Bio Medical Tracers Particles

8/3/2019 Bio Medical Tracers Particles

1/19

Particle Tracers

Use of natural or synthetic particles astracers includes at least 4 primaryapproaches:

Use of particles to visually demonstrate

molecular aggregates Attachment of particles to molecules or

cells to mark their locations

Attachment of particles to molecules orcells to allow them to be counted

Attachment of particles to molecules or

cells to allow them to be isolated


2/19

Molecular Aggregation

Many early immunologic assays werebased on molecular agglutination,

flocculation, or precipitation of molecular

aggregates to produce macroscopicallyvisible changes in previously uniform,

clear solutions. The aggregates were

formed by antigens interacting with

polyclonal antisera, e.g., the preciptinreaction, or spot - plate testing for blood

group substances or syphilis.


3/19

Molecular/Cellular Aggregation

Extension of the idea of aggregation to amore quantitative format was provided

by microscopic examination of cellular

aggregates (rosettes) formed by washed,tannic acid treated red blood cells,

coated with antibodies interacting with

antigen molecules or antigen - containing

cells. Other versions of this used carbonparticles, bacterial cells, white blood

cells, or chemically conjugated latex

(polystyrene) spheres.


4/19

Localization with Particles

Microscopic visualization also allowedlocalization of particle tagged cells or

molecules. As diffusion is inversely related to

molecular/particle size, however, smaller

particles were used in these contexts, e.g.,noble metal colloids (5 - 150 nm), large viruses

(TMV), bacterial cells (coccus sp.), small latex

spheres, & even small atomic chelate

collections such as decagold. Locale wasoften marked as a color change (reddish with

gold colloids) rather than direct visualization of

individual particles.


5/19

Adsorption of macromolecules to colloids

Colloid definition:

Conjugation of molecules to latex or bioparticles

involves the same kinds of chemistries already

discussed for linking biomolecules to antibodies,

lectins, or similar markers.

http://en.wikipedia.org/wiki/Colloid

http://books.google.com/books?id=rlcLQmcTADEC&pg=PA1136&lpg=PA1136&dq=noble+metal+colloids&source=bl&ots=yEk7E3Lj_j&sig

=Og5QFJqeoOt8o5K3LGeo9QM0tT8&hl=en&ei=rk3USdGGBYbglQf4y

sC-DA&sa=X&oi=book_result&ct=result&resnum=3

http://www.ias.ac.in/matersci/bmsjun2000/165.pdf

Characterization & modifications,

noble metal colloids:


6/19

Localization with Particles (cont.)

Localization & counting of individual small

particles, including small viruses like lamdaphage, can be done using transmission (TEM;

resolution to 1 - 10 nm) or scanning electron

microscopy (SEM; resolution to 10 - 50 nm).

Particles or particle conjugates may beintroduced into the live organism prior to fixation

& embedding for TEM or fixation & carbon/metal

coating for SEM. They may also be used after

fixation & washing for surface structures in SEM.Or after fixation, embedding, & sectioning for

TEM following etching with alkalis or treatment

with microwave heat to unmask antigens.


7/19

HCGBinding on MA-10 Cells by SEM

Problem: villi

& particles of

similar size.

MA-10cells in

culture


8/19

TEM of various particles:

www.tanaka.co.jp/english/products/html/f_6.html

www.devicelink.com/ivdt/archive/00/03/004.html

www.ansci.wisc.edu/.../microscopy/colloid.html

www.liv.ac.uk/Chemistry/Staff/brust.html


9/19

Automated Particle Counting

Counting of particle - tagged moleculesor cells may also be done automatically

so long as the particles can be detected

by size, color, fluorescence, electricalconductivity, or magnetic properties. The

Coulter counter evaluates size & number.

The Fluorescence Activated Cell Sorter

(FACS) detects laser - activatedfluorescence (often in attached latex

beads). Imaging software tracks

contrast, color, or dimensional profiles.


