Transcript Power Point Lecture Notes

Radiotracers
• Introduction
• Design of a Radiotracer Experiment

Molecule labeled at specific location

Physical processes
• Applications and techniques
• Basic premise

Radioactive isotope behaves the same as stable isotope

Radioactive isotope easier to follow and detect
 Dilution 10-6 to 10-12

Chemistry of element monitored by isotope behavior

Trace dynamic mechanisms

Also used to evaluate isotope effect
 Slight differences in kinetics due to isotopic mass differences
• Used in biology, chemistry
4-1
Radiotracer experiments
• Basic assumptions of experiments
• radioactive isotopes behave as the stable isotope

difference in masses can cause a shift in the reaction rate or
equilibria (the isotope effect)
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in most cases isotope effect does not significantly affect
radioisotope method

Isotope effect related to square root of the masses
 Largest in small masses (i.e., H)
* Not as reliable with H, C limited in intermolecular
reactions
• radioactivity does not change the chemical and physical properties
of the experimental system

Need to consider amount of activity
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Biological effects limited in short term
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Limit physical effects (i.e., crystal damage, radicals)
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Limited impact of daughter
 Different chemical form
4-2
Radiotracer experiment
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biological studies there is no deviation from the normal physiological state
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Chemical compound level should not exceed normal concentration
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specific activity of tracer must be sufficient
 Shorted lived isotopes better
Chemical and physical form of the radionuclide compound same as
unlabeled

Need to consider sorption to surfaces or precipitation
 Radionuclide often in concentration below saturation
 Precipitates due to presence of stable isotope
radionuclide and the stable nuclide must undergo isotopic exchange
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Redox behavior and speciation
Radiochemical purity
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Activity due to single isotope
Only labeled atoms are traced
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Radioisotope due to compound not free isotope or other chemical
form
4-3
Experimental considerations
• Suitable isotope
 Half-life
 Too short difficult to use
 Too long need to much isotope
 Decay mode
 Gamma eases experiments
 Availability
 Production method
 generator
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4-5
Labeled compounds
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Specifically labeled
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labeled positions are included in name of compound
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Greater than 95% of the radioactivity at these positions.
 i.e., aldosterone-1, 2-3H implies that <95% of the tritium label
is in the 1 and 2 positions.
Uniformly labeled
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compounds labeled in all positions in a uniform pattern.
 L-valine-14C (U) implies that all carbon atoms in L-valine are
labeled with equal amounts of 14C
Nominally labeled
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some part of the label is at a specific position
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no other information on labeling at other positions
 cholestrol-7-3H (N) some tritium is at position 7, but may also
be at other positions
Generally labeled
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compounds (usually tritium) with a random labeled distribution
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Not all positions in a molecule labeled
4-6
Synthesis
• Labeled compounds include
14C
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3H
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• Carbon
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Need to consider organic reactions for labeling
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Biosynthesis
 Photosynthetic
 Microbial
• Hydrogen
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reduction of unsaturated precursors
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Exchange reactions
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Gas reactions
4-7
Physical processes
• Location in a system
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Precipitation, sorption
 Measure change in
solution
concentration

