Radioimmuno Assay

Presented by Mamona WaheedPresented to Sir Alamgeer

Radioimmunoassay

bullRadioimmunoassay (RIA) is a very sensitive in vitro assay technique used to measure concentrations of antigens (for example hormone levels in the blood) by use of antibodies

bullRIA technique is extremely sensitive and extremely specific requiring specialized equipment it remains the least expensive method to perform such tests

bullThe technique was introduced in 1960 by berson and yalow as an assay for the concentration of insulin in plasmabull It represented the first time that hormone levels in the blood could be detected by an in vitro assay

bullThe technique of radioimmunoassay has revolutionized research and clinical practice in many areas eG bullBlood banking bullDiagnosis of allergies bullEndocrinology

Radioimmunoassay pros and consbull PRO versatility using the same principle almost any biomolecule can be assayedbull Fast (usually 2 days or less)bullSensitive (comparable to the most sensitive bioassays that is lt ngml)bull Large capacity thousands of samplesday specific (antibody-dependent)

Conbull Use of radioactivity hazardousbullExpensive equipment (gamma or betabullCounter)

bullPrinciple

bullThe technique is based on the ability of an unlabelled form of the substance to inhibit competitively the binding of a radioactively labelled substance by specific antibodies

Method

bullTo perform a radioimmunoassay a known quantity of an antigen is made radioactive frequently by labeling it with gamma-radioactive isotopes of iodine attached to tyrosine (hot)bullThis radiolabeled antigen is then mixed with a known amount of antibody for that antigen and as a result the two specifically bind to one another

bull Then a sample of serum from a patient containing an unknown quantity of that same antigen is addedbull This causes the unlabeled (or cold) antigen from the serum to compete with the radiolabeled antigen (hot) for antibody binding sites

bullAs the concentration of cold antigen is increased more of it binds to the antibody displacing the radiolabeled variant and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

Radioimmunoassay

bullRadioimmunoassay (RIA) is a very sensitive in vitro assay technique used to measure concentrations of antigens (for example hormone levels in the blood) by use of antibodies

bullRIA technique is extremely sensitive and extremely specific requiring specialized equipment it remains the least expensive method to perform such tests

bullThe technique was introduced in 1960 by berson and yalow as an assay for the concentration of insulin in plasmabull It represented the first time that hormone levels in the blood could be detected by an in vitro assay

bullThe technique of radioimmunoassay has revolutionized research and clinical practice in many areas eG bullBlood banking bullDiagnosis of allergies bullEndocrinology

Radioimmunoassay pros and consbull PRO versatility using the same principle almost any biomolecule can be assayedbull Fast (usually 2 days or less)bullSensitive (comparable to the most sensitive bioassays that is lt ngml)bull Large capacity thousands of samplesday specific (antibody-dependent)

Conbull Use of radioactivity hazardousbullExpensive equipment (gamma or betabullCounter)

bullPrinciple

bullThe technique is based on the ability of an unlabelled form of the substance to inhibit competitively the binding of a radioactively labelled substance by specific antibodies

Method

bullTo perform a radioimmunoassay a known quantity of an antigen is made radioactive frequently by labeling it with gamma-radioactive isotopes of iodine attached to tyrosine (hot)bullThis radiolabeled antigen is then mixed with a known amount of antibody for that antigen and as a result the two specifically bind to one another

bull Then a sample of serum from a patient containing an unknown quantity of that same antigen is addedbull This causes the unlabeled (or cold) antigen from the serum to compete with the radiolabeled antigen (hot) for antibody binding sites

bullAs the concentration of cold antigen is increased more of it binds to the antibody displacing the radiolabeled variant and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bullRIA technique is extremely sensitive and extremely specific requiring specialized equipment it remains the least expensive method to perform such tests

bullThe technique was introduced in 1960 by berson and yalow as an assay for the concentration of insulin in plasmabull It represented the first time that hormone levels in the blood could be detected by an in vitro assay

bullThe technique of radioimmunoassay has revolutionized research and clinical practice in many areas eG bullBlood banking bullDiagnosis of allergies bullEndocrinology

Radioimmunoassay pros and consbull PRO versatility using the same principle almost any biomolecule can be assayedbull Fast (usually 2 days or less)bullSensitive (comparable to the most sensitive bioassays that is lt ngml)bull Large capacity thousands of samplesday specific (antibody-dependent)

Conbull Use of radioactivity hazardousbullExpensive equipment (gamma or betabullCounter)

bullPrinciple

bullThe technique is based on the ability of an unlabelled form of the substance to inhibit competitively the binding of a radioactively labelled substance by specific antibodies

Method

bullTo perform a radioimmunoassay a known quantity of an antigen is made radioactive frequently by labeling it with gamma-radioactive isotopes of iodine attached to tyrosine (hot)bullThis radiolabeled antigen is then mixed with a known amount of antibody for that antigen and as a result the two specifically bind to one another

bull Then a sample of serum from a patient containing an unknown quantity of that same antigen is addedbull This causes the unlabeled (or cold) antigen from the serum to compete with the radiolabeled antigen (hot) for antibody binding sites

bullAs the concentration of cold antigen is increased more of it binds to the antibody displacing the radiolabeled variant and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bullThe technique was introduced in 1960 by berson and yalow as an assay for the concentration of insulin in plasmabull It represented the first time that hormone levels in the blood could be detected by an in vitro assay

bullThe technique of radioimmunoassay has revolutionized research and clinical practice in many areas eG bullBlood banking bullDiagnosis of allergies bullEndocrinology

