MA and elisa

MONOCLONAL ANTIBODIES

• WHAT ARE MONOCLONAL ANTIBODIES?

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• 1. Monoclonal antibodies are identical antibodies with the same tertiary structure (have the same antigen binding site )
• PRODUCED IN LARGE AMOUNTS FROM A SINGLE CLONE OF ANTIBODY-PRODUCING B CELLS
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• 2. USES OF MONOCLONAL ANTBODIES ARE - 1. target medication to specific cell types 2. medical diagnosis
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• 3. target medication to specific cell types by attaching therapeutic drug to an antibody (e.g cancer cells display different antigens to healthy cells )<---- can be targeted by Monoclonal antibodies with a toxic drug attached . Means cancer cells are killed and healthy cells are unaffected
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• 2. medical diagnosis eg-
• a) detect presence of specific antigens (i.e pathogens) in body fluids to diagnose if a person is infected with a particular disease
• b) detect presence of specific antibodies produced by a person against an antigen

ELISA TEST

• IMPORTANCE: method by which a medical diagnosis can be carried out
• ISSUES- new monoclonal antibody therapies tested on animals before they can move onto human (unethical)
• 1. what is it?
• a method of measuring the amount of antigen (direct elisa)- can determine if a pathogen is present in sample
• - amount of antibody (indirect elisa ) in a sample - can determine if an individual has antibodies against the pathogen indicating a previous or current infection

Stages of Direct elisa test

• stage 1- performed on a plastic tray divided into wells each has been coated with monoclonal antibody (produced specifically against the antigen of the pathogen to be detected)
• - monoclonal antibody is irreversibly bound to the plastic surface of the well
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• stage 2 - fluid sample to be tested (e.g blood, urine) is added to well, if molecules of the specific antigen are present, they will bind to the monoclonal antibodies to form antibody-antigen complexes
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• Stage 3- a second monoclonal antibody (also specific to same antigen) , which has an enzyme attached to it is added to the well
• - second antibody can only bind to the specific antigen to form an antibody-antigen complex (cannot bind directly to the first antibody or the plastic)
• - the well is washed - to remove any unbound second antibody so that any second antibody(with enzyme attached to it) will only remain in the well if the specific antigen is present and the second antibody attaches to it
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• Stage 4- suitable substrate (usually colourless) for the enzyme (attached to second antibody) is then added. If enzyme is till present in the well--> convert the colourless substrate to coloured product
• - coloured product- only be formed when the enzyme remains in the well showing that the second antibody attached to the specific antigen (indicate a + result for detection of the antigen (pathogen) in the sample being tested )
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• COLOURLESS- if the specific antigen is not present (all the second antibody with attached enzyme will be washed away during step 3 , no enzyme will be present to convert colourless substrate---> coloured product