Surface plasmon optical characterization of lipid monolayers at 5 μm lateral resolution

W. Hickel and W. Knoll

Citation: Journal of Applied Physics 67, 3572 (1990); doi: 10.1063/1.345307

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 Surface plasmon optical characterization of lipid monolayers
at 5 J.tm lateral resolution
W. Hickel and W. Knoll
Max-Planck-Institutfur Polymeiforschung, Ackermannweg 10, D-6500 Mainz,
Federal Republic of Germany
(Received 11 August 1989; accepted for publication 18 December 1989)
Surface plasmon microscopic pictures recorded as a function of the angle of incidence of the
coupling laser beam can be quantitatively evaluated and yield optical thicknesses of ultrathin
coatings with an areal resolution of 5 X 5 fLm 2 • This is demonstrated for a lipid monolayer
transferred to a solid support in its phase transition region. The coexisting amorphous and
crystalline domains can be analyzed simultaneously.

I. INTRODUCTION properties of thin-film samples is possible with a lateral reso-

Plasmon surface polaritons (surface plasmons or PSP lution of better than 5 pm. We show this for a lipid mono-
for short) are bound, nonradiative electromagnetic modes layer of DMPA transferred to a solid substrate by the Lang-
associated with a polarization charge wave propagating muir-Blodgett dipping technique. 14
along a metal-dielectric interface. I Their field intensity
peaks at the metal surface and decays exponentially normal II. EXPERIMENT
to the interface both into the metal and into the dielectric. 2 The schematic setup for SPM is shown in Fig. 2. Light
Due to dissipative and radiative loses these modes are also from a HeN e laser (2 m W at AL = 633 nm) is coupled to the
damped along their propagation direction. 3 .4 PSPs have surface plasmon mode at the Cr/Au layer surface (thick-
gained popularity in the past years owing to their field en-
hancement contributions in various surface spectroscopic
studies of adsorbates and thin coatings.s Recently, we could
40
show that these waves, in a very general way, can be consid-
ered as surface-specific "light,,6 showing all the optical phe- (a)
nomena that can be found in optics textbooks for plane elec-
tromagnetic waves: diffraction of PSP by a dielectric phase ~t"30r
grating, 7 interferometry between PSP and the driving pho- DMPA
ton field,s and surface plasmon microscopy (SPM)9 were
reported examples for this. The latter technique was shown lOr
to be particularly well suited for the imaging onow contrast
samples such as lipid monolayers. 10
These quasi-two-dimensional systems are well known to
10
+
undergo, when spread and compressed at the water-air inter- 0.5 0,6_ _ 0,7 0.8
A
face, a first-order phase transition from a liquid-expanded to
nm 2/Lipid
a condensed state. II A typical pressure area, 1T-A isotherm
(the equivalent to a three-dimensional van-der-Waals p-V
curve) for dimyristoyl phosphatidic acid (DMPA) on pure
H 20 is given in Fig. 1(a) and shows the broad coexistence
range of these two phases. If a suitable fluorescence dye is
added in small concentrations to the lipid layer, this coexis-
tence can be fluorescence microscopically observed 12, 13: Be-
cause of its low solubility in the condensed phase, the dye is
squeezed out of the growing crystalline domains which
hence appear dark in the microscopic picture. This is shown
in Fig. 1(b). In fact, this technique was a major break-
through in monolayer research as far as details of the crystal
growth and morphology were of interest. It does not enable
one, however, to further characterize the (optical) proper- FIG. l. (a) Pressure-area ('IT-A) diagram ofDMPA spread and compressed
ties of the two coexisting phases. at the water-air interface. The arrow marks the lateral density at which the
In the present letter we demonstrate that SPM (i) does monolayer was later transferred to a solid substrate. (b) Fluorescence mi-
croscopic picture taken at the water surface from a DMPA monolayer in the
not need an extra label as fluorescence probe, (ii) that it phasdransition range, doped with 0.5 mol % of a fluorescence dye (NBD-
allows for more than just imaging of low-contrast samples PEl. The label is squeezed out of the crystalline domains which hence ap-
and that instead (iii) quantitative analysis of the optical pear dark.