Radioimmunoassay pros and consbull PRO versatility using the same principle almost any biomolecule can be assayedbull Fast (usually 2 days or less)bullSensitive (comparable to the most sensitive bioassays that is lt ngml)bull Large capacity thousands of samplesday specific (antibody-dependent)

Conbull Use of radioactivity hazardousbullExpensive equipment (gamma or betabullCounter)

bullPrinciple

bullThe technique is based on the ability of an unlabelled form of the substance to inhibit competitively the binding of a radioactively labelled substance by specific antibodies

Method

bullTo perform a radioimmunoassay a known quantity of an antigen is made radioactive frequently by labeling it with gamma-radioactive isotopes of iodine attached to tyrosine (hot)bullThis radiolabeled antigen is then mixed with a known amount of antibody for that antigen and as a result the two specifically bind to one another

bull Then a sample of serum from a patient containing an unknown quantity of that same antigen is addedbull This causes the unlabeled (or cold) antigen from the serum to compete with the radiolabeled antigen (hot) for antibody binding sites

bullAs the concentration of cold antigen is increased more of it binds to the antibody displacing the radiolabeled variant and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bullThe technique of radioimmunoassay has revolutionized research and clinical practice in many areas eG bullBlood banking bullDiagnosis of allergies bullEndocrinology

Radioimmunoassay pros and consbull PRO versatility using the same principle almost any biomolecule can be assayedbull Fast (usually 2 days or less)bullSensitive (comparable to the most sensitive bioassays that is lt ngml)bull Large capacity thousands of samplesday specific (antibody-dependent)

Conbull Use of radioactivity hazardousbullExpensive equipment (gamma or betabullCounter)

bullPrinciple

bullThe technique is based on the ability of an unlabelled form of the substance to inhibit competitively the binding of a radioactively labelled substance by specific antibodies

Method

bullTo perform a radioimmunoassay a known quantity of an antigen is made radioactive frequently by labeling it with gamma-radioactive isotopes of iodine attached to tyrosine (hot)bullThis radiolabeled antigen is then mixed with a known amount of antibody for that antigen and as a result the two specifically bind to one another

bull Then a sample of serum from a patient containing an unknown quantity of that same antigen is addedbull This causes the unlabeled (or cold) antigen from the serum to compete with the radiolabeled antigen (hot) for antibody binding sites

bullAs the concentration of cold antigen is increased more of it binds to the antibody displacing the radiolabeled variant and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

Radioimmunoassay pros and consbull PRO versatility using the same principle almost any biomolecule can be assayedbull Fast (usually 2 days or less)bullSensitive (comparable to the most sensitive bioassays that is lt ngml)bull Large capacity thousands of samplesday specific (antibody-dependent)

Conbull Use of radioactivity hazardousbullExpensive equipment (gamma or betabullCounter)

bullPrinciple

bullThe technique is based on the ability of an unlabelled form of the substance to inhibit competitively the binding of a radioactively labelled substance by specific antibodies

Method

bullTo perform a radioimmunoassay a known quantity of an antigen is made radioactive frequently by labeling it with gamma-radioactive isotopes of iodine attached to tyrosine (hot)bullThis radiolabeled antigen is then mixed with a known amount of antibody for that antigen and as a result the two specifically bind to one another

bull Then a sample of serum from a patient containing an unknown quantity of that same antigen is addedbull This causes the unlabeled (or cold) antigen from the serum to compete with the radiolabeled antigen (hot) for antibody binding sites

bullAs the concentration of cold antigen is increased more of it binds to the antibody displacing the radiolabeled variant and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

Conbull Use of radioactivity hazardousbullExpensive equipment (gamma or betabullCounter)

bullPrinciple

bullThe technique is based on the ability of an unlabelled form of the substance to inhibit competitively the binding of a radioactively labelled substance by specific antibodies

Method

bullTo perform a radioimmunoassay a known quantity of an antigen is made radioactive frequently by labeling it with gamma-radioactive isotopes of iodine attached to tyrosine (hot)bullThis radiolabeled antigen is then mixed with a known amount of antibody for that antigen and as a result the two specifically bind to one another

bull Then a sample of serum from a patient containing an unknown quantity of that same antigen is addedbull This causes the unlabeled (or cold) antigen from the serum to compete with the radiolabeled antigen (hot) for antibody binding sites

bullAs the concentration of cold antigen is increased more of it binds to the antibody displacing the radiolabeled variant and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bullPrinciple

bullThe technique is based on the ability of an unlabelled form of the substance to inhibit competitively the binding of a radioactively labelled substance by specific antibodies

