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Hematology - NANSLO Lab Activity

Page history last edited by Sue Schmidt 9 years, 3 months ago

NANSLO REMOTE LAB ACTIVITY

 

SUBJECT SEMESTER: XXXXXXXXXXXXXXXXXXXXX

TYPE OF LAB:  XXXXXXXXXXXXXXXXXXXX

 

 

 

 

 

 

TITLE OF LAB:  Hematology

 

Hematology NANSLO Lab Activity in Word Format last updated April 30, 2014.

 

Lab format: This lab is a remote lab activity.

 

Relationship to theory (if appropriate):  In this lab you will learn to identify the cellular components of blood and how perturbations in its structure or composition can cause physiological problems.  

 

Instructions for Instructors:  This protocol is written under an open source CC BY license. You may use the procedure as is or modify as necessary for your class.  Be sure to let your students know if they should complete optional exercises in this lab procedure as lab technicians will not know if you want your students to complete optional exercises.

 

Instructions for Students:  Read the complete laboratory procedure before coming to lab.  Under the experimental sections, complete all pre-lab materials before logging on to the remote lab.  Complete data collection sections during your online period, and answer questions in analysis sections after your online period.  Your instructor will let you know if you are required to complete any optional exercises in this lab.

 

Remote Resources:  Primary – Microscope, Secondary – Hematology slide set.

 

LEARNING OBJECTIVES:

 

After completing this laboratory experiment, you should be able to do the following things:

 

  1. Know the function of each formed element in the blood.
  2. Identify and distinguish between the different kinds of formed elements in the blood.
  3. Measure the relative size of certain cells.
  4. Identify, distinguish, and measure the size of healthy red blood cells and sickle-celled ones.
  5. Identify, distinguish, and measure the size of healthy red blood cells and Thalassemia majors ones.
  6. Identify, distinguish, and measure the size of healthy white blood cells and infectious mononucleosis ones.
  7. Explain the basis for blood typing and matching for blood donation and identify appropriate blood donors for a given recipient.

 

BACKGROUND INFORMATION:

 

Blood is the fluid that transports oxygen and nutrients to the cells and carries away carbon dioxide and other waste products. Technically, blood is a transport liquid pumped by the heart (or an equivalent structure) to all parts of the body after which it is returned to the heart to repeat the process.  Blood is the only tissue whose matrix is a complete fluid.  It is a tissue, because it is a collection of similar specialized cells that serve particular functions. These cells are suspended in a liquid matrix (plasma) which makes the blood a fluid.  If blood flow ceases, death will occur within minutes because of the effects of an unfavorable environment on highly susceptible cells.


Blood cells known as formed elements are suspended in the plasma.  Often clinical evaluations of the blood characteristics of the patient are important in assessing the condition of the patient.  Blood characteristics vary according to age, sex, metabolic condition, health conditions, and genetic variations.

 

Plasma serves as a transport medium for delivering nutrients to the cells of the body and for transporting waste products derived from cellular metabolism to the kidneys, liver, and lungs for excretion. It is also a transport system for blood cells, and it plays a critical role in maintaining normal blood pressure. Plasma helps to distribute heat throughout the body and to maintain homeostasis, or biological stability, including acid-base balance in the blood and body.


Plasma is derived when all the blood cells—red blood cells (erythrocytes), white blood cells (leukocytes), and platelets (thrombocytes)—are separated from whole blood. The remaining straw-colored fluid is 90–92 percent water, but it contains critical solutes necessary for sustaining health and life.  Important constituents include electrolytes such as sodium, potassium, chloride, bicarbonate, magnesium, and calcium.  In addition, there are trace amounts of other substances including amino acids, vitamins, organic acids, pigments, and enzymes.  Hormones such as insulin, corticosteroids, and thyroxin are secreted into the blood by the endocrine system.  Plasma concentrations of hormones must be carefully regulated for good health.

 

Figure 1:  Makeup of Blood
Figure 1: Blood is made up of multiple components, including red blood cells, white blood cells, platelets, and plasma.

Platelet: blood components". Art. Encyclopædia Britannica Online. Web. 27 Apr. 2014. http://www.britannica.com/EBchecked/media/113906/Blood-is-made-up-of-multiple-components-including-red-blood


 

Red blood cells, also called erythrocytes, are the cellular component of blood.  Millions of red blood cells in the circulation of vertebrates give the blood its characteristic color and carry oxygen from the lungs to the tissues. The mature human red blood cell is small, round, and biconcave.  It appears dumbbell-shaped in profile. The cell is flexible and assumes a bell shape as it passes through extremely small blood vessels.  It is covered with a membrane composed of lipids and proteins, lacks a nucleus, and contains hemoglobin — a red, iron-rich protein that binds oxygen.

 

White blood cell, also called leukocytes, are cellular components of the blood that lack hemoglobin, have nuclei, are capable of motility, and defend the body against infection and disease by ingesting foreign materials and cellular debris, by destroying infectious agents and cancer cells, or by producing antibodies (also called immunoglobulins, protective proteins produced by the immune system in response to the presence of a foreign substance, called an antigen. Antibodies recognize and latch onto antigens in order to remove them from the body.)   A healthy adult human has between 4,500 and 11,000 white blood cells per cubic millimeter of blood. Fluctuations in white cell number occur during the day.  Lower values are obtained during rest and higher values during exercise.  White cell count also may increase in response to convulsions, strong emotional reactions, pain, pregnancy, labor, and certain disease states, such as infections and intoxication.

