves vacuoles (or open-ings) in the cytoplasm. Basophils are the least common of the white blood cells and it is not unusual to see none in a differential count (normal is 0-1%). Basophilia is seen in myxedema, hypothyroid conditions, ulcerative colitis, certain types of anemia, and cancers. Basophils are histamine producers

Formed elements (Blood)

Buffy Coat -1% -Leukocytes and platelets Erythrocytes -45%

The percentage of whole blood that is composed of blood cells:

Hematocrit

HEMATOCRIT (%)

Hematocrit or Packed Cell Volume (PCV): -The percentage of RBCs in whole blood -In the blood tube it is the lower section of fluid (refer to the centrifuged blood tube picture or diagram. -Don't forget units (%) Hematocrit %: -National Institute of Health (NIH) Values -Male 40.1 - 51.0 % -Female 34.1 - 44.9 %

The iron-containing pigment molecule in hemoglobin that binds to oxygen

Heme

With large numbers of erythrocytes dying and being replaced every day, what happens to the heme released from the erythrocytes?

Heme is catabolized into two pigments, biliverdin and bilirubin. Submit

HEMOGLOBIN

Hemoglobin is a measure of the protein that makes up the oxygen carrying portion of the RBC's, composed of four globulin proteins: -Abbreviation: Hb or Hgb -Don't forget units! (g/dl) NIH Values: Male 13.7 - 17.5 g/dl Female 11.2 - 15.7 g/dl

High Hemoglobin

-Anabolic steroids -COPD -Dehydration -Emphysema -EPO -Heart failure -Kidney cancer -Living at high altitude -Polycythemia vera -Pulmonary fibrosis -Smoking

FORMED ELEMENTS

-ERYTHROCYTES -PLATELETS -LEUKOCYTES (PERCENTAGE OF TOTAL WBC'S) -Granulocytes: Neutrophils (60 to 70%) Eosinophils (2 to 4%) Basophils (< 0.5%) -Agranulocytes: Lymphocytes (25 to 33%) Monocytes (3 to 8%)

Cardiac muscle

-Intercalated disks -Involuntary -Only found in the heart

HEMOLYTIC DISEASE OF THE NEWBORN (HDN)

-Normally effects second pregnancy, unless prior exposure to rH positive blood -Father is (+), mother is (-) -Rhogam is the treatment given to prevent miscarriage

EOSINOPHILS

-Nucleus: Bi-lobed -Granules: Red Cytoplasmic -Size: 10 to 14 micrometers -Number: 100 to 400/mm3 -Development: 14 Days -Life Span: 8 to 12 days -Function: Worms (parasites) Antigen-antibody complexes inflammatory and allergies

BASOPHILS

-Nucleus: Lobed -Granules: Large blue-purple Cytoplasmic -Size: 10 to 14 micrometers -Number: 20 to 50/mm3 -Development: 1 to 7 Days -Life Span: A few hours to a few days -Function: Release Histamine Contain Heparin

NEUTROPHILS

-Nucleus: Multilobed -Granules: inconspicuous -Size: 10 to 12 micrometers -Number: 3000 to 7000/mm3 -Development: 14 Days -Life Span: 6 hours to a few days -Function: Phagocytize bacteria; Elevated due to Bacterial Infection

Layers of the heart

-Pericardium -Epicardium -Myocardium -Endocardium

BLOOD TYPING

-Universal Donor: O- -Universal Recipient: AB+ -Don't forget the - or + on the test, it will be wrong!

Low Hemoglobin

-Vitamin and iron deficiencies -Acute or chronic bleeding -Hormone deficiencies, such as thyroid hormone or testosterone -Chronic diseases, such as kidney failure, cancer and Crohn's disease -Autoimmune diseases, such as lupus -Side effect of certain medications, such as chemotherapy drugs

Plasma

-Water (92%) -Proteins: Albumin, Globulins, Fibrinogen -Nutrients: Glucose, A.A.'s, Lactic Acid, Lipids, Iron, Trace elements, Vitamins -Electrolytes: Na+, Ca++, K+, Mg++, Cl-, HCO3-,HPO4--,SO4-- -Nitrogenous Wastes: Urea, Uric acid, Creatinine, Creatine, Ammonia, Bilirubin -Others: Dissolved CO2, O2, N2 Enzymes Hormones