Separations
 Ratio of isotope in
the separation
process
* Ion exchange,
solvent
extraction
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Reaction mechanisms
 Intermediate
reaction molecules
 Molecular
rearrangements
4-8
Isotope effects
• Based on kinetic differences or equilibrium differences
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0.5 mv2
 Mass is different
• Distillation
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Mass difference drives different behavior
• Effects can be seen approaching equilibrium
• Kinetic isotope effects are very important in the study of chemical
reaction mechanisms
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substitution of a labeled atom for an unlabeled one in a
molecule causes change in reaction rate for Z < 10
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change can be used to deduce the reaction mechanism
• change in reaction rate due to changes in the masses of the
reacting species due to differences in vibrational frequency along
reaction coordinate in transition state or activated complex
• Experimentally straightforward to measure the existence and
magnitude of kinetic isotope effects
4-9
Biological experiments
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Autoradiography
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oldest method
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radioactive sample is placed on photographic emulsion
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After period of time film is developed
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precise location of the radioactive matter in sample is found
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autoradiography used to locate radionuclides in a sample or chromatogram
Radioimmunoassay (RIA)
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sensitive method of molecules in biological samples
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based on the immunological reaction of antibodies and antigens
 antigen or antibody labeled with a radiotracer
 limited amount of antibody is available, antigen will compete for
binding sites
 Start with a certain amount of radiolabeled antigen, any additional
antigen added will displace some the radiolabeled antigen
 Measure activity of the supernatant
* amount of unbound antigen
 mix the same amounts of antibody and radiolabeled antigen together
with unknown stable antigen sample
 stable antigen will compete with the radiolabeled antigen for binding
sites on the antibody molecules.
Some of the radiolabeled antigen will not be able to bind
constructing a calibration curve that shows the amount of radioactivity present in
the supernatant after adding standard
4-10
Biological experiments
• DNA analysis
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extract the DNA from a sample
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DNA is cut into pieces using enzymes that cut either side of a
repeated sequence
 DNA mixture of segments of differing size
 Electrophoresis is used to sort the fragments by size
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spatially separated fragments are allowed to react with
radiolabeled gene probes
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gene probes contain radiolabeled specific DNA fragments of
DNA bind only to DNA segments containing a nucleotide
sequence that is complementary to its own (matching strand
in the DNA double helix
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original DNA fragments identified by the radiolabeled DNA
that has reacted
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physical pattern the autoradiograph is pattern of the DNA
sequences and sizes
4-11
Environmental and industrial
• Environmental processes
 Flow
 Dispersion
 In atmosphere
and hydrosphere
 Short lived isotopes
 Isolated from
other systems
4-12
4-13
Industrial uses of Radiation
• Radiation
 Imaging
 Density
 Analysis
 Curing
Requires source, detector, data analysis, and
shielding
4-14
Measurement with neutrons and
photons
• Radiography
• Tomography
• Density
 Tracers in wells
 Am/Be source (1 Ci to 0.1 Ci)
 137Cs (around 1 Ci)
• Used in determining
 flow
- industrial production
 moisture content
-airplane maintenance
 images
4-15
Uses in Medicine
• Radiology
 anatomical structure (x-rays)
• Nuclear Medicine
 analyze function
 therapy
• MRI
 1H, 13C, 17O
Equipment
• Detectors
 gamma
 coordinated to produce images
• Isotopes
 Need to produce and purify
4-16
Isotope Production
• Reactor produced
 n,g reaction
• Cyclotron produced
 p,x reactions
 PET radionuclides
• Generators
 long lived parent, short lived daughter
(99mTc from 99Mo)
 Ion exchange holds parent, daughter is eluted
• Natural
4-17
 212Bi from natural decay chain
Tools for Nuclear Medicine
• Hot Atom Chemistry
 formation of different molecule upon decay or
production
• Organic chemistry
 synthesis of labeled compounds
MoAb with ligand
complex which can pass through barriers
complex similar to biological molecule
 must be biologically active
• Medical
 metabolism
 diagnosis
 therapy
4-18
Isotope
51Cr
59Fe
67Ga
75Se
99mTc
111In
123I
131I
133Xe
186Re
205Tl
Isotopes
Half-life
27.7 days
44.5 days
78.3 hours
119.8 days
6.02 hours
67.3 hours
13.2 hours
8.05 days
5.25 days
89.3 hours
73.5 hours
Use
blood and spleen scan
Fe metabolism
tumors and infections
pancreatic scanning
many uses
blood, bone
thyroid
thyroid
lung
bone pain
blood, heart
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External Sources
• X-rays
 oldest use discovered in 1895
travel through soft tissue, attenuated by bone
 barium as contrast media
 tomography
Computerized axial tomography
• Radiotherapy