 

Granulocytes, the most numerous of the white cells, rid the body of large pathogenic organisms such as protozoans or helminths and are also key mediators of allergy and other forms of inflammation. These cells contain many cytoplasmic granules, or secretory vesicles, that harbor potent chemicals important in immune responses. They also have multilobed nuclei, and, because of this, they are often called polymorphonuclear cells. On the basis of how their granules take up dye in the laboratory, granulocytes are subdivided into three categories: neutrophils, eosinophils, and basophils.


The most numerous of the granulocytes—making up 50 to 80 percent of all white cells—are neutrophils. They are often one of the first cell types to arrive at a site of infection where they engulf and destroy the infectious microorganisms through a process called phagocytosis.  Eosinophils and basophils typically arrive later.  The granules of basophils contain a number of chemicals, including histamine and leukotrienes, that are important in inducing allergic inflammatory responses.


Eosinophils destroy parasites and also help to modulate inflammatory responses.


Neutrophils are characterized histologically by their ability to be stained by neutral dyes and functionally by their role in mediating immune responses against infectious microorganisms.  Neutrophils move with amoeboid motion.


Neutrophils are actively phagocytic.  They engulf bacteria and other microorganisms and microscopic particles. The granules of the neutrophil are microscopic packets of potent enzymes capable of digesting many types of cellular materials.


An abnormally high number of neutrophils circulating in the blood is called neutrophilia. This condition is typically associated with acute inflammation, though it may result from chronic myelogenous leukemia, a cancer of the blood-forming tissues.  An abnormally low number of neutrophils is called neutropenia. This condition can be caused by various inherited disorders that affect the immune system as well as by a number of acquired diseases, including certain disorders that arise from exposure to harmful chemicals. Neutropenia significantly increases the risk of life-threatening bacterial infection.

 

Figure 2: Two neutrophils among many red blood cells.
Figure 2: Two neutrophils among many red blood cells. The normal functions of neutrophils are compromised in chronic granulomatous disease. "Chronic granulomatous disease". Photograph. Encyclopædia Britannica Online. Web. 27 Apr. 2014. http://www.britannica.com/EBchecked/media/126495/Two-neutrophils-among-many-red-blood-cells

 

Eosinophils are characterized histologically by their ability to be stained by acidic dyes (e.g., eosin) and functionally by their role in mediating certain types of allergic reactions.  Eosinophils contain large granules, and the nucleus exists as two non-segmented lobes.

 

Figure 3: White blood cells in a field of red cells.
Figure 3: White blood cells in a field of red cells. (Top left) Monocyte, (top centre) basophil, (top right) platelets, (bottom left) two neutrophils, (bottom right) lymphocyte and eosinophil, respectively.
A. Owczarzak/Taurus Photos, Inc., "Eosinophil: white blood cells in field of red cells". Photograph. Encyclopædia Britannica Online. Web. 27 Apr. 2014. http://www.britannica.com/EBchecked/media/767/White-blood-cells-in-a-field-of-red-cells-Monocyte

 

 

Basophils are characterized histologically by its ability to be stained by basic dyes and functionally by its role in mediating hypersensitivity reactions of the immune system.  Basophils are the least numerous of the granulocytes and account for less than one percent of all white blood cells occurring in the human body.


Agranulocytes do not contain any cytoplasmic granules or secretory vesicles.  We distinguish two types: lymphocytes and monocytes.


Lymphocytes, which are further divided into B and T cells, are responsible for the specific recognition of foreign agents and their subsequent removal from the host.

    • B lymphocytes secrete antibodies.
    • Typically, T cells recognize virally infected or cancerous cells and destroy them, or they serve as helper cells to assist the production of antibody by B cells.

 

Monocytes move from the blood to sites of infection where they differentiate further into macrophages.  These cells are scavengers that phagocytose whole or killed microorganisms and are therefore effective at direct destruction of pathogens and cleanup of cellular debris from sites of infection. Neutrophils and macrophages are the main phagocytic cells of the body, but macrophages are much larger and longer-lived than neutrophils.


Platelets also called thrombocytes, are colorless, non-nucleated blood component that are important in the formation of blood clots (coagulation). Platelets are found only in the blood of mammals.


Platelets are formed when cytoplasmic fragments of megakaryocytes, which are very large cells in the bone marrow, pinch off into the circulation as they age. They are stored in the spleen. Some evidence suggests platelets may also be produced or stored in the lungs where megakaryocytes are frequently found.


Platelets play an important role in the formation of a blood clot by aggregating to block a cut blood vessel and provide a surface on which strands of fibrin form an organized clot by contracting to pull the fibrin strands together to make the clot firm and permanent and, perhaps most important, by providing or mediating a series of clotting factors necessary to the formation of the clot. Platelets also store and transport several chemicals, including serotonin, epinephrine, histamine, and thromboxane.   Upon activation, these molecules are released and initiate local blood vessel constriction which facilitates clot formation.