Low Hematocrit Values

-blood loss -cancers -kidney and liver diseases -malnutrition -vitamin B12 and folic acid deficiencies -iron deficiency -pregnancy -systemic lupus erythematosus -rheumatoid arthritis -peptic ulcer disease

High Hematocrit Values

-severe burns -diarrhea -shock -Addison's disease -Dehydration

PROPERTIES (Blood)

4 to 6 liters Plasma 55%

The migration of white blood cells from capillaries to surrounding tissues

Diapedesis

Eosinophils

Eosinophils are characterized by large round or oval granules that pick up the eosin dye, so that they appear red-orange. The nucleus usually stains slightly less intense than a neutrophil and frequently contains only two lobes and rarely more than three. The granules are generally spread evenly over the cell. Normally eosinophils will make up about 3-5% of the white blood cells. An increase in eosinophils (eosinophilia) is often seen in allergies and parasitic worm infestations.

Hemolytic Disease of the Newborn (HDN)

Hemolytic disease of the newborn (HDN) or erythroblastosis fetalis is a disease where the mother's an-tibodies (IgG) cross the placenta and lyse (rupture) the fetus' RBCs. The disease can be mild (as usually found in ABO incompatibility) or severe (usually Rh incompatibility), and fetal death from heart failure (hydrops fetalis) can occur. When the disease is moderate or severe, many erythroblasts (early red blood cells) are present in the fetal blood because the bone marrow is trying to replace the destroyed RBCs, thus the name erythroblastosis fetalis. HDN can occur with an incompatibility of any blood group, but involvement of the ABO and Rh sys-tems is the most frequent. HDN is the most serious when the mother is Rh negative and the fetus is Rh positive (father is Rh positive). Because there are no pre-formed antibodies with the Rh system, the first baby usually does not have a problem. However, childbirth, abortions, ruptures in the placenta during pregnancy, or medical procedures carried out in the pregnancy can cause the mother to be exposed to the fetus' Rh positive RBCs. The mother's immune system will develop anti-D and the antibody will circulate in the mother's plasma. This antibody is small enough to cross the placenta and attack any subsequent pregnancies that are Rh positive. To combat this problem, RhoGAM is given.** RhoGAM is IgG anti-D and the antibodies will lyse any D positive fetal cells before the maternal immune system can react. **Rho is the blood bank's designation for the D antigen and GAM stands for gamma globulin (IgG).** To prevent the development of Rh antibodies, RhoGAM is usually administered during the 28th week of the first pregnancy, and then again within 72 hours after delivery, if the baby is Rh positive. For every subsequent pregnancy after the first, RhoGAM is administered at regular intervals, especially during the second half of the pregnancy. It is also extremely important for Rh-negative women to receive this injec-tion after any miscarriage or abortion in order to avoid future pregnancy complications.

White blood cells are able to move from the blood into the surrounding tissues. What is the functional significance of this ability?

It allows them to travel to the site of an infection and respond quickly to exogenous pathogens.

idEntiFying LEUKoCytEs

Leukocytes are divided into two basic categories, depending on whether there are visible granules in the cytoplasm of the cell. Granulocytes have granules, while agranulocytes generally lack granules (they do have some but they are small and not very conspicuous). Another difference between granulocytes and agranulocytes is the shape of the nucleus of the cell. The size of the cells also differs greatly, so it is important when trying to differentiate the leukocytes, that one observes the size of the cell (as compared to an erythrocyte), the shape of the nucleus, and the color of any granules.

BLOOD VALUES

Many labs/hospitals/clinics, etc.. Have their own normal values. For example normal range for hemoglobin in Florida is going to be different from normal values in Quito Ecuador whose altitude is 12000 feet! Their normal hemoglobin levels our higher than those living in Florida! What you need to understand is the basic range, these are not 100% exact numbers so don't get carried away with exact numbers for blood values.

Mature red blood cells do not contain all of the usual organelles. Red blood cells lack a nucleus and do not contain mitochondria, endoplasmic reticulum, or centrioles. Based on this information, which of the following is true?

Mature red blood cells do not divide and therefore have a short life span.

