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Micro Labs 17-20


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Lab 17
Describe the concept and general procedure for using serologic testing to identify unknown antigens (direct serologic testing).
The test is based on the fact that antigen-antibody reactions are very specific (antibodies usually only react with the antigen that stimulated their production). A suspension of unknown antigen is mix with a known antibody and a reaction between them is looked for.
Lab 17
Describe how to determine serologically whether an organism is a subgroup A, B, C, or D Shigella.
The unknown antigen (suspected Shigella) is placed on each of the four circles on a serology slide and a different antiserum for each of the subgroups is then add to each circle. A reaction is positive when clumping is apparent.
Lab 17
Correctly interpret the results of the following serological tests:
a. serological typing of Shigella
b. serological typing of streptococci
c. serological testing for pregnancy
a. Clumping in one of the serology wells indicates a positive reaction with one of the subgroup antibodies.
b. Clumping of the latex in the wells indicates a positive for that Lancefield group.
c. A red line crossing the blue line indicates a positive; HGC (the hormone being tested for) binds to antibodies along a strip in the test card, other antibodies bound to latex then bind to HGC turning the strip red and indicating a positive.
Lab 17
Define serology.
Refers to using atigen-antibody reactions for diagnostic purposes
Lab 17
Define antigen and state what may act as an antigen.
A substance that reacts with antibody molecules and antigen receptors of lyphocytes. Can be large molecular weight proteins or polysaccharides.
Lab 17
Define antiserum.
The known preparation of antibodies used in direct serological testing.
Lab 17
Describe one way of producing known antiserum.
Monoclonal antibody technique: The spleen of an animal infected with the antigen that the desried antibodies bind to is removed. The desired antibody is isolated by finding the plasma cell that produces it and fusing it with a myeloma (cancer) cell creating a hybirdoma cell that will grow out side of the body and secrete the desired antibodies.
Lab 18
State the difference between a qualitative serological test and a quantitative serological test.
Qualitative test only determines whether the antibodies are present and is used for screening purposes. Quantitative test give the titer or amount of the antibody in the serum and is used to moniter progress of a disease.
Lab 18
State what disease the RPR and the FTA-ABS procedures test for. Indicate which of these is a presumptive test, which is a confirming test, and why.
RPR: tests for syphilus. It is a presumptive test because it detects antibodies made against the body's own lipids (nontreponemal) that may be present during other diseases, as well. FTA-ABS: tests for syphilus. Is a confirming tests because it test for antibodies made specifically against the syphilus-causing spirochete Treponema Pallidum(treponemal).
Lab 18
State the significance of nontreponemal antilipid (reagin) antibodies in serological testing.
The antibodies are not found is normal serum but are present in a patient's serum if they have syphilus. They are not, however, unique to syphilus infections.
Lab 18
State the significance of heterophile antibodies in serological testing.
During infections caused by EBV the body produces nonspecific heterophile antibodies. These antibodies can be detected because of a cross reaction with horse or sheep blood (serving as known antigen) that causes the red blood cells to clot.
Lab 18
State the significance of anti-deoxyribonucleoprotein antibodies in serological testing.
During the autoimmune disease SLE (lupus) the body makes specific antibodies against its oun DNA (anti-deoxyriboneucleoprotein antibodies). In testing for the antibodies the known antigen is DNA attached to latex so clumps will be visible.
Lab 18
Briefly describe the EIA test for HIV antibodies and state the significance of a positive HIV antibody test.
Wells to which HIVs have been adsorbed are first filled with the patient's serum and then washed to remove any serum compontents that did not bind to the HIVs. Anti-human gamma globulin (anti-HGG)antibodies are added to the wells, after which the wells are washed to remove any of these antibodies that didn't bind to the serum antibodies. Finally a substrate that will react with the enzyme bound to the anti-HGG is added which will change color if the test is positive, indicating the patient has HIV antibodies and therefore HIV. If a test is positive it is immediately repeated to ensure accuracy.
Lab 18
State the most common reason for a false-negative HIV antibody test.
Usually occur because the patient has been recently infected and has not had time to make sufficiant amounts antibodies to be detected.
Lab 18
Interpret the results of the following serological tests:
a. serologic test for infectious mononucleosis
b. serologic test for SLE
c. FTA-ABS test
a. Positive will show agglutination of horse erythrocytes.
b. Positive will show cluping a DNA attached to latex (for visibility) by anti-deoxyribonucleoprotein.
c. Positive reaction will show florescing spirochetes because antibodies against the Treponema pallidum antibodies have been attached to florescent particles.
Lab 19
Define the following terms: sterilization, disinfection, static, and cidal.
Sterilization: destruction of all life. Desinfection: reduction or elimiation of pathogens. Static: inhibits growth. Cidal: kills microbes.
Lab 19
State whether moist or dry heat is more effective in controlling microorganisms, and indicate why.
Moist heat is the most effective in killing because of its ability to penetrate cells.
Lab 19
State specifically how moist heat kills microorganisms.
Moist heat kills by denaturing proteins and possibly melting lipids.
Lab 19
State two methods of applying moist heat.
Autoclaving (steam under preasure) or boiling water.
Lab 19
Briefly describe the process of autoclaving (pressure, time, and temperature).