 

Figure 4:  A micrograph of a round aggregation of platelets Figure 4:  A micrograph of a round aggregation of platelets (magnified 1000x). 
Dr. F. Gilbert/Centers for Disease Control and Prevention (CDC) (Image Number: 6645)
"Platelet". Photograph. Encyclopædia Britannica Online. Web. 27 Apr. 2014. http://www.britannica.com/EBchecked/media/116860/A-micrograph-of-a-round-aggregation-of-platelets

 

 

Blood is a germ-free or axenic fluid; however, it is susceptible to infections by eukaryotic parasites, bacteria or viruses, and some of these infections range from mild to fatal. For example:


Thalassemia is group of blood disorders characterized by a deficiency of hemoglobin, the blood protein that transports oxygen to the tissues. Thalassemia (Greek: “sea blood”) is so called because it was first discovered among peoples around the Mediterranean Sea among whom its incidence is high. 

In the mild form of the disease, Thalassemia minor, there is usually only slight or no anemia and life expectancy is normal.  Occasionally, complications occur involving slight enlargement of the spleen.

 

Thalassemia major is characterized by severe anemia, enlargement of the spleen, and body deformities associated with expansion of the bone marrow.

Mononucleosis is a viral infection causing fever, sore throat, and swollen lymph glands, especially in the neck.  Mononucleosis, or mono, is often spread by saliva and close contact.  It is known as "the kissing disease" and occurs most often in those ages 15 to 17. However, the infection may develop at any age.  Mono is usually linked to the Epstein-Barr virus (EBV), but can also be caused by other organisms such as cytomegalovirus (CMV).  Mono may begin slowly with fatigue, a general ill feeling, headache, and sore throat. The sore throat slowly gets worse. Your tonsils become swollen and develop a whitish-yellow covering. The lymph nodes in the neck are frequently swollen and painful.

References:

 

  1. "blood." Encyclopedia Britannica. David H. Yawn, M.D.  Last updated 6/27/2013.  http://www.britannica.com/EBchecked/topic/463483/plasma 
  2. "plasma."  Encyclopedia Brintannica.  David H. Yawn, M.D. Last updated 6/27/2013.  http://www.britannica.com/EBchecked/topic/463483/plasma
  3. MedlinePlus, US Library of Medicine. http://www.nlm.nih.gov/medlineplus/ency/article/000591.htm
  4. "thalassemia." Encyclopedia Britannica. The Editor of the Encyclopedia Britannica. Last.  Updated 10/27/2013.  http://www.britannica.com/EBchecked/topic/589769/thalassemia

 

 

EQUIPMENT:

  • Paper
  • Pencil/pen
  • Slides
    • Human blood smear
    • Human sickle cell anemia
    • Thalassemia majors smear
    • Infectious mononucleosis
  • Computer with Internet access (for the remote laboratory and for data analysis)

 

EXPERIMENTAL PROCEDURE:

 

Once you have logged on to the remote lab system, you will perform the following laboratory procedures.  See Preparing for the Microscope NANSLO Lab Activity below.

 

PRE-LAB EXERCISE 1:  Microscopic Examination of the Formed Elements of Blood

 

In this exercise you will:

 

  • Use the microscope to distinguish between the different types of formed elements in blood.
  • Identify the red blood cells from different white blood on images you have taken.
  • Measure and compare the relative size of red blood cells and the monocytes.
  • Compare the nucleus shape of the different white blood cells.

 

Pre-Lab Questions:

  1.  Using what you know about the formed elements in blood, do you think the size and shape of the red blood cells will be bigger, smaller, or the same as the ones of the monocytes?
  2. Rewrite your answer to question #1 in the form of an If . . . Then . . . hypothesis.

 

EXERCISE 1:  Microscopic Examination of the Formed Elements of Blood

 

Data Collection:

  1. Select the human blood smear slide (Slide Cassette 3: #21) from the slide loader.  Using the 10X objective, identify the blood cells and bring them in to focus.
  2. Carefully working your way through all the objectives focusing with each one until you reach the 60X objective, capture an image.  Insert your images of normal red blood cells, neutrophils, eosinophils, basophils, lymphocytes and monocytes below.
  3. Label the three different parts of a cell (nucleus, cytoplasm and cell membrane) and the granules of the white blood cells.
  4. Describe the shape of the red blood cells and the different white blood cells and their nuclei.
  5. Were you able to see the nucleus of the red blood cells? Explain your answer.
  6. Next we are going to measure the size of the red blood cells and the monocytes.  To determine the size of the cells, we are going to use the ratio method.  In order to do this, you will need one piece of information which is the width of your field of view.  On our microscopes, the field of view is 205µm.
  7. Now if we use the image in figure 1 we can see that the total width of the field of view is 13.6 cm or 136 mm (Image A).  The cell (Gray) is 3.7 cm or 37mm (Image B).


    Figure 5:  Measurements

  8. Now if we divide 37mm/136mm = 0.272 which we multiply the total length of the field of view by so 0.272 * 205µm = 55.77 µm rounded for significant figures gives us a cell size of 56µm.
  9. Measure the difference in the size of the red blood cells and compare that to the size of the monocytes.
  10. Are your results in correlation with what you have predicted earlier?
  11. Rewrite your hypothesis in light of our new information you collected in this exercise.