Neutrophils

Neutrophils are generally close to twice the size as an erythrocyte (average is around 12 μm). Because of the light staining of the granules, the nucleus is clearly seen. In a normal, healthy individual about half of the neutrophils will have three lobes. Decreased number of lobes (a shift to the left) means that the bone marrow is releasing these cells early. Increased number of lobes (hypersegmentation) is seen in megaloblastic or iron deficiency anemia. Neutrophils are phagocytes, and are the primary white blood cells that respond to bacterial infections. Because of their immune system function, neutrophils usually make up 50-70% of the white blood cells. In an infection, the bone marrow will respond by releasing an increased number of young neutrophils, resulting in neutrophilia (increased number of neutrophils and increased number of neutrophils with fewer or no lobes). Neutrophilia is also seen in acute inflammation (e.g., appendicitis, burns, heart attacks, etc.).

DIFFERENTIAL DIAGNOSIS

Quick concept: -A patient comes into the doctor, doctor orders blood work and just for example their hemoglobin comes back as below normal range. -The next step is to figure out what the cause is? Hopefully with your years of experience and medical education, you start to formulate ideas as to what caused your patients low hemoglobin. This is Differential Diagnosis: - It doesn't tell you what is wrong it merely tells you what the most likely option are for your patients' condition. - Sometimes its obvious other times more tests need to be run to get to the exact diagnosis, other times you can narrow it down to a couple of diagnoses and you do your best to pick the correct one! -The next couple of slides will show you the most common conditions that will cause high/low hemoglobin and hematocrit

PLATELETS

Nucleus: None Granules Stain Deep Purple Size: 2 to 4 micrometers Number: 150,000 to 400,000/mm3 Development: 4 to 5 Days Life Span: 5 to 10 Days Function: Seal small tears in blood vessels Blood clotting

LYMPHOCYTES

Nucleus: Spherical or indented Granules: None Size: 5 to 17 micrometers Number: 1500 to 3000/mm3 Development: Days to Weeks Life Span: Hours to Years Function: Mount immune response by direct cell attack or by antibodies

MONOCYTE

Nucleus: U or Kidney Shaped Granules: None Size: 14 to 24 micrometers Number: 100 to 700/mm3 Development: 2 to 3 Days Life Span: Months Function: Phagocytosis (Develop into macrophages in tissue)

PERICARDIAL LAYERS

Pericardium: -Fibro serous sac that encloses the heart and the roots of the great vessels -Layers of the Pericardium: Fibrous Serous Pericardial Cavity

Cell fragments involved in blood clotting and the repair of damaged blood vessels

Platelets

PLATELEtS

Platelets are fragments of a much larger cell called a megakaryocyte. Platelets are involved in the clot-ting process and form the initial platelet plug to stop bleeding. A platelet count that's lower than normal (thrombocytopenia) or higher than normal (thrombocytosis) is often a sign of an underlying medical condition, or it may be a side effect from medication. **Note: The above three cells were studied in API Lab and should be reviewed.*

ABO System

The ABO group is determined by the presence (or absence) of A and/or B antigens on the surface of the RBC. The presence of an antigen determines the ABO group (i.e., if there are A antigens on the red cell, then the individual is said to have A blood type). As you learned in General Biology, the ABO system is a codominant system, which means if both the A and B antigens are present, both are expressed

Rh System

The Rh system is another important blood group. Although there are more than fifty antigens in the sys-tem, the one people are most familiar with is the D antigen. The D antigen is a dominant antigen and if you possess at least one D antigen, the Rh type is said to be "positive." Rh negative individuals lack the D antigen. D negative individuals (or Rh negative) can develop anti-D if transfused with D positive blood. D negative blood can be given to both D positive and negative individuals. The Rh system does NOT have pre-formed antibodies.

CBC and Differential

The blood of almost all patients with major illnesses is examined because of the importance of determin-ing the presence of anemia and/or changes in leukocytes. Therefore, one of the most common blood tests done in the laboratory is the CBC (complete blood count) and DIFF (short for differential). The CBC is comprised of a WBC (white blood cell), RBC (red blood cell), HGB (hemoglobin), HCT (hemocrit), platelet count, three indicies (MCV, MCH, MCHC), and a white cell differential.

Jaundice is a symptom where the patient exhibits a yellow discoloration of the skin and eyes. What causes jaundice?

The liver is unable to process bilirubin, so its level in the blood rises.

