The nanotechnology for life sciences, nano-biotechnology,
is the convergence of nanoscience/technology and biological
science, leading to new "eyes and hands" for
understanding and manipulating biological systems. In
this field, atomic force microscopy (AFM) has attracted
keen attentions, because it is compatible with non-conductive
materials, does not require labeling, and can be operated
under the physiological condition, while allowing single
molecular level analysis. Therefore, AFM has become
a central tool for nano-biotechnology. Also, imaging
surfaces with the nanoscale resolution, probing local
mechanical properties, and measuring a variety of interaction
forces from nN to pN are possible with the instrument.
In particular, the microscopy measuring the force of
a picoNewton range has been utilized for a variety of
single molecule measurements, and the examples include
protein unfolding process, DNA-DNA, protein-protein,
and ligand-receptor interaction. At the same time, a
significant growth of the genome and proteome studies
for drug discovery, as well as for disease diagnosis
and prevention, has placed a strong demand for advanced
biomolecular recognition probes with high sensitivity
and enhanced specificity. It is therefore clear that
this type of approach finds important applications not
only within basic life science research, but also within
emerging areas for the analysis of larger biomolecular
When the force measurement between individual biomolecules
is carried out with AFM, the accuracy and the resolution
of the force value is a critical factor for the reliability.
In order to enhance the reliability, it is necessary
to control density and orientation of the immobilized
biomolecules on surface. It used to be hard to control
small scale variations due to spatial differences in
surface topography and chemistry, and such factor is
some of the remaining bottle-necks to the future progress
of such force-based approaches. To resolve such issues,
NSB POSTECH offers a new surface modification of AFM
tips and substrates for measuring specific biomolecular
interaction forces reliably (J. Am. Chem. Soc. 2007,
9349; Adv. Mater. doi:10.1002/adma.200801323). The dendron-modification
of the surfaces can optimize the density of immobilized
biomolecules, remove steric hindrance between interacting
biomolecules, and avoid unwanted nonspecific binding
and/or the formation of multiple biomolecular complexes.
Therefore, dendron modified-AFM tip of NSB POSTECH provides
a superb option to the existing biomolecular surface
immobilization methods. We believe that thus-modified
AFM tips are ideal for studying the interactions between
DNA and DNA/RNA, ligand and protein, protein and protein,
as well as receptor on cell surface and ligand.
Figure 1. A schematic drawing of the experimental setup
employing the dendron-modified
AFM tip and substrate.
1) DNA-DNA Interaction at Single Molecule Level
(J. Am. Chem. Soc. 2007, 9349)
The dendron modified-AFM tip simplifies the force-distance
curves for the specific biomolecular interaction, and
enhances the reliability of the analysis. We confirmed
effect of the spacing provided by the dendron layer
on the force analysis by using dendrons of different
generations. In case of DNA force measurement, the use
of the 9-acid gave a narrow histogram in comparison
with the cases of the lower generation dendrons (3-acid).
It is clear that the control of the spacing between
the biomolecules is essential for the fine analysis.
Figure 2. Effect of spacing on the adhesion forces.
Distributions of adhesion forces (recorded with a retract
velocity of 0.10 ¥ìm/s) obtained for 30 mer oligonucleotide
functionalized with (a) the first generation dendron
(3-acid), (b) the second generation dendron (9-acid).
The sequence of the DNA is 5'-NH2-GCT GCT ATG GAG ACA
CGC CCT GGA ACG AAG-3', and its complementary DNA sequence
is 5'-NH2-CTT CGT TCC AGG GCG TGT CTC CAT AGC AGC-3'.
The adhesive events were observed by a chance of 50
- 80 %, and the retraction traces with a single clean
pull-off event were recorded by using a substrate and
an AFM tip with the optimal lateral spacing. The adhesive
force histograms displayed the narrow distribution in
comparison with the case other immobilization methods
Figure 3. Distribution of adhesive forces recorded
at 0.10 ¥ìms-1 for all of the fully complementary sequences.
The y-axis shows the probability of observing the specific
force for each case.
In particular, the attractive force associated with
the specific molecular binding event is observed reproducibly
during the approach mode.
Figure 4. A typical force-distance curve for the complementary
30-mer DNAs. Approach and retract traces are in red
and blue, respectively.
2) Protein-Protein Interaction at Single Molecule
3) Imaging of mRNA on a Tissue Surface (Nucleic
Acids Research doi:10.1093/nar/gkn965 (2008)).
We demonstrated successfully the feasibility of mapping
mRNA distribution in a biological tissue sample by measuring
DNA-RNA interaction forces. For this end, we utilized
a DNA probe attached to a dendron-modified AFM tip to
measure the specific adhesive force to the complementary
RNA and Pax6 mRNA of 802 bases, and to map the Pax6
mRNA distribution on the surface of sectioned mouse
Figure 5. Spatial distribution of the Pax6 mRNA in
a section of mouse embryonic tissue. VZ and CP stand
for ventricular zone and cortical plate, respectively.
4) Ultrasensitive Analytical Scanning of Biochips
5) AFM Probes tethering a Single Molecule at the
The dendron modified-AFM tip of NSB POSTECH is readily
adaptable to other biomolecular species, such as proteins,
oligopeptides, RNAs, aptamers, PNAs, etc. Therefore,
dendron modified-AFM tip of NSB POSTECH should not only
be of interest to researchers using AFM for studies
of label-free DNA hybridization, but also those utilizing
a range of force measurement approaches for basic life
sciences research, as well as researchers who are interested
in developing new methods for biomolecular screening.
NSB POSTECH recommends AFM tips of a small spring constant
for studying DNA-DNA, DNA-RNA, RNA-RNA, and protein-protein
interaction. When we modified an AFM tip from Veeco
(MSCT-AUNM), the spring constant changed from 10 pN/nm
to a value between 10 pN/nm to 16 pN/nm. NSB POSTECH
is more than happy to coat other AFM tips with the dendron
upon customer's request.
Please contact firstname.lastname@example.org
for inquiry and quotation.