Analysis:

 

  1. Describe the function of each cell type in the table blow.

    Cell Type
    Function
    Erythrocytes
     
    Neutrophils
     
    Eosinophils
     
    Basophils
     
    Monocytes
     
    Platelets
     

     

 

PRE-LAB EXERCISE 2:  Normal Red Blood Cells and Sickled Cells Slide Observations

 

In this lab exercise you will measure and compare the relative size and shape of normal red blood cells with the sickled celled ones.

Pre-lab 2 Questions:

  1. Do you predict the size of normal red blood cells to be smaller, bigger, or the same as the sickled celled ones?
  2. Rewrite your answer to question one in the form of an If … Than … hypothesis.

 

EXERCISE 2:  Normal Red Blood Cells and Sickled Cells Slide Observations

 

Data Collection:

 

  1. Select the Sickle Cell Anemia slide (Slide Cassette 3: #22)  from the slide loader. Using the 10X objective, identify the blood cells and bring them in to focus.
  2. Carefully working your way through all the objectives focusing with each one until you reach the 60X objective, capture an image.  Insert your picture of sickled cells below.

 

Analysis:

 

  1. Utilizing your normal blood image form exercise 1 and the method from exercise 1 determine the length of the normal and sickle red blood cells.
  2. Based on your observation, describe the difference between the shape of normal red blood cells versus sickled cells.
  3. Are your results in correlation with what you have predicted earlier?
  4. Rewrite your hypothesis to take into account the new information you have learned in this exercise.
  5. What is the impact of sickle cell anemia on oxygen transport?

PRE-LAB EXERCISE 3:  Normal Red Blood Cells and Thalassemia majors Slide Observations

 

In this lab exercise you will measure and compare the relative size and shape of normal red blood cells with Thalassemia major ones.

Pre-lab 3 Questions:

  1. Do you predict the size of normal red blood cells to be smaller, bigger or the same as Thalassemia majors ones?
  2. Rewrite your answer to question one in the form of an If … Than … hypothesis.

 

EXERCISE 3:  Normal Red Blood Cells and Thalassemia majors Slide Observations

 

Data Collection:

 

  1. Select the Thalassemia majors slide (Slide Cassette 3: #32)  from the slide loader.  Using the 10X objective, identify the blood cells and bring them in to focus.
  2. Carefully working your way through all the objectives focusing with each one until you reach the 60X objective, capture an image.  Insert your picture of Thalassemia major cells below.

 

Analysis:

 

  1. Utilizing your normal blood image form exercise 1 and the method from exercise 1 to determine the diameter of the normal red blood cells and the Thalassemia majors ones.
  2. Based on your observation, describe the difference between the size of normal red blood cells versus the Thalassemia majors cells.
  3. Are your results in correlation with what you have predicted earlier?
  4. Rewrite your hypothesis to take into account the new information you have learned in this exercise.
  5. What is the impact of large pale red blood cells on the blood circulation and the overall body health?  Explain your answer.

 

PRE-LAB EXERCISE 4:  Normal and Infectious Mononucleosis Blood Observations

 

In this lab exercise you will measure and compare the relative size and shape of normal white blood cells with infectious mononucleosis ones.

Pre-lab 4 Questions:

  1. Do you predict the size of normal white blood cells to be smaller, bigger or the same as the infectious monocudeosis ones?
  2. Rewrite your answer to question one in the form of an If … Than … hypothesis.

 

EXERCISE 4:  Normal and Infectious Mononucleosis Blood Observations

 

Data Collection:

 

  1. Select the infectious mononucleosis slide (Slide Cassette 3: #25)  from the slide loader. Using the 10X objective, identify the blood cells and bring them in to focus.
  2. Carefully working your way through all the objectives focusing with each one until you reach the 60X objective, capture an image.  Insert your picture of infected white blood cells below.

 

Analysis:

 

  1. Utilizing your normal blood image form exercise 1 and the method from exercise 1, determine the diameter of the normal white blood cells and the infectious mononucleosis ones.
  2. Based on your observation, describe the difference between the size of these cells.
  3. Are your results in correlation with what you have predicted earlier?
  4. Rewrite your hypothesis to take into account the new information you have learned in this exercise.
  5. What is the impact of the infectious mononucleosis on the overall function of the immune system?  Explain your answer.

 

EXERCISE 5:  ABO-RH Blood Typing

 

Blood typing is the classification of blood in terms of distinctive inherited characteristics that are associated with the antigens located on the surface of red blood cells (erythrocytes). The ABO and the Rh blood groups are among those most commonly considered.  Identification of these determinants has become indispensable in connection with blood transfusion, because the recipient and donor must have the same, or compatible, blood groups. Otherwise, hemolysis (destruction) or coagulation (clotting) results from interaction of an antigen on the red blood cells of one with an antibody in the serum of the other.


ABO blood group system is the classification of human blood based on the inherited properties of red blood cells (erythrocytes) as determined by the presence or absence of the antigens A and B which are carried on the surface of the red cells. Persons may thus have type A, type B, type O, or type AB blood. The A, B, and O blood groups were first identified by Austrian immunologist Karl Landsteiner in 1901.