Because of their short lifespan, a huge number of erythrocytes are recycled by macrophages. Which of the following is NOT a function of the macrophage?

The macrophage synthesizes a small amount of hemoglobin and transports it to the bone marrow.

Monocytes

The monocyte is the largest cell in normal blood. It is generally two to three times the size of the RBC. The nucleus of the monocyte may be any shape, but is frequently oval or round or indented to form a horseshoe shape. The nucleus stains less densely than other leukocytes and has a looser chromatin pat-tern than the lymphocyte. The cytoplasm is a blue-gray color and frequently contains tiny red to purple granules. These granules are not to be much smaller and finer than in the granulocytes and should not be confused. Because the monocyte is a phagocyte, the cytoplasm is frequently seen with vacuoles. Monocytes give rise to macrophages and dendritic cells in the immune system. Because of the monocytes' role in the immune system, increased numbers may be found in many condi-tions, including autoimmune diseases, gastrointestinal disorders, several viral and bacterial infections, post-splenectomy, inflammatory bowel disease, and some chronic infections such as tuberculosis, syphilis, and Rocky Mountain spotted fever. Autoimmune disorders such as lupus or rheumatoid arthritis might also lead to monocytosis. Normally monocytes will make up about 4-6% of the white blood cells **Mnemonic to Remember the Concentration of White Blood Cells From greatest to least concentration: • "Never Let Monkeys Eat Bananas" • Neutrophils, Lymphocytes, Monocytes, Eosinophils, Basophils

RBC

The normal red blood cell count differs between the sexes. For females, it is approximately 3.90-5.03 million cells/μL and 4.32-5.72 million cells/μL for males. In anemia, the red cell count is lower than normal. Anemia causes fatigue and weakness. Anemia has many causes, including low levels of iron, certain vitamins (e.g., B12), blood loss, or other underlying conditions, such as cancer. A red blood cell count that is higher than normal (erythrocytosis), or high hemoglobin or hematocrit levels, could point to an underlying medical condition, such as polycythemia vera or heart disease.

WBC

The normal white cell count is around 4,500-10,000/μL (normal blood cell ranges differ slightly by the instrument used and the population). The cells are then examined to determine the percentage of the various types of white blood cells. The total WBC count is determined by a blood cell analyzer and the different types of WBCs can be determined by the analyzer or a blood smear can be made and stained with Wright's stain. One hundred cells are counted and are divided (differentiated) into the five basic types of cells. If the white blood cell count is higher than normal (leukocytosis) it may indicate an infection or inflam-mation, an immune system disorder, or a bone marrow disease. The differential becomes especially helpful in determining the diagnosis. If the white blood cell count is lower than normal (leukopenia) it is associated with acute viral infections, chemotherapy treatment, and other infections.

Lymphocytes

The smallest of the leukocytes, the normal lymphocyte has a large, dark-staining nucleus with little pale blue cytoplasm. In normal situations, the coarse, dense nucleus of a lymphocyte is approximately the size of a red blood cell and is generally round in shape. There are three types of lymphocytes: natural killer cells, T cells, and B cells. Experienced hematologists can differentiate between the different types of lymphocytes, but students are not required to do so. Lymphocytosis is seen in infancy, exophthalmic goiter, mumps, rubella, infectious mononucleosis, sunburn, pertussis, and viral infections. Lymphopenia is associated with increased rates of infection after surgery or trauma as well as chemotherapy treatment. Normally lymphocytes will make up about 25 to 35% of the white blood cells.

diFFEREntiAl diAgnosis

This is as good a time as any to introduce this concept. If you have watched the popular TV show House you have already been introduced to this process. When House is discussing what unique condition the patient has and he is writing all the possibilities on the white board, that is differential diagnosis: all the possibilities you can use to explain the condition your patient is displaying. In this section you will be presented with two differential diagnosis lists, one for low and high hemoglobin and one for hematocrit.