Method

bullTo perform a radioimmunoassay a known quantity of an antigen is made radioactive frequently by labeling it with gamma-radioactive isotopes of iodine attached to tyrosine (hot)bullThis radiolabeled antigen is then mixed with a known amount of antibody for that antigen and as a result the two specifically bind to one another

bull Then a sample of serum from a patient containing an unknown quantity of that same antigen is addedbull This causes the unlabeled (or cold) antigen from the serum to compete with the radiolabeled antigen (hot) for antibody binding sites

bullAs the concentration of cold antigen is increased more of it binds to the antibody displacing the radiolabeled variant and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

Method

bullTo perform a radioimmunoassay a known quantity of an antigen is made radioactive frequently by labeling it with gamma-radioactive isotopes of iodine attached to tyrosine (hot)bullThis radiolabeled antigen is then mixed with a known amount of antibody for that antigen and as a result the two specifically bind to one another

bull Then a sample of serum from a patient containing an unknown quantity of that same antigen is addedbull This causes the unlabeled (or cold) antigen from the serum to compete with the radiolabeled antigen (hot) for antibody binding sites

bullAs the concentration of cold antigen is increased more of it binds to the antibody displacing the radiolabeled variant and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bull Then a sample of serum from a patient containing an unknown quantity of that same antigen is addedbull This causes the unlabeled (or cold) antigen from the serum to compete with the radiolabeled antigen (hot) for antibody binding sites

bullAs the concentration of cold antigen is increased more of it binds to the antibody displacing the radiolabeled variant and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bullAs the concentration of cold antigen is increased more of it binds to the antibody displacing the radiolabeled variant and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

The bound antigens are then separated from the unbound ones and the radioactivity of the free antigen remaining in the supernatant is measured using a gamma counter Using known standards a binding curve can then be generated which allows the amount of antigen in the patients serum to be derived

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

The principle of RIA

bull The amount of Ab per tube is kept constant the amount of antigen added (known or unknown) is the variable parameterbull The added antigen will be distributed between a bound (B) and a free (F) fraction This distribution is governed by the association constant (KA) of the Ab

Ab + Ag 1048698 AgAb and K = [AbAg] [Ab][Ag]

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bullConclusion If total Ab input is kept constant the value of BF is a measure for the total Ag input

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bullTo measure this distribution B-F bullA small but constant amount g p of labeled antigen (tracer) is added to the reactionbullEventually there will be a competition reaction between this small but constant amount of tracer and the cold antigen for a limited amount of antibody

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

Requirements for the development of an RIA

1 Pure antigen for - standards (μg)

- Tracer production (tens of μg)- Ab production (hundreds of μg)

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

2 Tracer self-made or commercial

3 Specific high-affinity antibody self-made or commercial

4 A method to separate bound and free antigen

5 (Optional) A system to extract the antigen from the sample

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

Separating Bound from Free Antigen

bull Precipitate the antigen-antibody complexes by adding a second antibody directed against the first For example if a rabbit igg is used to bind the antigen the complex can be precipitated by adding an antirabbit-igg antiserum (eG Raised by immunizing a goat with rabbit igg) This is the method shown in the diagram above

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bull The antigen-specific antibodies can be coupled to the inner walls of a test tube

After incubation bull The contents (free) are removed bull The tube is washed (bound) bull The radioactive of both is measuredbull The antigen-specific antibodies can be coupled to particles like sephadex Centrifugation of the reaction mixture separates the bound counts (in the pellet) from the free counts in the supernatant fluid

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

Radioimmunoassay is widely-used because of its great sensitivity

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bullThe greater the specificity of the antiserum the greater the specificity of the assay

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

Drawback

bullExpense and hazards of preparing and handling the radioactive antigen

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bullBoth 125I or 131I emit gamma radiation that requires special counting equipment bullThe body concentrates iodine atoms mdash radioactive or not mdash in the thyroid gland where they are incorporated in thyroxine (T4)

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

RIA as a major clinical tool

It is used to assay plasma levels of most of our hormonesbullDigitoxin or digoxin in patients receiving these drugsbullCertain abused drugsbull For the presence of hepatitis B surface antigen (hbsag) in donated bloodbullAnti-dna antibodies in systemic lupus erythematosus (SLE)

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

bull Narcotics (drug) detectionbullBlood bank screening for the hepatitis (a highly contagious condition) virusbull Early cancer detectionbullMeasurement of growth hormone levels tracking of the leukemia virusbullDiagnosis and treatment of peptic ulcers research with brain chemicals called neurotransmitters

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28

Thanks

• Radioimmuno Assay
• Radioimmunoassay
• Slide 3
• Slide 4
• Slide 5
• Radioimmunoassay pros and cons
• Slide 7
• Principle
• Method
• Slide 10
• Slide 11
• Slide 12
• The principle of RIA
• Slide 14
• Slide 15
• Slide 16
• Slide 17
• Requirements for the development of an RIA
• Slide 19
• Separating Bound from Free Antigen
• Slide 21
• Slide 22
• Slide 23
• Drawback
• Slide 25
• RIA as a major clinical tool
• Slide 27
• Slide 28