Vishy- Ag/ Ab reactions

Question 1

IgM have ____ binding sites for antigen. ____ refers to minimal conc of antibodies that can succ bind to antigens and cause a detectable test reaction.

Answer 1

10; Titers

Question 2

Describe agglutination reactions

Answer 2

1. Direct agglutination: Belong to IgM; form lattice. If they react with cell surface antigens, they aggregate and form lattice.
2. Indirect agglutination: Participating Ab belong to IgG –> monomers. 2 binding sites for Ag. They dont aggregate as well and dont form lattice. Have to add secondary Ab –> anti Ab that bind to primary Ab and form lattice.

Question 3

what are the two types of Ag/Ab interactions

Answer 3

1. Agglutination reactions

2. Precipitation reactions

Question 4

Describe the single diffusion types of Precipitation reactions

Answer 4

1. Single diffusion: When Ab bind to soluble Ag in liquids. They form immune complexes and precipitate. Ab are included along with molten agar poured on a glass plate. When agar solidifeids, a well is created and Ag are introduced into the well and keep diffusing in agar where it meets Ab. There is formation of precipitate ring. The farther away the precipitate ring is away from well = conc of Ag is higher. If precipate ring is closer to well, the conc of Ag is low. This technique is quantative.

Question 5

What is equivalence zone

Answer 5

The conc of both Ab and Ag are optimal and there is formation of a detectable precipitation reaction

Question 6

what is zone of ab excess

Answer 6

no detectable precipiation reaction bc of excess Ab

Question 7

what is zone of Ag excess

Answer 7

no detectable precipitation reaction bc of excess Ag

Question 8

Describe double diffusion

Answer 8

Molten agar poured on plate. Multiple wells are created. In some wells ag is introduced in some wells Ab are introduced. Once Ag diffuse and they bind Ab they form a precipitate ring, if there is cross reactvity, there iwll be partial binding and precipiate rings. This is qualitiative! Ab

Question 9

What is immunoelectrophoresis

Answer 9

Measures serum proteins. Proteins present in serum go towards anode/cathode dep. on charge. In the wells, ab are introduced and ag and ab diffuse. If Ab are directed towards Ag there will be preciptate rings.

Question 10

What is radio immunoassay

Answer 10

used to measure levels of hormone in human blood. Based on competition bet radio labeled antigen and unlabeled Ag in human blood sample for Ab.

Question 11

Less radioactivity means ____hormones in patients sample. High radioactivity means _____hormones in patient’s sample

Answer 11

more; less

Question 12

What is ELISA

Answer 12

Measure the levesls of Ag or Ab. Secondary Ab is commercial and conjugated with an enzyme which could be alkaline phosphatase or horse radish peroxidase. It binds to primary Ab. Wash to remove unbound Ab. When you add substrate, there will be color formation. Positive color formation means positive test reaction

Question 13

What are advantage of ELISA

Answer 13

1. Sensitive: nanogram levels or lower
2. Reproducible
3. Minimal reagents
4. Qualitative and Quantitative
5. Greater scope: Wells can be coated with Ag or Ab
6. Suitable for automation –> high speed
7. NO radiation hazards

Question 14

Describe Western Blotting

Answer 14

Proteins are run in electrophoresis and undergoe resolution based on molecular weight. High on top, low on bottom. Transfer all proteins to a membrane and incubate it with patients serum and add secondary Ab and add substrate. You will then be able to see bands.

Question 15

What is immuno flourescence

Answer 15

Qualitative! Detects the expression of Ag on cell surface of human cells or bacterial cells. ab are conjugated with flouresence dye and observe under flouresece microscope. If flourescene under microscope = +

Question 16

The flourescene markers PE (red) or flourescein (green)

Answer 16

Ab can be covalently linked to these flourescence markers.

Question 17

Describe direct and indirect immunoflourescence

Answer 17

Direct: Primary Ab conjugated with flourescence molecule

Indirect: secondary Ab are conjugated with flourescence dye.