 

Blood containing red cells with type A antigen on their surface has in its serum (fluid) antibodies against type B red cells. If, in transfusion, type B blood is injected into persons with type A blood, the red cells in the injected blood will be destroyed by the antibodies in the recipient’s blood. In the same way, type A red cells will be destroyed by anti-A antibodies in type B blood. Type O blood can be injected into persons with type A, B, or O blood unless there is incompatibility with respect to some other blood group system also present.


To perform a blood typing test, anti-A and anti-B sera are each separately mixed with a drop of sample blood and observed for agglutination (or clumping).

 

Rh blood group system is a system for classifying blood groups according to the presence or absence of the Rh antigen, often called the Rh factor, on the cell membranes of the red blood cells (erythrocytes). The designation Rh is derived from the use of the blood of rhesus monkeys in the basic test for determining the presence of the Rh antigen in human blood. The Rh blood group system was discovered in 1940 by Karl Landsteiner and A.S. Weiner. Since that time, a number of distinct Rh antigens have been identified, but the first and most common one, called RhD, causes the most severe immune reaction and is the primary determinant of the Rh trait.


The Rh antigen poses a danger for the Rh-negative person, who lacks the antigen, if Rh-positive blood is given in transfusion. Adverse effects may not occur the first time Rh-incompatible blood is given, but the immune system responds to the foreign Rh antigen by producing anti-Rh antibodies. If Rh-positive blood is given again after the antibodies form, they will attack the foreign red blood cells, causing them to clump together, or agglutinate. The resulting hemolysis, or destruction of the red blood cells, causes serious illness and sometimes death.

 

A similar hazard exists during pregnancy for the Rh-positive offspring of Rh-incompatible parents when the mother is Rh-negative and the father is Rh-positive. The first child of such parents is usually in no danger unless the mother has acquired anti-Rh antibodies by virtue of incompatible blood transfusion. During labor, however, a small amount of the fetus’s blood may enter the mother’s bloodstream. The mother will then produce anti-Rh antibodies which will attack any Rh-incompatible fetus in subsequent pregnancies. This process produces erythroblastosis fetalis, or hemolytic disease of the newborn, which can be fatal to the fetus or to the infant shortly after birth. Treatment of erythroblastosis fetalis usually entails one or more exchange transfusions. The disease can be avoided by vaccinating the mother with Rh immunoglobulin after delivery of her firstborn if there is Rh-incompatibility. The Rh vaccine destroys any fetal blood cells before the mother’s immune system can develop antibodies.


Reference: http://www.britannica.com/EBchecked/topic/69880/blood-typing

In the table below, predict the agglutination reactions for each of the following blood types.

 

Blood Type

Agglutination Reaction (Clumping – yes or no)

Anti-A Serum

Anti-B Serum

Anti-Rh Serum

A +

 

 

 

A –

 

 

 

B +

 

 

 

B –

 

 

 

AB +

 

 

 

AB –

 

 

 

O +

 

 

 

O –

 

 

 

 

SUMMARY QUESTIONS:

 

Answer the following questions.

 

  1. Which of the formed elements are referred to collectively as granulocytes?
  2. Which of the formed elements are referred to collectively as agranulocytes?
  3. When a patient is admitted to the hospital, one of the first procedures that health professionals perform is a blood draw. One lab value that is checked is a white blood cell (WBC) count. If the WBC count is elevated, this could indicate the presence of inflammation. Further analysis (called a differential) is performed to determine the relative prevalence of different types of white blood cells and the potential cause of the inflammation. What might be causing the inflammation if (use your textbook for reference:)
    1. Neutrophils were elevated?  Explain your reasoning.
    2. Lymphocytes were elevated?  Explain your reasoning.
    3. Eosinophils were elevated?  Explain your reasoning.
  4. If someone is diagnosed with "microcytic, hypochronic anemia," how would you describe the hemoglobin content and size of his or her red blood cells?
  5. Iron is necessary to produce hemoglobin.  In iron-deficiency anemia, would you expect the red blood cells to be hyperchronic, normochronic, or hypochronic?  Explain.
  6. What is the difference between antigen and antibody?
  7. If your blood type is A-, which antigens are present on your red blood cells?  What if your blood type is AB+?
  8. With the ABO blood types, individuals will have circulating antibodies to antigens that are not present in their own blood.  Identify the antibodies (A, B, both A and B, or none) produced by the following individuals:
    1. An individual with type A blood:
    2. An individual with type B blood:
    3. An individual with type O blood:
    4. An individual with type AB blood:
  9. Based on your results for each patient, identify the blood types he/she could receive using the following choices.  Note:  Some patients may be able to receive several types of blood.

    A+
    B+
    AB+
    O+
    A
    B
    AB
    O


    Patient 1 could receive _____________________________
    Patient 2 could receive _____________________________
    Patient 3 could receive _____________________________

    Patient 4 could receive _____________________________

  10. What is a universal donor?  Which of these patients would be considered universal donor?
  11. What would happen to a type O patient if he receives type A or B blood?

 

PREPARING FOR THIS NANSLO LAB ACTIVITY:

 

Read and understand the information below before you proceed with the lab!

  

Scheduling an Appointment Using the NANSLO Scheduling System

 

Your instructor has reserved a block of time through the NANSLO Scheduling System for you to complete this activity. For more information on how to set up a time to access this NANSLO lab activity, see www.wiche.edu/nanslo/scheduling-software.  

