you perform using a sumber of plant tissue. The peroxide 0.2. You performed tiss
ID: 270416 • Letter: Y
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you perform using a sumber of plant tissue. The peroxide 0.2. You performed tissue printing in the laboratory enzyme was observed in all the plants plants. Some pl prts studied, however the expression level was not the same in all the expressed a ants much higher level of perox ide then others? Please explain how the expression level of peroxidase will relate to the physiology of the plant Q.3. ELISA test is routinely done to test for AIDS virus infection. By using a simple flow diagram to explain the procedure you 'll undertake to take to test for the presence of AlDS virus in a human blood sample?Explanation / Answer
Peroxidases (E.C. 1.11.1.7) are heme-containing enzymes that oxidize a variety of hydrogen donors at the expense of hydrogen peroxide. The activity of peroxidase (POD) has been correlated with plant physiological processes such as lignification, suberization, auxin metabolism, wounding, and disease resistance. The enzyme is ubiquitous and occurs in multiple isoforms in plants. The enzyme is involved in a large number of biochemical and physiological processes and may change quantitatively and qualitatively during growth and development. It acts as a signal molecule and regulates activities such as growth, morphogenesis and development. Plants respond to various environmental challenges with diverse biochemical changes such as lignification. The enzyme oxidizes phenolic monomers to form lignin, function in H2O2 production. An increase in peroxidase activity has been reported as an early response to different stresses such as salinity, drought, and exposure to toxic compounds such as heavy metals. Increased activity provides resistance against formation of H2O2. The activity of the different peroxidase isoenzymes depends on factors such as temperature, season.
Enzyme-Linked Immunosorbent Assays (ELISA) is the most commonly test to screen for HIV infection because of its simple methodology, high sensitivity, and suitability for testing large numbers of blood samples. Enzyme conjugates bind to specific HIV antibody, and substrates/chromogens produce color in a reaction catalyzed by the bound enzyme conjugate. The method for ELISA is as represented as different steps
Step 1 Take a 96-well microtiter plate.
step 2 Add diluted serum (400-fold)
step 3Add antigen (coated solid support)
step 4 Reaction with antibody (primary) for 30 minutes at 37º C or 40º C
step 5 Wash to remove unbound serum components
step 6 Addition of a conjugate (an antihuman immunoglobulin with a bound enzyme)
step 7 Binding of specific antibody (secondary) to the antigens (present on the solid phase).
Step 8 Washing
Step 9 Addition of substrate
Step 10 Color development
Step 11 Measurement of color by spectrophotometer
Amount of color is proportional to the amount of antibody bound (i.e. antibody concentration).
Alternate ELISA methodologies are also done. It is based on a competitive format in which specific HIV antibody in the sample competes with an enzyme-bound antibody reagent for antigen sites on the solid phase. Here the color development is inversely proportional to specific HIV antibody concentration.
Recently antigen sandwich method has been developed in which enzyme (alkaline phosphatase or horseradish peroxidase) is conjugated to an HIV antigen (similar to the immobilized antigen on the solid phase). The antibody is "sandwiched" between two antigen molecules, one that is immobilized on the solid phase and other containing the enzyme. The addition of substrate results in color development which is directly related to concentration of antibody. The method is more sensitive and effective in detecting all isotypes of antibody (including IgM).
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