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Understanding the method used for a test provides a broader context for understanding your results. This article provides brief explanations of several common laboratory methods mentioned on this site.

Accordion Title
검사 방법(Laboratory Methods)
  • Overview

    Laboratories use a variety of methodologies to test the countless analytes that are of interest to the medical community. Understanding the method used for a test provides a broader context for understanding your test results. Below are links to explanations of several common laboratory methods mentioned on this site.

    Laboratory methods are based on established scientific principles involving biology, chemistry, and physics, and encompass all aspects of the clinical laboratory from testing the amount of cholesterol in your blood to analyzing your DNA to growing microscopic organisms that may be causing an infection. Such methods are much like the recipes in a cookbook, defining the procedures or processes that are used to test biological samples for particular analytes or substances. The laboratory scientist follows step-by-step procedures until the end product, a test result, is achieved.

    Some methods, like some recipes, are much more complicated and labor-intensive than others and require varying degrees of expertise. Often, there may be more than one method that can be used to test for the same substance. Consequently, the same analyte may be tested differently in different laboratories, a fact that is crucial when comparing test results.

    The descriptions of the methods listed below attempt to give some insight into the scientific principles used and the steps that are required to produce a result. Explanations of the methods – and their differences – are provided to give you a better understanding of some of the tests that you may undergo. These bulleted items are not intended to be a comprehensive list of available methodologies, but do represent some of those that are mentioned on this web site.

  • Immunoassay

     

    Immunoglobulins are proteins produced by the immune system to recognize, bind to, and neutralize foreign substances in the body. Immunoassays are tests based on the very specific binding that occurs between an immunoglobulin (called an antibody) and the substance that it specifically recognizes (the foreign molecule, called an antigen). Immunoassays can be used to test for the presence of a specific antibody or a specific antigen in blood or other fluids.

    When immunoassays are used to test for the presence of an antibody in a blood or fluid sample, the test contains the specific antigen as part of the detection system. If the antibody being tested for is present in the sample, it will react with or bind to the antigen in the test system and will be detected as positive. If there is no significant reaction, the sample tests negative. Examples of immunoassay tests for antibodies include Rheumatoid Factor (which tests for the presence of autoimmune antibodies seen in patients with rheumatoid arthritis), West Nile Virus (which tests for antibodies that a person made in response to an infection with that virus) or antibodies made in response to a vaccination (such as tests for antibodies to Hepatitis B to assure that the vaccination was successful).

    When immunoassays are used to test for the presence of antigens in a blood or fluid sample, the test contains antibodies to the antigen of interest. The reaction of the antigen that is present in the person’s sample to the specific antibody is compared with reactions of known concentrations and the amount of antigen is reported. Examples of immunoassay tests for antigens include drug levels (like digoxin, vancomycin), hormone levels (like insulin, TSH, estrogen), and cancer markers (like PSA, CA-125, and AFP).

    Sources
    (© 2006). Immunoassay Detection Technologies, Chapter 2. Abbott Diagnostics Scientific Resources Learning Guide [On-line information]. PDF available for download through http://www.abbottdiagnostics.com.

    Clarke, W. and Dufour, D. R., Editors (2006). Contemporary Practice in Clinical Chemistry, AACC Press, Washington, DC. Harris, N. and Winter, W. Chapter 10, Immunoassays. Pp. 117-119.

  • Enzyme-linked immunosorbent assay (ELISA)

    This testing method is a type of immunoassay. It is based on the principle that antibodies will bind to very specific antigens to form antigen-antibody complexes, and enzyme-linked antigens or antibodies can be used to detect and measure these complexes.

    To detect or measure an antibody in a person’s blood, a known antigen is attached to a solid surface. A solution containing the patient sample is added. If the patient’s sample contains antibody, it will bind to the antigen. A second antibody (against human antibodies) that is labeled with an enzyme is then added. If the enzyme-linked antibody binds to human antibodies, the enzyme will create a detectable change that indicates the presence and amount of the antibody in the patient sample.

    Sources
    (© 1994-2006). Introduction to Antibodies – Enzyme-Linked Immunosorbent Assay (ELISA). Millipore Corporation [On-line information]. Available online at http://www.chemicon.com/resource/ANT101/a2C.asp through http://www.chemicon.com.

    (2001). Gerostamoulos, J. et. al. (2001). The Use Of Elisa (Enzyme-Linked Immunosorbent Assay) Screening In Postmortem Blood. TIAFT, The International Association of Forensic Toxicologists [On-line information]. Available online at http://www.tiaft.org/tiaft2001/lectures/l13_gerostamoulos.doc through http://www.tiaft.org.

    Clarke, W. and Dufour, D. R., Editors (2006). Contemporary Practice in Clinical Chemistry, AACC Press, Washington, DC. Harris, N. and Winter, W.

  • Western blot

    This is an immunoassay test method that detects specific proteins in blood or tissue. It combines an electrophoresis step with a step that transfers (blots) the separated proteins onto a membrane. Western blot is often used as a follow-up test to confirm the presence of an antibody and to help diagnose a condition. Examples of its use include confirmatory HIV and Lyme disease testing.

    To perform a western blot test, a sample containing the protein is applied to a spot along one end of a layer of gel. Multiple samples and a control may be placed side by side along one end of the gel in separate “lanes.” An electrical current causes the proteins in the sample(s) to move across the gel, separating the proteins by size and shape and forming bands that resemble the steps of a ladder. These sample and control ladders are then “blotted” (transferred) onto a thin membrane that is put in contact with the gel. Labelled or tagged antibodies are then used in a one or two step process to detect the proteins bound to the membrane. For example, to confirm HIV or Lyme antibody tests, the proteins separated are those of the causative organism. A patient’s sample is then added to the blot and any antibodies to the organism are bound and later detected by labeled antibodies to human immunoglobulins. The presence of the certain proteins is interpreted by comparison with known negative or positive control samples in the other lanes.

    Sources
    Khalsa, G. Western blotting. Arizona State University, School of Life Sciences, Mama Ji’s Molecular Kitchen [On-line information]. Available online at http://lifesciences.asu.edu/resources/mamajis/western/western.html through http://lifesciences.asu.edu.

    Tietz Textbook of Clinical Chemistry and Molecular Diagnostics. Burtis CA and Ashwood ER, Bruns DE, eds. 4th edition St. Louis: Elsevier Saunders; 2006.