 

Students Accessing a NANSLO Lab Activity for the First Time

 

You must install software on your computer before accessing a NANSLO lab activity for the first time. Use this link to access instructions on how to install this software based on the NANSLO lab listed below that you will use to access your lab activity – www.wiche.edu/nanslo/lab-tutorials.

 

  1. NANSLO Colorado Node -- all Colorado colleges.
  2. NANSLO Montana Node -- Great Falls College Montana State University, Flathead Valley Community College, Lake Area Technical Institute, and Laramie County Community College.
  3. NANSLO British Columbia Node -- Kodiak College.

 

Using the Microscope for a NANSLO Remote Web-based Science Lab Activity

 

We've provided you with three ways to learn how to use the microscope for this NANSLO lab activity:

 

  1. Read these instructions.
  2. Watch this short video. 
  3. Print off these instructions to read (PDF version of the instructions.)

 



NOTE:   The conference number in this video tutorial is an example.  See “Communicating with Your Lab Partners” below to determine the toll free number and pin to use for your NANSLO lab activity.

 

MICROSCOPE RWSL LAB INTERFACE INSTRUCTIONS

 

The Remote Web-based Science Lab (RWSL) microscope is a high quality digital microscope located at the NANSLO Node.  Using a web interface as shown below, you can control every function of the microscope just as if you were sitting in front of it. 

 

The equipment control software shown below is written using the LabVIEW software from National Instruments. The user interface is presented as a LabVIEW control panel which will be referred to as the lab interface for the remainder of the document.

Figure 1:  RWSL Microscope Lab Interface
Figure 1:  Remote Web-based Science Lab (RWSL) Microscope Lab Interface

 

COMMUNICATING WITH YOUR LAB PARTNERS

 

As soon as you have accessed this lab interface, call into the toll free conference number shown on the control panel to communicate with your lab partners and with the Lab Technicians.  Use the PIN code noted to join your lab partners.  Only one person can be in control of the equipment at any one time so talking together on a conference line helps to coordinate control of the equipment and creates a more collaborative environment for you and your lab partners.

 

GAINING CONTROL OF THE MICROSCOPE

 

Right click anywhere in the grey area of the lab interface and choose “Request Control of VI” from the dialogue box that appears when multiple students are using the microscope at the same time,.  After you request control, you may have to wait a short time before you actually receive control and are able to use the features on this lab interface.

 

Figure 2:  Right click anywhere in the grey area to "Request Control of VI" 

Figure 2:  Selecting "Request Control of VI"

 

RELEASING CONTROL OF THE MICROSCOPE

 

To release control of the microscope so that another student can use it, right click anywhere in the grey area of the lab interface and choose "Release Control of VI" from the dialogue box that appears. 

 

 

Figure 3:  Right click anywhere in the grey area and select "Release Control of VI" to release control of the lab interface

Figure 3:  Selecting "Release Control of VI" 

 

MICROSCOPE CONTROLS

 

 The Stage Controls allow you to adjust the visual of the specimen that has been placed on the stage of the microscope, select lenses with various magnifications, and select whether or not the condenser lens is in the light beam.  Below are more specific instructions on using these controls.  When using the arrows on this lab interface, click and hold the arrow until the desired effect is achieved or click and wait to view the result before clicking again. Quick clicks on the arrows may cause the system to lock up.

 

Figure 4:  Microscope controls including stage controls, objectives and condenser

Figure 4:  Microscope Controls - Stage, Objective & Condenser

 

Stage Controls: Using the left and right and up and down arrows found to the right of the microscope image in the Stage Control area, moves the microscope stage which holds the specimen.  These arrows allow you to precisely control the position of the specimen on the stage. 

 

  1. Use the "Right" and "Left" arrows to move the Stage so that you can view the specimen from left to right.
  2. Use the "Backward" and "Forward" arrows to move the Stage so that you can view the top, middle or bottom of the specimen.
  3. Use the "Up" and "Down" arrows to move the stage closer or farther away from the objective lens to bring a specimen into focus.   BE CAREFUL!  Don't move the stage too close to the lens.

 

When selecting the button between the "Up" and "Down" arrows, you can toggle between “Coarse” and “Fine” focus.  When the button is dark green and “Coarse/Fine” is displayed to the right of the button, the microscope is in “Coarse” focus.  When the button is bright green and “Fine” is displayed, the microscope is in “Fine” focus.   Typically, you will start with coarse focus which moves the stage in large increments and then use fine focus to complete your final focusing as it moves the stage in smaller increments.  There is no difference between the course and fine focus when using the 60X objective

 

NOTE:  When you click on these arrows, the specimen appears to move in the opposite direction.  Since the objective stays fixed, the image moves in the opposite direction of the stage.  This is how these controls work on most microscopes so the "feel" of the microscope is preserved over the web.

