Biology Reference
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equilibrium state, often designated as Δ
.
33,34
A detailed treatment
of the force spectroscopy of single and multiple bond dynamics was given
by Evans and Williams.
35
In the next sections, we present early work in which cell surface
molecules were mapped using modiied AFM probes, discuss the use of the
colloidal probe method for mapping vitronectin (VN) and prostaglandin
receptors, describe measurements of the binding strength of transferin
receptors and discuss the issue of unbinding versus uprooting of membrane
proteins. We then provide a brief literature survey of recent protein
mapping studies and discuss the possibility of mapping intracellular mRNA.
x
or just
x
12.2 PIONEERING EXAMPLE OF CELL SURFACE MAPPING
As an early example of cell surface mapping, using an AFM probe
coated with a sugar-speciic lectin, concanavalin A, Gad
mapped
the presence or absence of mannan molecules over the surface of live
Saccharomyces cerevisiae
et al.
36
yeast cells on a glass surface covered with covalently immobilized
concanavalin A which is reactive with mannan on the cell surface. Cells
were adsorbed to the glass surface in a highly packed condition, which
was convenient for later mapping of mannan. After imaging closely packed
yeast cells with a bare probe, a new probe that was coated with covalently
immobilized concanavalin A was brought on top of one of the cells, and the
interaction between the probe and the cell surface was measured by the
force volume method. This AFM mode simultaneously gave the topography
and 16 × 16 force curves recorded over 3 μm × 3 μm regions. The force
curves obtained in this way showed extensions of very lexible molecules
up to 1 to a few micrometres, which was interpreted as the extension of
almost randomly coiled mannan molecules covalently attached on the yeast
cell wall. The downward delection of the cantilever as shown in
Fig. 12.1a
was considered to be due to the tensile force exerted by the lexible mannan
chains to the cantilever. The sharp upward jumps (observed three times
here) were interpreted as the unbinding events of concanavalin A from the
surface mannans. The magnitude of the jump
E
measured in nanometres
was converted to the unbinding force
F
by multiplying it with the force
constant of the cantilever
, assuming the Hookean linear behaviour of the
cantilever. The spring constant
k
was determined to be approximately 0.025
nN/nm by pushing a small cantilever cut out from a thin gold foil. Since the
nominal spring constant supplied by the manufacture was 0.02 nN/nm, the
measured value was within an allowable error range.
k
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