Expt2 - B I/CH 368 Laboratory E xperiment 2 H eart Attack A...

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1 BI/CH 368 Laboratory Experiment 2 Heart Attack? Analysis of Isozymes in Blood Samples from ER Patients Case Background You and your lab partner are third-year medical students in a coronary care unit. The previous day, there were two admissions of patients with suspected myocardial infarction (heart attack). Both patients were in their late forties and had suffered intense chest pains. Patient #1 (male) had taken ill at a local restaurant, while Patient #2 (female) had just returned home after a strenuous running workout. Two blood samples were taken from each patient, at approximately 2 hours and 24 hours after the onset of the chest pains, for assay of blood serum lactate dehydrogenase (LDH). Now you are faced with another acute admission (totally unrelated to the first two cases) and there is no available bed in the coronary unit!!! Bearing in mind that an initial myocardial infarction may be followed shortly by another, is it possible from the serum LDH data (or from additional data) to decide whether either Patient #1 or Patient #2 could be moved safely to a general medical ward (or even released) to make room in your coronary care unit for the latest admission? What are your deciding factors? Experimental Background Cytosolic proteins are normally retained efficiently inside cells. Tissue damage may compromise membrane integrity sufficiently to allow release of these cytosolic proteins. The insult does not necessarily have to be large enough to result in cell death. Release of tissue proteins into the extracellular environment may occur because of exposure to toxic chemicals, for example in cirrhosis of the liver due to alcohol abuse, or release could be because of some other tissue shock or damage. Assay for certain isozymes not normally found in the bloodstream can be a useful diagnostic tool, as you will explore in this project. Experimental Procedure Week 1. LDH Isozyme Analysis What would you expect to have happened to blood LDH levels in heart attack patients? During this portion of the project, you will use native gel electrophoresis to separate the different isozymes of LDH that are present in the blood samples of the two patients. By using a specific “activity stain” following this kind of electrophoresis, only the LDH subunits will be displayed, and a very clear interpretation of the makeup of your particular LDH sample can be made. Hopefully it will prove helpful to compare the different LDH’s from the blood samples of your patients. Patient 1 suffered chest pains while dining out. Patient 2 suffered chest pains after a run.
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BI/CH 368 Laboratory – Experiment 2 – Spring 2012 2 Part I. Separation of LDH isozymes by charge Differences in charge between proteins will result in differing electrophoretic mobilities, assuming that differences in size do not become important as well. One way to avoid this is to run such charge-separation gels in a matrix with huge pores, such that the size of the protein has no bearing on its mobility. Such is the case when proteins are run through an agarose gel.
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