 

 

Figure 5:  Right/Left & Backward/Forward Stage Controls

    Figure 5:  Right/Left & Backward/Forward Stage Controls

    Figure 6:   Up/Down Stage Control & Coarse/Fine Focus Control       Figure 6:  Up/Down Stage Controls &
      Coarse/Fine Focus Control

                        

Objective:  A microscope mounts an objective lens very close to the object to be viewed.  Depending on need, different lenses with different power will be used on the microscope. This microscope feature multiple objectives, each with different power, mounted on a rotating turret.  The larger the magnification numbers the greater the magnification.  For example, if a specimen is viewed through a 40X objective lens, the magnifier in that lens displays the specimen 40 times larger than an equivalent view as seen by the unaided eye.  Remember that the ocular or other lenses also add to the magnification.

 

This microscope has five lenses – 4X, 10X, 20X, 40X, and 60X.  Use the arrows below the objective lens box that indicates the magnification of the current objective lens to move to a higher or lower magnification lens.  If you have activated the “Picture-in-Picture” Preset 2 (see below) you will be able to see the objective lens move when you select a new magnification.

 

Condenser:  The condenser controls whether or not the condenser lens is in the light beam. You want to have the condenser OUT for the 4x objective but IN for all the others.

 

SELECTING A CASSETTE AND LOADING SLIDES ONTO THE STAGE

 

There are two tabs on the lab interface.  When you first access the lab interface, the "Microscope" tab is displayed by default.  Click on the Slide Loader tab at the top of the screen to access the controls for the Slide Loader robot.  There can be up to four cassettes available on the Slide Loader.  These cassettes are used to store slides, and each can hold up to 50 slides.  The cassettes available to you are dependent on the lab activity to be completed.  Once a cassette has been selected, you will use the drop-down list to select your slides.

 

Figure 7:  Slide Loader Tab
Figure 7:  Select the Slide Loader Tab to select a cassette and slides.

 

 

EXAMPLE OF HOW TO LOAD SLIDES

 

In this example, we have selected Cassette #1. Using the drop-down menu, we have selected "1:  Colored Threads Whole Mount."  Then, we selected the "Load" button.  A message indicates that the slide is loading.  Using the picture-in-picture camera, you can watch this happening.  The robotics selects the slide and places it on the microscope stage.

 

Figure 8:  Selecting 1: Colored Threads While Mount Slide
Figure 8:  Selecting the slide
"1: Colored Threads Whole Mount" from Cassette #1

 

Notice that when a slide is actually on the microscope (or when it is being loaded or unloaded), the cassette controls are greyed out so you cannot load a second slide until the first is removed.  Once the slide is on the microscope stage, it will be listed in the "Current Slide on Stage" box.  The only thing that the Slide Loader robot can do is return it to the cassette when the "Return Slide to Cassette" button is selected. 

 

Figure 9:  Slide is loaded and Slide Loader Slide Selector is greyed out
Figure 9:  "LOADING SLIDE ... PLEASE WAIT" is displayed
in the "Current Slide on Stage" window

 

Select the "Microscope" tab to perform the NANSLO lab activity.  Once you are finished with the slide, select the "Slide Loader" tab and select "Return Slide to Cassette" button.  Once the slide is returned to the cassette, the Slide Loader controls are again available to select another slide from the cassette.

 

ENHANCING THE MICROSCOPE IMAGE

 

The digital camera mounted on the microscope has a camera control unit that is equipped with a series of image processing functions that enable you to quickly and easily correct imaging problems that arise from low or high contrast, poor focus, insufficient or uneven illumination, sample shading or discoloration and noise.  The most common reason for uneven elimination is a light source that does not completely fill the field of view on lower magnifications.  The White Balance should be used only if the image appears to be brown or gray, and you think you might need to adjust it (although it won't hurt anything to click this button).


A choice of color modes can be selected in the Microscope Image area and are used to display the image in different color palettes in order to highlight certain features.  The default setting is "Normal."

 

Figure 10:  Microscope Image Effects

 

Figure 10:  Microscope Image Special Effects and Other Image Controls for Camera

 

Here is a description of each option:

  1. In the “Normal” mode, the sample is displayed in its true  colors.
  2. In the “Negative” mode, the sample is displayed in a color-inverted form, where red, green, and blue values are converted into their complementary colors. The technique is useful in situations when color inversion can be of benefit in exposing subtle details or in quantitative analysis of samples.
  3. In the “Blue Black” mode, the black portions of a grayscale negative sample are displayed in blue. This mode is often useful to reveal details in samples having a high degree of contrast.  The “Blue Black” filter can aid you in examining a wide spectrum of difficult samples.
  4. In the “Black & White” mode, a grayscale image of the sample is displayed.
  5. In the “Sepia” mode, a brown scale (black and white) image of the sample is displayed. Although typically this filter is of little utility, it can be employed to alter image color characteristics to improve the visualization of sample detail.
  6. At times, the sample may have an unacceptable color quality.  Use “White Balance” calibration to remove the color cast.  This process is often referred to as white balancing.
  7. Auto Exposure is on automatically.  You do not need to do anything with Auto Exposure unless you are adjusting the luminance.  If you are doing so, you should turn off Auto Exposure by clicking on the button.  The green light is now off.  Now adjust the luminance.  See explanation below.

 

Reference:  http://www.microscopyu.com/articles/digitalimaging/dn100/correctingimages.html

 

Auto Exposure is normally turned on, but you can turn it off if you want to play around with the brightness of the light source and not have the microscope camera automatically adjust it.  It is usually best, though, to leave it turned on. 

