General Objectives

The student shall develop the following:

1. An understanding of the test methods used in the transfusion lab and their uses.
2. Knowledge and understanding of the quality assurance needed to control test methods in order to ensure a high level of patient safety.

Specific Objectives

The student shall be able to do the following:

Antibody Detection

1. For saline-phase antibody detection methods:

   1. discuss the importance of serum/cell ratios.
   2. discuss the relevance of test temperatures (4°C, RT, 37°C) to detection of clinically significant antibodies.
   3. state the immunoglobulin class(es) of antibodies detected and provide a list of antibody specificities that react in saline tests.
   4. state the concentration, molarity, and pH of saline used in serological tests.
   5. state the molarity of LISS and discuss its mechanism of action, advantages, and disadvantages.

2. For enzyme-phase antibody detection methods:

   1. list four enzymes used in blood banking and state their sources and mechanisms of action (two).
   2. list four uses of enzymes in the blood bank.
   3. differentiate between one-stage and two-stage enzyme methods, including their advantages and disadvantages.
   4. state the immunoglobulin class(es) of antibodies detected and provide a list of antibody specificities detected by enzyme tests.
   5. name seven antibodies usually NOT detected by enzymes and explain why.
   6. explain the main reason why many blood banks no longer use enzyme phases routinely.
   7. describe the quality control required to control that enzyme-treated cells have not been over- or under-treated.

3. For albumin-phase antibody detection methods:

   1. discuss the mechanism of action of bovine albumin in facilitating agglutination by IgG antibodies.
   2. state the immunoglobulin class(es) of antibodies detected in albumin phases and provide a list of antibody specificities detected.
   3. discuss the effectiveness of albumin phases (compared to other phases) in detecting antibodies.
   4. describe the method used to perform the albumin phase.

4. For antiglobulin-phase antibody detection methods:

   1. discuss the principle of the direct antiglobulin test (DAT) and the indirect antiglobulin test (IAT).
   2. describe the principle of preparation of antihuman globulin serum (AHG serum).
   3. state the constituents of polyspecific AHG serum.
   4. describe the methods used to perform IAT and DAT tests.
   5. discuss the importance of washing red cells prior to addition of AHG serum.
   6. discuss two reasons why IgG sensitized red cells are added to negative antiglobulin tests.
   7. differentiate between the DAT and the IAT in terms of method and uses.
   8. discuss the use of IAT positive and negative controls when doing antiglobulin tests.
   9. state the immunoglobulin class(es) of antibodies detected in the IAT phase and provide a list of antibody specificities detected.
   10. methods and indications for use of saline, albumin, LISS, and enzyme IAT phases.
   11. use of monospecific antisera.
   12. daily quality control.

5. For the following antibody detection methods done at 37^oC (saline , LISS, albumin addition, one-stage enzyme, two-stage enzyme), discuss details of the technique, including:

   1. principle of the test
   2. method (procedural steps)
   3. controls required
   4. interpretation of results (hemolysis and agglutination)
   5. antibodies best detected
   6. advantages and disadvantages

6. For the indirect antiglobulin test (IAT), including saline-IAT, enzyme-IAT, and LISS-IAT, discuss the following:

   1. principle of the test
   2. method (procedural steps)
   3. controls required
   4. interpretation of results
   5. antibodies best detected
   6. implications of types of specimens and antiglobulin sera used to detection of complement
   7. causes of false positive and negative results

7. For these antibody screen methods (gel, polyethylene glycol (PEG), solid phase immune adherence assays [SPAA], ABS 2000), discuss:
   1. principle of the test
   2. potential for semi-automation and full automation
   3. advantages and disadvantages
   4. limitations

Miscellaneous Methods

8. Describe the methods used to perform absorptions (cold and warm) and list four applications.

9. Discuss antibody elution methods as follows:
   1. methods used to obtain an eluate for antibody identification (Rubin diethyl ether; Landsteiner heat; chloroform; xylene; acid elutions): general method, advantages and disadvantages, applications (HDN, AIHA, etc.), quality control of the last wash.
   2. methods used to obtain a negative DAT (modified heat; chloroquine diphosphate, ZZAP): general method; advantages; disadvantages; applications (prior to warm autoabsorption; prior to antigen typing).
   3. reasons for the number of elutions in a) and b).

10. Discuss antibody titration methods under the following headings:

    1. define titre
    2. define prozone phenomenon and state its cause
    3. applications (list five)
    4. describe titration methods
    5. limitations
    6. ways to minimize variables (list three)
    7. significant increase in titre

11. For prewarm technique:

    1. describe its method (in general).
    2. discuss its purpose and application to sera containing cold autoantibodies and cold alloantibodies.
    3. explain its implication for the need to antigen type donor blood.

12. For reading and recording hemagglutination tests:

    1. discuss four reasons why tests are graded (- to 4+).
    2. explain why tests are read for both agglutination and hemolysis.
    3. explain why tests are read both macroscopically and microscopically.
    4. explain why tests must be handled gently when reading them.
    5. explain the importance of making legible entries on worksheets when recording tests, and why results are recorded using a pen, with corrections initialled.
    6. explain why microscopic readings are made on approximately 4 - 6 fields.
    7. explain why a test result should be recorded immediately after reading.
    8. discuss the importance of paying careful attention to clerical details.

Assessment for Internet-based modules is based on a Pass/Fail status. To receive credit for a module, you must pass each assignment, demonstrate satisfactory performance for participation in the module, and meet module deadlines.

More specifically, assessment is based on these criteria:

• Work of satisfactory quality must be submitted for assignments, with the pass mark set at 70%. Besides evaluation, assignments are meant to enhance learning. If work quality is inadequate, an exercise will be returned as unacceptable with comments and suggestions for improvement. You will then have time to improve and resubmit it by a mutually agreeable due date. If the revised work is acceptable, you will then receive credit for it.

• Participation in class discussions must be of satisfactory quantity and quality. Participating in class discussions is mandatory. This is achieved by exercises that require you to post comments and questions to the class mailing list.

• Assignments must be submitted according to due dates and within the time limit set for the module. Late work will not be accepted unless a delay has been negotiated with the instructor.

Learning Objectives