1. Immobilization Techniques for DNA biosensor
LECTURE OF SUBJECT:
Dr. sharafaldin Al-musawi
College of Biotecholgy
LECTURE: 6
SUBJECT: Biosensors & Biochips
LEVEL: 4
Immobilization Techniques for DNA biosensor

2. Immobilization Techniques for DNA biosensor

3. Functional group for DNA Immobilization
A functional group is a specific group of atoms within a molecule that is responsible for characteristic chemical reactions of that molecule.
There are different covalent attachment chemistries involving various functional groups on the surface of functionalization chip for DNA immobilization.
Among this factor we can refer to:
Carboxyl
Aldehyde
Epoxy
Isothiocyanate
Maleimide
Mercaptosilane

4. Carboxyl group
The carboxyl group is an organic functional group consisting of a carbon atom double bonded to an oxygen atom and single bonded to a hydroxyl group.
Another way to view it is as a carbonyl group (C=O) that has a hydroxyl group (O-H) attached to the carbon atom.
The carboxyl group is commonly written as -C(=O)OH or -COOH.

5. Reaction of carboxyl group with amine

6. The aminated DNA can Interact with immobilized carboxylate via EDC [1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide] OR Sulfo-NHS (N-hydroxysulfosuccinimide).

7. Simple method of immobilization High surface coverage of DNA’s
The advantages and disadvantages of the reaction of carboxy functionalized surface with aminated DNA
Advantages:
Simple method of immobilization
High surface coverage of DNA’s
Easy coupling reaction
Disadvantages:
Efficiency of immobilization depends on: pH
Concentration
Ionic strength
Reaction time.

8. Aldehyde group
Aldehyde: any of a class of organic compounds, in which a carbon atom shares a double bond with an oxygen atom, a single bond with a hydrogen atom, and a single bond with another atom or group of atoms.
The double bond between carbon and oxygen is characteristic of all aldehydes and is known as the carbonyl group.

9. Chemical bonding with aminated-DNA
The aminated DNA can Interact with immobilized Aldehyde group in bio reception surfaces.
DNA
Aminated DNA
Aldehyde group
Chemical bonding with aminated-DNA
+ H2O

10. Long hybridization time High binding strength
The advantages and disadvantages of the reaction of aldehyde functionalized surface with aminated DNA
Advantages:
Good stability
Long hybridization time
High binding strength
Disadvantages:
Limits the absolute signal intensity
Stable enough for long term use
High hybridization temperature

11. Epoxy group
An organic chemical that contains a group consisting of an oxygen atom bound to two already connected atoms, usually carbon.
A  functional group that containing an oxygen atom joined to two different groups that are themselves joined to other groups.

12. Epoxy groups can intract with DNA via chemical (covalent) bonding with hydroxyl, amine and sulfhydryl groups.
Double-strand DNA can be immobilized on the epoxy surface Because guanine bonds easily react with epoxy group.

13. Easy protocol for immobilization Good stability High binding strength
The advantages and disadvantages of the reaction of epoxy functionalized surface with aminated DNA
Advantages:
Easy protocol for immobilization
Good stability
High binding strength
Stable enough for long term use
Disadvantages:
Reactions between DNA and epoxy are extremely slow.
Low Immobilization density

14. Isothiocyanate
Isothiocyanate is the chemical group –N=C=S, formed by substituting the oxygen in the isocyanate group with a sulfur.

15. Isothiocyanate groups can interact with DNA via chemical (covalent) bonding with its aminated form.

16. Stable enough for long term use Disadvantages:
The advantages and disadvantages of the reaction of Isothiocyanate functionalized surface with aminated DNA
Advantages:
Well-ordered surface
Re-usability
High density DNA/area
Stable enough for long term use
Disadvantages:
High non-specific hybridizations
Long hybridization time

17. Maleimide group
Maleimide is a chemical compound with the formula H2C2(CO)2NH.

18. Maleimide group can interact with DNA via chemical (covalent) bonding with its sulfhydrylated form.

19. Faster immobilization reaction Re-usability Good stability
The advantages and disadvantages of the reaction of maleimide functionalized surface with sulfated DNA
Advantages:
Faster immobilization reaction
Re-usability
Good stability
High binding strength
Disadvantages:
High non-specific interaction
Degradation in aqueous solutions

20. Mercaptosilane group Mercaptan
any of a class of odiferous sulfur containing compounds having the type formula RSH, where R is any hydrocarbon group.
Also called thiol.

21. Mercaptosilane can perform chemical (covalent) bonding with DNA-SH.

22. Stable enough for long term use Re-usability Good stability
The advantages and disadvantages of the reaction of mercaptosilane functionalized surface with sulfated DNA
Advantages:
Stable enough for long term use
Re-usability
Good stability
High binding strength
Disadvantages:
High non-specific interaction
High hybridization temperature