 

When you turn off the Auto Exposure, the button turns dark green.  Some new controls appear that let you turn the LED off or on, and also adjust the intensity of the light source.  The intensity of the light source can be increased or decreased manually with the dial that now appears next to the Objective control when Auto Exposure is turned off.

 

Figure 11:  When auto exposure is turned off, LED controls are available

Figure 11:  Additional controls available when Auto Exposure is turned off

 

 

CAPTURING AND SAVING A MICROSCOPE IMAGE

 

When the “Capture Image” button is pressed, a high-resolution image of what is currently in the field of view of the objective is captured.  While the image is being captured, the button will be illuminated bright green.  The capture is complete when the light turns off.  Be patient as this may take several seconds to complete.


 After the Capture Image light turns off, select the “View Captured Image” tab on the bottom of this control panel to view the image.

 

Figure 12:  Select the Capture Image button and then select the View Captured Image Tab

Figure 12:  Click the capture image button (#1), wait till the green light goes off,
and then select the View Captured Image tab (#2)

 

After opening this image through the View Captured Image tab, you will need to take a snapshot of it and save it to your computer.  There are several ways to do this, depending on your operating system.


WINDOWS:

  1.  Pressing the two keys ALT and Print Screen simultaneously will copy the active window into your computer clipboard.  Then you can past it into a document.
  2. Windows 7 and above has a Snipping Tool program under Programs/Accessories which can capture selected areas of the screen. 
  3. Right click on it and select "Copy" from the menu presented.  After right clicking and selecting Copy, just open a document and right click and select Paste.  You can either paste it directly into your lab report document or into another one for safe keeping until you use it later.  You can use drawing tools in your word processing editor to annotate this image so you can show your instructor that you know what you were suppose to be looking for!

 

Figure 13:  Right click and then select copy to paste the image into a document

Figure 13:  Right click and select Copy to paste the image into a document

 

MAC:

  1. Press these three keys simultaneously MAC Keys to capture image.  This will change your cursor icon into a little cross. 
  2. Now press the spacebar, and the icon becomes a camera.  Click in the image window you want to take a snapshot of, and it will save the image to a file on your desktop.

 

There are lots of free screenshot utilities you can also use to capture this image.

 

If you are familiar with saving a document to your computer, you also can select “Save Image As” from the pop-up menu,  give the image a name and then select a location on your computer where you want this image to be saved for future use. 

 

MICROSCOPE IMAGE VIEW WINDOW

 

The Image View Window displays the real-time video feed from the digital camera “looking through” the microscope. 

 

Figure 14:  Image View Window
Figure 14:  Image View Window

 

PICTURE-IN-PICTURE CONTROLS - CAMERA PRESET POSITIONS AND PAN-TILT-ZOOM CONTROLS

 

When you click on the "Picture-in-Picture" button, it turns bright green.  A second real-time video feed from another digital camera appears in the Image View Window.  The controls shown in Figure 15 are all operational when the Picture-in-Picture feature is selected. 

 

Figure 15:  Picture-in-Picture Image Controls

Figure 15:  Picture-in-Picture Image Controls

 

CAMERA PRESETS

 

There are six camera preset positions.

 

Figure 16:  Picture-in-picture Preset 1 and 6Figure 16:  Picture-in-picture Camera Preset 1 and 6 - Displays the microscope, microscope camera,
and a camera control unit projecting the sample on the Stage. 

 

 

 

 

 

 

 

 

 

 

Figure 17: Picture-in-picture Camera Preset 2Figure 17:  Picture-in-picture Camera Preset 2:  Displays a closeup of the objective lens.

 

 

 

 

 

 

 

 

 

 

 

Figure 18: Picture-in-picture Camera Preset 3Figure 18:  Picture-in-picture Camera Preset 3 - Displays a closeup of the camera control unit
projecting the sample on the Stage.

 

 

 

 

 

 

 

 

 

 

Figure 19:  Picture-in-picture Camera Present 4 

Figure 19:  Picture-in-picture Camera Preset 4 - Displays the microscope eye piece and
the camera mounted to the microscope.

 

 

 

 

 

 

 

 

 

 

Figure 20: Picture-in-picture Camera Present 5Figure 20:  Picture-in-picture Camera Preset 5 - Displays the Condenser Lens underneath the Stage that focuses the light on the sample.  The Condenser Lens controls the width of the beam.  In some instances you will want a tighter beam while in other cases you will want a broader beam to control the image quality.  This setting has been optimized for you.

 

 

 

 

 

 

 

 

PAN, TILT, ZOOM CONTROLS FOR PICTURE-IN-PICTURE

 

For each camera preset view, additional camera options are available. 

  1. Use the up and down arrows to tilt the camera up or down. 
  2. Use the right and left arrows to pan right or left.
  3. Use the left "Zoom OUT" arrow and right "Zoom IN" arrow to zoom out and in.


Figure 21:  Picture-in-picture Zoom Capability
Figure 21:  Picture-in-picture Camera - Example of "Zoom In" capability

 

 

For more information about NANSLO, visit www.wiche.edu/nanslo.

 

 

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Creative Commons Attribution 3.0 United States License 3  

 

 

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