PERiPhERAL sMEAR AnALysis

To perform a manual differential, a drop of blood is spread thinly onto a glass slide that is then stained. Wright's stain is a mixture of eosin and other dyes, including methylene blue. Eosin is an acidic red-orange dye and structures that take up that dye are referred to as eosinophilic. Structures that take up basic dyes (e.g., methylene blue) are referred to as basophilic. Structures that take up a combination of both dyes are called neutrophilic. The differential is performed by counting one hundred WBCs on the stained slide. In the past, a blood smear was prepared with every CBC. However, modern automated blood cell counting analyzers are able to perform most differentials. However, if the results from an automated cell count and/or the differential indicates the presence of abnormal white blood cells (WBCs), red blood cells (RBCs), and/or platelets or if there is reason to suspect that abnormal cells are present, then a manual blood smear will be made and read. A blood smear is used to categorize and/or identify conditions that affect one or more types of blood cells and to monitor individuals undergoing treatment for these conditions. There are many diseases, disorders, and deficiencies that can affect the number and type of blood cells produced, their function, and their lifespan. **In this lab, you will be required to identify the different types of white blood cells. For the practical exam, only mature WBCs will be chosen. Reading differentials takes time, so be sure to allow more than one lab session to review the blood smears

FUNCTIONS OF BLOOD

Transportation Protection Regulation

hEMogLoBin (hgB)

Unlike hematocrit, which is a relative measure of the RBCs (%), hemoglobin is a direct measure in (grams/ dL). The measure of hemoglobin under normal conditions is directly related to the oxygen-carrying capacity of the blood and therefore a measure of how well body tissues are being oxygenated. A commonality between hemoglobin and hematocrit is that they are significantly different between males and females. Male hemoglobin ranges from 13.8 to 18.0 g/dL and females 12.1 to 15.1 g/dL. We will not be performing a lab test for hemoglobin but you are expected to know its basic function/purpose, normal values, and what could cause abnormally high or low values

tRAnsFUsions—RECEiVing BLood

When blood is required, generally packed red blood cells are transfused. Packed RBCs are made by re-moving the plasma and platelets from donor blood. This reduces blood volume and the chance of adverse reactions to antibodies and substances that may be circulating in the donor's plasma. Because the plasma and platelets have been removed, just the antigen present on the RBC surface is transfused. This donor antigen CANNOT be the same as the circulating antibody of the recipient (receiver). For example, look-ing at the chart above, if the donor is an A, the donor blood could not be given to group B or O patients because of their circulating anti-A. Generally speaking, patients can receive their own blood group. If you notice in the chart on the following page, AB can receive from all blood groups (because there are no antibodies circulating in the recipient plasma that could react with donor antigens). This is the basis of the statement "AB is the universal recipient." However, one must understand that the statement only applies to the transfusion of RBCs. Plasma transfusions are the opposite!

The Blood Type

When referring to the blood type, both the ABO group and the Rh type are included. Thus, if both A and D antigen are found on the RBCs, the blood type is said to be "A positive" and that individual can receive A positive, A negative, O positive, and O negative blood or can donate to A positive and AB posi-tive individuals.

Granulocytes

Young granulocytes are called blasts. Blasts are large cells, with little cytoplasm. As the granulocyte matures, the size of the cell shrinks. The nucleus of the cell, which as a blast is generally round in shape, begins to pinch in to more of a kidney shape. The pinching continues until two lobes are formed with a connecting filament. (This is the process of segmentation.) The older the cell, the more lobes that will form. If one observes the nucleus, the blast has a fine texture, while the nucleus of the mature granulo-cyte will appear denser. As the cell matures the ratio of nucleus to cytoplasm changes and the amount of cytoplasm increases. Agranulocytes do NOT go through this particular maturation process, so if one observes a nucleus with lobes and connecting filaments, the cell is a granulocyte. As granulocytes mature, the granules develop and will be easily seen when dyed. Young granulocytes are called blasts. Blasts are large cells, with little cytoplasm. As the granulocyte matures, the size of the cell shrinks. The nucleus of the cell, which as a blast is generally round in shape, begins to pinch in to more of a kidney shape. The pinching continues until two lobes are formed with a connecting filament. (This is the process of segmentation.) The older the cell, the more lobes that will form. If one observes the nucleus, the blast has a fine texture, while the nucleus of the mature granulo-cyte will appear denser. As the cell matures the ratio of nucleus to cytoplasm changes and the amount of cytoplasm increases. Agranulocytes do NOT go through this particular maturation process, so if one observes a nucleus with lobes and connecting filaments, the cell is a granulocyte. As granulocytes mature, the granules develop and will be easily seen when dyed.