 kill tumor from outside
 intersection of a few beams
4-20
Diagnostic Nuclear Medicine
• Obtaining medical images
 gamma rays can be used to produce image
1st used with thyroid with 131I (fission product, half-life
of 8 days)
Measure of uptake and metabolic activity
observed for hours (dose to high 3 rads/µCi, 1-10 µCi)
• Need to have isotope accumulate in a specific organ
• Spatial pattern of emissions gives a 3-D picture
 Collimated detector needed
 single energy g best for collimator
99mTc (140 keV)
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Positron Emission Tomography
• ß+ produces two 511 keV g
• Identify line where decay occurred
• Possible to reconstruct distribution
• Useful isotopes include:
Isotope
Half-life
15O
2 minutes
13N
10 minutes
11C
20 minutes
18F
110 minutes
• PET shows dynamic events
 blood flow
 respiration (lung to brain)
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Therapeutic Nuclear Medicine
• Uses ionizing radiation to kill tissue
 radical production
• Oxygen effect
 O2 has a large electron affinity
O2 + e- --> O2• High LET
 alpha particles
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Clinical Applications
• Endocrine System
 Thyroid
• Central Nervous System
 Brain
 Eye
• Musculoskeletal System
• Gastrointestinal System
 Stomach
 Pancreas
• Cardiovascular System
 Dynamics
- Adrenals
- CFS
- Intestines
- Liver
-Disease
4-24
More clinical applications
• Urinary system
• Hematopoietic system (Blood)
 First done by Lawrence in 1938 on leukemia
• Lymphatic system
• Tumors
4-25
Thyroid
Anterior and posterior images
from whole body I-131
scintigram
30 mCi I-131 (sodium iodide)
600 rad to lung
imaging for papillary
carcinoma of the
thyroid
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Thyroid
papillary
carcinoma of
the thyroid
status post total
thyroidectomy
200 mCi I-131
sodium iodide
Dose > 30 mCi
requires
hospitalization4-27
Brain
• 20 mCi Tc-99m DTPA
• No activity
4-28
Brain
• 20 mCi
Tc-99m
DTPA
• Brain
Activity
4-29
Skeletal
• 18.2 mCi Tc-99m
MDP
• Only bone uptake,
should have soft
tissue, bladder and
renal uptake
4-30
Skeletal
•Tc-99m MDP (Bone Study)
•In-111 labeled White Blood Cells (Sickle
cell)
No spleen uptake seen
•Tc-99m Sulfur Colloid (Marrow uptake)
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Skeletal and Soft tissue
• Tc-99m
pyrophosphate
• Electrical injury
4-32
Skeletal, error
• Tc-99m DTPA and Tc99m MDP
• The outer package was
labeled MDP, but was
really DTPA
• MDP is
• methylenediphosphon
ate
(contains C-P-C bonds)
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Liver
• 5.2 mCi Tc-99m sulfur colloid i.v. (SPECT)
• 1.8 rad to liver, 0.1 rad to whole body
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Lung
• Xe-133
ventilatio
n image
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Lung
• 4.2 mCi Tc-99m MAA i.v. and 10.4 mCi
Xe-133 gas by inhalation
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Tumor
• 15 mCi F-18 fluorodeoxyglucose (FDG)
• 0.59 rad whole body
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Tumor
14.8 mCi F-18
fluorodeoxyglucos
e i.v
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Tumor
• 11.0 mCi F-18 fluorodeoxyglucose (FDG) i.v
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Tumor
• 10.8 mCi F-18 fluorodeoxyglucose i.v.
4-40
Isotope dilution analysis
• quantitative analysis based on measurement of isotopic abundance of a
nuclide after isotope dilution
• Direct dilution
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determine the amount of some inactive material in a system
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define unknown amount as x grams
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To the system with x grams of inactive A, add y grams of active
material A* of known activity D
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know the specific activity of the added active material, S1
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Change specific activity
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basic equation of direct isotope dilution analysis
unknown amount x of material A given in terms of amount y of
added labeled material A* and the two measured specific activities
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S1 and S2
Example
• A protein hydrolysate is to be assayed for aspartic acid
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5.0 mg of aspartic acid, having a specific activity of 0.46 Ci/mg
is added to hydrolysate
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From the hydrolysate, 0.21 mg of highly purified aspartic acid,
having a specific activity of 0.01 Ci/mg, can be isolated
• How much aspartic acid was present in the original hydrolysate?
• We say that
• x=number of mg aspartic acid in original hydrolysate
• y=5.0 mg
• S1= 0.46 Ci/mg
• S2=0.01 Ci/mg
4-42
Inverse IDA
• simple variant on the basic direct IDA
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inverse IDA measure the change in specific activity of an unknown
radioactive material A* after diluting it with inactive A
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assume have q mg (where q is unknown) of a radioactive substance
A* whose specific activity is known
 (i.e., Sq=D/q)
 (Sq can be measured by isolating a small portion of A*,
weighing it, and measuring its activity)
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add r mg of inactive A to A* and thoroughly mix the A and A
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isolate and purify the mixture and measure its specific activity Sr.
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Sr=D/(q+r)
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