Biodefense; anew option against terrorism atack

Biodefenseproject

A.D.N.

ARTIFICIAL DOG NOSE

Terrorism and biodefense

ARTIFICIAL DOG NOSE• Could we imagine the possibility of

detecting only one molecule?

ARTIFICIAL DOG NOSE

ARTIFICIAL DOG NOSE

• Narcotics• Explosives

ACTUAL SOLUTIONS

ACTUAL SOLUTIONS

Narcotics and explosives

ACTUAL SOLUTIONS

BACTERIA AND VIRUSES

ACTUAL SOLUTIONS• Up to 40 explosives• Most frequent narcotics • Up to 8 microorganisms• Viruses

• This requires quantities in NANOGRAMS

IDEA• MOLECULE, smallest AMOUNT • TO IDENTIFY ONE MOLECULE• BIOLOGICAL RECEPTOR

– BIOLOGICAL– NON BIOLOGICAL SUBSTANCES

ANTIBODY

Inmunology conceptClonal selection theory: MacFarlane Burnet 1957

1.Each lymphocyte bears a single type of receptor of unique specificity.2.Antigen interaction leads to lymphocyte activation.3.Daughter cells bear identical antigen specificity to the parent cell.

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1.RECEPTOR CONSTRUCTION

• Antibodies are a class of proteins secreted by B-Cells in response to the presence of a foreign substance or Antigen.

• Antigen: foreign substances which can combine with TCR or BCR or Ab and have the potential of inducing immune response (>10,000Dlt).

– binding interactions are: van der Waals forces, hydrophobic interactions, and H-bonds

ORGANICS RECEPTORS

• Hapten: substances which can combine with Ab, but cannot induce immune response independently. In another word, hapten only possess immunoreactivity.

• Carrier: enhance the immunogenicity of hapten, can be a protein, bacteria, cell...

ASPIRIN

ORGANICS RECEPTORS

• Hapten needs:– Covalent union

with a large protein.

– Cell’s surface (eritrocytes).

– MO’s surface.

Mechanism of Hapten-Carrier

• Hapten recognized by Ig receptor on B cell• Hapten-carrier endocytosed by B & APC• Carrier processed and presented on class II

MHC to Th cell • Activated Th & APC cell produces cytokines• Cytokines enable B cell to be activated to

produce anti-hapten antibodies

The three main types of armed effector T cell produce distinct sets of effector molecules.

THYMUS-DEPENDENT

THYMUS-INDEPENDENT

CYTOKINES

A second signal is required for B-cell activation either by thymus-dependent or by thymus-independent antigens.

B CELL

T CELL

2º INTERACTIONCYTOKINES

...or by thymus-independent antigens

APC

2º INTERACTION

HAPTEN

CARRIER

• Th, and APC cells recognize carrier, B cells recognize hapten

• Th, APC & B cells cooperate by interacting

• Interactions are class II self-MHC restricted

T Cell-B Cell Interactions(hapten-carrier effect)

CD40

Immunoglobulinreceptor

MHC II

B7B7 CD28

TCRTCRT helpercell

Antigen

1. Antigen presentation toTh cell

2. B7 expressed 3. Th cell is activated

and expresses CD40 ligand,

Cytokines secreted

CD40 ligand

Cytokine

Cytokine receptor

Bcell

Bcell

Bcell

T helpercell

4. Cytokine binds to cytokine receptor,

CD40 ligand binds to CD40

Bcell

Bcell

Bcell

5. B cell activated

6. B cells proliferate, differentiate, secrete Ig

Class II MHCAPCTh cell B

cellTh cell

Bcell

Bcell

B cell takes up and presents antigen

Th cell Th cellB

cell

Th cells are primed by antigen-presenting cell

B-T cell cooperationB cells receive signals from T cells

B cells divide

Bcell

Bcell

Bcell

Bcell

Antibody formingcell

Antibody formingcell

Antibody formingcell

Bmemory

cell

ORGANIC RECEPTORS

• Hapten needs:

ORGANIC RECEPTORS• Hapten needs:

CRYSTAL

APC

ORGANIC RECEPTORS• Hapten needs:

COCAINE HEROIN

ATROPINE HEPARINE

ORGANIC RECEPTORS

PENICILLIN

TNT

WHY NOT?TNT

ORGANIC RECEPTORS

ORGANIC RECEPTORS

Artificial-carrier

PROBABLY, THE SMALLEST INTELLIGENT CARRIER POSIBLE

ONE ANTIGENIC STRUCTURE FOR SMALL HAPTENS

THEORICAL PROCESS

CONTACT LENS

1.RECEPTOR CONSTRUCTION• MONOCLONAL ANTIBODIES

InmunizationSpecific Ag

AntibioticsAnimal serumhipoxantine

Isolation of monoclonal antibodies

Nourishing stratus

*Hammerling, o. J.; u. Hámmerling; j. F. Kearny (editores): monoclonal antibodies and t-cell hybridomas.Perspectives and tk(hnical advances. Research monog. In lmmunology vol. 3, amsterdam, elsevier/north holland biomed press, 1981

Mouse tumor

HAT medium

Ab

summary• ANTIBODIES = MOLECULAR

RECEPTORS• HAPTENS BECAME INUNOGENICS• NATURAL SELECTIONS OF

ANTIGENICS STRUCTURES• THERE IS NOT HISTOCOMPATIBILITY

PROBLEMS

• FACED WITH…. EXPLOSIVES• WITH TOXICS• WITH BACTERIA AND VIRUSES• WITH A WIDE RANGE OF MOLECULAR STRUCTURE

• QUANTITIES OF • 10 12 TIMES MORE SENSITIVE

• OF NANOGRAMS 10 -9 gr• ZEPTO O YOCTO GRAMOS 10 -23

ANTIBODIES TYPES

Biochips• Micromatrix of biologic macterial.• Massive obtention of information.• Classification• design• Clustering/data mining

Ab’s Microarrays

TYPESFluorescence image of SAM-Biotin capture membrane subsequent to antibody treatment

SAM-Membrane was incubated with FITC-labelled monoclonal anti-phosphotyrosine antibody PT-66 (Sigma, Order# F3145) (dilution 1:1000 in TBS) for 150 min followed by washings (5 times) with TBS-buffer. Fluorescence image was generated using FLA 3000 reader (Fuji, excitation at 473 nm, fluorescence filter 520 nm).

Chemoluminescence

Comparison of a microarray experiment. The red dots represent transcripts found with AB's microarray that are either absent or not detectable on Affymetrix whole genome microarrays.

CHEMOLUMINISCENCE• This micrograph of the Germanium detector shows an active area

about 30 µm wide, sandwiched between two interdigitated electrodes on top of a silicon dioxide base.

• optical micrograph of an array of GeOI photodetectors. The detectors at the top of the photograph are 100 µm wide, while those at the bottom measure just 10 µm

Ab’s Microarrays

Scanner molecular

ARTIFICIAL DOG NOSE

ARTIFICIAL DOG NOSE

ARTIFICIAL DOG NOSE• Molecular sensitivity• High specificity• Up to 360 different molecules at one

time• Independently

– Explosives– Toxics– Bacteria– viruses

ARTIFICIAL DOG NOSE

• LIMITATIONS– EXPERIMENTAL TECHNOLOGY– HIGH COST R and D– DIAGNOSIS TIME TOO LONG

“Dreams come true, without that possibility,

nature would not incite us to have them”John Updike

ORGANICS RECEPTORS