10/19

The first Coulter Counter

High Speed Automatic Blood Cell

Counter and Cell Size Analyzer

Wallace H. Coulter

Coulter Electronics, Chicago,

Illinois

:1034-1042, 1956

High Speed Automatic Blood Cell

Counter and Cell Size Analyzer

Wallace H. Coulter

Coulter Electronics, Chicago,

Illinois

:1034-1042, 1956

J.Paul Robinson, Professor of Immunopharmacology, Professor ofBiomedical

Engineering, Schools ofVeterinary Medicine & Engineering, Purdue University


11/19

Automated Particle Counting

http://www.spectrex.

com/html_files2/pc2

200.php

All the FACS & nothingbut the FACS:

http://www.cyto.purdue.edu/fl

owcyt/educate/pptslide.htm


12/19

www.bio.davidson.edu/COURSES/

Bio111/FACS1.html

http://www.bio.davidson.edu/cour

ses/genomics/method/FACS.gif


13/19

Some modern FACS systems such as theBD Aria can examine as many as 9 or

even 13 different signals simultaneously.

Dye Sensed Laser - Ex/Em Laser Detector

FSC FSC Blue FSC

SSC SSC Blue F

FITC B-530/30 Blue E

PE B-576/26 Blue DPE-Texas Red B-610/20 Blue C

PE-Cy5.5 B-695/40 Blue B

PE-Cy7 B-780/60 Blue A

APC R-660/20 Red C

Alexa 680 R-710/20 Red B

APC-Cy7 R-780/60 Red A

Alexa 405 V-450/40 Violet BAlexa 430 V-530/30 Violet A

www.microbiology.emory.edu/.../p_pfc.shtml


14/19

Magnetic & latex particles from BangsLaboratories:

http://www.bangslabs.com/learning

Formation of mini-emulsionscontaining carbon or magnetic cores:

http://www.mpikg-golm.mpg.de/kc/landfester/

Microscopy of latex & latex films:

http://www.lehigh.edu/~ols0/intro.html

Magnetic particles from Seradyn:http://www.thermo.com/com/cda/landingpage/0,,2348

,00.html


15/19

Quantum Dots (Last but not least!)

QdotProteinAconjugatewithUV

http://www.invitrogen.com/site

/us/en/home/brands/Molecular-Probes/Key-Molecular-

Probes-

Products/Qdot/Technology-

Overview.html

Tunability ofQdot

output by

adjustment of

particle sizewww.immunology.utoronto.ca/..

./FlowIntro.htm

www.aist.go.jp/.../hot_line/

hot_line_22.html


16/19

www.ceac.aston.ac.uk/research/staff/as/research/


17/19

a,b, Quantumdotshavingdifferentmoleculesfortarget-specificinteraction, and, attachedtothesurface,

paramagneticlipids (a, basedondescriptionsinref.15; themoleculesareshownattachedtopartofthedot

forclarity, butextendoverthewholesurface)andchelators (b, basedondescriptionsin Refs5,18,19)for

nuclear-spinlabelling.c, Basedonthedescriptioninref.3.thesilicaspherehasQDsandparamagnetic

nanoparticlesinsideandtarget-specificgroupsattachedtotheoutside.d, ThestructureofamultimodalQDprobe, basedonsilica-shelledsingle-QDmicelles.Adaptedfromref.20, copyright (2006)ACS.

Figure 1 - Various designs of multimodal QD probes.

From the article

Designing quantum-dot

probes

Rumiana Bakalova, Zhivko

Zhelev, IchioAoki & Iwao

Kanno

Nature Photonics 1, 487 -

489 (2007)

doi:10.1038/nphoton.2007.

150

www.nature.com

/.../nphoton.2007

.150_F1.html


18/19

Quantum dot conjugates can simultaneously reveal the fine details of many cell

structures. Here, the nucleus is blue, a specific protein within the nucleus is pink,

mitochondria look yellow, microtubules are green, and actin filaments are red. Soon the

technique may be used for speedy disease diagnosis, DNA testing, or analysis of

biological samples.

QUANTUM DOT CORP., HAYWARD, CA

publications.nigms.nih.gov/.../chapter1.html


19/19

Other New Uses:

New uses for particles are modifications of

those already mentioned.

Uniform latex or gold particles can be used

to calibrate electron or visual microscopes.

Labeling of blots using latex or colloid

tagged antibodies or nucleic acids

generate colors without added enzyme

reactions; gold or silver colloid stains can

also be intensified using silver salts.

Quantum dots are semiconductor

aggregates with high optical activity &

electron density.

Date post:	06-Apr-2018
Category:	Documents
Upload:	ashok-kumar
View:	220 times
Download:	0 times

Bio Medical Tracers Particles

Documents