Autoclaving is raising the boiling point of water to 121 degrees C by raising the preasure to 15 lbs/square inch. 15-45 minutes is sufficient time for it to be cidal.
Lab 19
State whether or not boiling is an effective means of sterilization, and indicate why.
No, since boiling will not destroy endospores and some viruses can survive extensive boiling.
Lab 19
State two methods of applying dry heat.
Hot air sterilization and incineration.
Lab 19
Define pasteurization.
Mild heating of materials to kill particular spoilage or pathogenistic organisms.
Lab 19
State whether low temperature has a static or cidal effect on microorganisms, and indicate why.
It has a static effect on microbes since it inhibits growth but doesn't kill.
Lab 19
State whether desiccation has a static or a cidal effect on microorganisms, and indicate how it affects the cell.
It has static effect on cells. Dehydration of a cell inhibits the microbial enzymes.
Lab 19
Describe osmosis in terms of water flow through a semipermeable membrane.
Osmosis is simple defustion of water through a membrane from areas of higher water concertration to lower water concentration.
Lab 19
Define the following terms: a)hypotonic, b) hypertonic, c) isotonic, d) plasmoptysis, and e) plasmolysis.
a) solute concentration is greater inside of the cell than outside.
b) solute concentration is greater outside of the cell than inside.
c) solute concentration is equal inside and outside of the cell.
d) bursting of a cell.
e) shrinking of the cytoplasmic membrane due to hypertonic environment; causes dehydraton.
Lab 19
State whether hypertonicity has a static or a cidal effect on microorganisms.
Has a static effect. (Plasmolysis)
Lab 19
State how the wavelength and the length of exposure influence the bacteriocidal effect of UV light.
The longer the bacteria is exposed to UV light between 260-270 nm the greater the cidal effect.
Lab 19
Describe specifically how UV light kills microorganisms.
UV light is absorbed by DNA and causes thymine-thymine dimers, resulting in premature termination of DNA base pairing at the site of the dimer. During the bodies efforts to fix the DNA even more mutation takes place, causing more damage.
Lab 19
State why UV light is only useful as a means of controlling surface contaminants and give several practical applications.
Because of its poor penetrating power it is used only for disinfecting hospital rooms and sinks, microbiological hoods, or on food processing equipment. It is also harmful to humans.
Lab 19
State why filters are preferred over autoclaving for such materials as vaccines, antibiotic solutions, sera, and enzyme solutions.
It can sterilize solutions like vaccines and vitamins that would be damaged by autoclaving.
Lab 20
Define the following terms: disinfection, disinfectant, antiseptic.
Disinfection: reduction or elimination of pathogenic microbes or materials. Disifectant: chemicals used to disinfect inanimate objects. Antiseptic: nontoxic disinfectant suitable for animal tissue.
Lab 20
State why chemical agents are usually unreliable for sterilization.
They don't quickly and reliably kill all viruses, microbes and endospores.
Lab 20
List five factors which may influence the antimicrobial action of disinfectants and antiseptics.
1. concentration of agent
2. temperature at which agent is used
3. types of microbes present
4. number of microbes present
5. nature of material carrying the microbes (dirt or other materials might inhibit agents).
Lab 20
Describe two modes of action of disinfectants and antiseptics (i.e., how they harm the microorganisms).
1. Damage the lipids and/or proteins of the cyto membrane causing cellular leakage.
2. Denature microbial enzymes and proteins blocking metabolism.
Lab 20
State why the results of an in vitro test to evaluate chemical agents may not necessarily apply to in vivo situations.
The conditions of the in vivo test may vary from the conditions of the in vitro test.
Lab 20
Define transient flora and resident flora and compare the two groups in terms of ease of removal.
Resident flora are normal flora of the skin. Transient flora are aquired from physical contact. Many normal flora are impossible to completely remove because they live in sweat glands or deep in the skin. Transient flora are mostly removed with hand washing.
Lab 17
Define antibody and state where they are primarily found in the body.
Specific protein configurations produced by B-lymphocytes and plasma cells in reponse to specific antigens and capable of reacting with that antigen. They are found mainly in the plasma portion of the blood.
Lab 17
Define: a) direct serologic testing and b) indirect serologic testing.
a) the use of known antibodies, called antiserum, to identify an unknown antigen.
b) unknown antibodies being made against an antigen associated with a particular disease are detected using a known antigen.
Lab 18
State the principle and the general procedure behind indirect serologic testing.
This type of testing is based on the fact that antibodies are only produced in response to a specific antigen. A patients serum is mixed with the known antigen and the antibody-antigen reaction is looked for.
Lab 18
Define titer.
Indicates how much it is possible to dilute a patients serum and still have detectable amounts of antibody in the serum (the amount of a certain antibody in the serum). The more antibodies present the more it is possible to dilute the serum and still ditect the antibodies.
Lab 19
Describe how ionizing radiation kills microorganisms and state several common applications.
It ionizes water and other molecules to form radicals that can break DNA. Used to sterilize desposable medical supplies like gloves or syringes or to retard spoilage in meats or fruit.
Lab 20
Name two chemical agents which are reliable for sterilization.
Ethylene oxide gas and aldehyde.

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