Immunodiffusion/ELISA

Question 1

Immuno is associated with…

Diffusion is linked with…

Answer 1

Antibodies and antigens

Agar plates (antibodies/antigens dffuse from high concentrations to low)

Immunodiffusion determines presence of antigens and/or antibodies - solutions diffuse precipitin forms if reaction occurs.

Question 2

Antibodies are

Answer 2

Glycoproteins - proteins that have sugars on it as well as blood proteins [immunoglobulins].

Produced by plasma cells - activated B cells due to a response to an antigen.

Y binds to antigen (& targets them for destruction by other immune system cells)

Question 3

Antigens are

Answer 3

Any type of molecule that elicits an immune response

Ex: carbs, proteins, DNA, lipids can serve

•U•

Question 4

Epitope

Answer 4

Where antibody binds to antigen

1:1 specificity - 1 antibody recognizes one antigen

Can be varied shapes because can be recognized by diff antibodies.

Question 5

Antibody:

 light chain \    / light chain

                      ^
        Variable portion

Heavy chain | | heavy chain

                      ^
          Constant portion

Answer 5

Composed of 2 heavy chains+ 2 light chains,
constant region - used by antibody to bind to cell of immune system; 5 classes: IgM,A,D,G,E,
variable region- place where antibody binds to antigen with epitope

Question 6

IgG - monomer

Answer 6

Major antibody in serum
Crosses placenta
Long-lasting antibody/immunity
mom gives during pregnancy

Question 7

IgA- monomer (blood) or dimer

Answer 7

Secretions - breast milk, genital, mucosal membranes, digestive tract, ears, eyes.
Protection as baby breastfeeds
Protect body surfaces exposes to outside foreign substances

Question 8

IgM - monomer (receptor on B cells) or pentamer (units in blood)

Answer 8

First antibody we produce

Eliminates pathogens in early stages of B cell mediated immunity before IgG is sufficient

Question 9

IgD - Monomer (receptor on B cells)

Answer 9

Antigen from environment and presents to immune system.

Can have IgD and IgM on B cell

Question 10

IgE monomer

Answer 10

Do not have Allergies - low IgE

Seasonal, food, etc Allergies- high IgE

Binds to mast cells, which granulate, histamines released - watery eyes, hives, etc.

Question 11

Serum

Answer 11

Clear to part of blood

• contains antibodies

No clotting factor

Question 12

Precipitin

Answer 12

Rxn between antibody and antigen

Forms line

Question 13

ELISA

Answer 13

Enzyme Linked Immunosorbent Assay

Question 14

Positive result in ELISA

Answer 14

Antibody present within patient’s blood - associated w/ disease presence or encounter and recovery.

Question 15

True or False - the body after recovery from disease can STILL make antibodies as an immune system response and prevention.

Answer 15

True

Question 16

Amount of color in sample

Answer 16

Quantity of enzyme (amount of antigen present in OG sample)

Question 17

1. Centrifuge

Answer 17

Separates test sample - obtain fluid serum w/ antibodies

Question 18

2. Prepare Serial Dilution

Answer 18

Determine concentration of antibody in test sample’s serum.

3 diff. solutions to perform 3 serial dilutions (serum from patient and buffer)

Question 19

3 & 4. ELISA Pretreated

Answer 19

With SLE antigens to bind to surface of well acting as protective coat - prevents antibody from sticking to wells.

When antibodies bind to antigen they do it without binding to wells themselves due to this step.

Question 20

Positive Control

Answer 20

Primary antibody - first antibody to detect foreign molecule (antigen)

Question 21

Negative control

Answer 21

Buffer

Question 22

2 controls

Answer 22

R/o error and problems with reagents changing w/time & mistakes with ELISA procedure.

Question 23

What happens if 2 positive responses are produced by both controls

Answer 23

Null results and assay must be completed again to produce accurate results

Question 24

5. Incubate

Answer 24

Allow sufficient binding between antibody - antigen to properly proceed w/ effective timing for.

Question 25

6. Wash plate

Answer 25

Unreacted antibodies need to be removed; prevent leftover antibody from binding w/ secondary antibody. (False positive)

Question 26

7. Secondary Antibody

Answer 26

Binds to primary antibody / source: diff. species than primary .

Secondary antibody distinguishes primary antibody as foreign.