Question 18

What is FACS

Answer 18

Used to quantify the number of CD4 + T cells. When there is loss of these types of cells = person at risk for tuberculosis infection. Take immune cells from body and stain for CD4 marker with FITC and pass cells through fax machine.

Question 19

How can you determine phagocytic efficiency of monocytes

Answer 19

if you see intracellular organisms the macrophages have normal ability to ingest microorganisms.

Question 20

How would you determine if lymphocytes have normal capacity to multiply in response to Ag

Answer 20

collect peripheral blood mononucleal cells. Throw Ag or mitogen. If Ag then monocytes will present Ag to T cells or B cells then …add radioactive thymidine. Once T cells multiply radioactive thymidine get incorporated in DNA. High radioactive counts amounts means cells have normal capacity to multiply in response to Ag or mitogen

Question 21

how do you det cytotoxic ability of NKs

Answer 21

take target cells that have low exp of MHC molecles, add NKs and label target cells with radioactive chromium. RC gets incorporated into cytoplasmic proteins of target cells. IF NK have normal bailty to kill target cells they will release perforin and granulysin and this will result in rupture of target cells which will release radio active chromium into supernatant. Collect supernatant and measure radioactivity. High radioactivity means NK have normal ability to kill target cells that are abnormal

Question 22

People with A BG will have ____ antibodies. People with B BG will have ____ antibodies. People with AB wont have either Ab. People with O blood group will have ____ and ____ antibodies.

Answer 22

B; A; AB

Question 23

What is blood group antigens

Answer 23

carbohydrate molecules exp on cell surface of RBcs and WBC’s and is commonly referred to as H antigen.

Question 24

People with O blood group have Nac glucosamine, ____ and ______

Answer 24

galactose; fucose

Question 25

People with A blood group have N ac glucosamine, ____, galactose and ____

Answer 25

fucose, NAc galactose

Question 26

People with B blood group have N ac glucosamine, fucose, galactose, and _____

Answer 26

galactose

Question 27

If mother is Rh - and the fetus is Rh + what happens

Answer 27

during delivery there will be mixing of blood, mothers immune system will see Rh antigen and produce anti Rh antibodies. If mother is pregnant the second time and fetus is Rh +, anti Rh antibodies present in mother will travel to placenta and there will be Ag/Ab reaction and can result in death of fetus.

Question 28

What is Type I hypersensitivy reactioysn

Answer 28

immediate hypersensitivity reaction (occurs within mins to hours) IgE; basophils and mast cells

Question 29

What is Type II hypersensitivy rxn

Answer 29

complement activation in response Ab binding to cell surface antigens (occurs within hours to days) IgG or IgM Ab’s–> bind to cell surface Ag. There will be activation of complement. Has localized effect

Question 30

What is Type II hypersensitivity rxn

Answer 30

complement activation due to soluble Ag/Ab complexes (occurs within hours to days) Ab belong to IgG or IgM bind to soluble Ag’s. There is formation of Immune complex which can be carried to any part of body, where there will be complement activation. Has a systemic effect

Question 31

What is Type IV

Answer 31

Delayed type hypersensitivity rxn (takes 2-4 days)

CD4 + T cells, memory cells and DC’s. Ab dont participate in type Iv

Question 32

What is a complement fixation test

Answer 32

Used to det if a person has immune complex circulating in circulation (blood, serum, or lymph). Indicator system consists of RBc’s, Ab against RBC’s, as well as complement molecules. In assay you introduce pt’s serum which may/may not comtain immune complex. Ab against RBC will bind to RBC, there is complement activation, and RBC ruptures whci means that patients serum sample doesnt have immune complex. If patients serum has IC then complement has a choice to bind with IC from patient or wth RBC/ RBC Ab and there will be less hemolysis which means there is more IC in patients serum.

Question 33

Ab dont participate in delayed hypersensitivity reaction ___ and ___ do. In latent Tuberculosis, Ag is introduced in skin and taken up by DCS and presented to sensitized T cells which migrate from lymph node to skin.

Answer 33

CD4 + T cells; DC’s

Question 34

Direct agglutination Ab belongs to _____, Indirect agglutination belongs to ____

Answer 34

IgM; IgG