3572 J. App!. Phys. 67 (8), 15 April 1990 0021-8979/90/083572-04$03.00 @ 1990 American Institute of Physics 3572

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Screen ICamela
Lens
' - - - - - Prism
Cooting
Evanucent
Field 01 PSP
FJG. 2. Schematic of the SPM setup.
neses 5.5 and 41.0 nm, respectively) evaporated onto the
base of a 90° BK-7 glass prism in the Kretschmann configu-
ration. ls This combination of materials, thicknesses, and la-
ser wavelength ensured a lateral resolution of < 5 .urn by
reducing the propagation length of the PSP corresponding-
ly.16 The plasmons that were scattered by the sample and
coupled out via the prism were collected (Fourier-backcon-
verted) by a lens if = 2.5 em) to form an image of the inter-
face. This could be recorded by a TV camera (CeD) and
stored on a magnetic tape for later analysis, By evaporating
Si02 (d = 18 nm) onto the Au surface a clean hydrophilic
substrate was obtained. This was mounted in the Langmuir
trough slightly below the lipid monolayer at the water air
interface in a near horizontal position. After the DMPA film
had been compressed to the coexistence range [see arrow in
Fig. 1(a)] the substrate was withdrawn by a mechanical
sample lift and the monolayer thereby transferred in a so-
called "head down" configuration, 1 I
Fresnel fits (full lines) calculated with literature values for
the complex dielectric functions of the two metals. 17,18 From
the shift of the resonance induced by the monolayer relative
to the bare substrate (.6.e = OS) one obtains an average
thickness of the monomolecular film of do = 1.45 nm assum-
ing n = 1.55 as index of refraction. This agrees with ellipso-
metric data reported earlier. 19
Now, Fig. 4 shows various pictures of the monolayer
taken by SPM at different angles of incidence, e, close to the
minimum of the reflected intensity (see Fig. 3). Each picture
shows about 410 X 350 ,urn. 2 One can clearly see that as one
tunes into the resonance the light intensity in general de-
creases but at the same time the contrast, i.e., the intensity
difference between different phases at first increases because
the amorphous (formerly liquid-expanded) areas darken
faster than the crystalline dentridic domains. On further in-
creasing e the contrast vanishes and then inverts, i.e., now
the crystallites are darker than the amorphous domains.
Eventually, at again increasing light intensity, the contrast
slowly goes to zero for stilI larger angles. The residual inter-
ference pattern observable in the brighter pictures arises
from insufficient spatial filtering of the laser beam.
The different reflectivity behavior of the two phases can
be quantitatively evaluated by a simple image-analyzing pro-
gram that calculates intensity histograms of the pixels in a
well-defined area. This is shown in Fig. 5 for two rectangular
frames located in different areas corresponding to the two
coexisting phases [see thick arrows in Figs. 5(b) and (c)].
The histograms given above, (a), for an amorphous area and
below, Cd), for a crystalline domain demonstrate that it is
indeed possible to derive a quantitative measure of the aver-
age intensity reflected from areas as sman as 5 X 5 f1m 2 as a

m. RESULTS AND DISCUSSION

Prior to SPM pictures the usual reflectivity versus angle
of incidence (9-29) scans were performed for the bare sub-
1.6.30 0 1.9.00 0
strate (BK-7/Cr/ Au/Si0 2 ) and for the coated regions. The
data points are given in Fig. 3 together with the respective

;:.; . ".~:>::"~" :::. ~~r: .,'~

~6.600 1.8.10°
0~
..... (..~' ...
J?: :........... ...... ...·x

'"c
QJ 50
C
'0 1.6.90° ~ 7.80°
1!!
u
QJ

"ai'-

Q
1,5 50 S5 [,7.50"
external angle/deg

FIG. 3. Refiected intensity vs external angle of incidence, e. of the bare

substrate (-8--) and coated with a monolayer (-0-). Fun lines are
Fresnel fits with prism: €BK7 = 2.3; Cr:€c, = - 6.3 + i 9.9, d = 5.5 mn; FIG, 4. Series ofSPM picturt,'S taken at various angles of incidence, as indi-
AU:EAu = - 12.4 + i 1.43; d = 41,0 lim; Si02 :0: = 2.13, d = 18.1 urn. cated. Each frame represents an area of 410X 350 pm!. No electronic con-
DMP A monolayer:E = 2.40; d = 1.45 nm. trast enhancement was applied,

3573 J. Appl. Phys., Vol. 67, No. S, 15 April 1990 W. Hickel and W, Knoll 3573

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 function of the angle of incidence. For better clarity the 100
frames in Fig. 5 were chosen to be - 20X 20 11m2. No sub-
stantial increase in the noise of the histogram was found, oj
however, for smaller areas. The mean intensity values on an *.....
>-
8-bit gray scale properly rescaled in order to account for III
C
camera nonlinearities and background illumination (both 'E
OJ

measured separately) are plotted in Fig. 6(a) as a function 50

of e and may be compared directly with the data of Fig. 3.
As expected, the resonance curve for the amorphous areas is
-
"0
OJ
U
w

less shifted than for the crystalline domains, the macroscopi-

fi
cally (laser spot size) averaged curve (b) in Fig. 3 being in
between. The Fresnel fits are based on the substrate data of
0
Fig. 3(a) and allow now for the first time for a separate but 20
simultaneous analysis of the two coexisting phases. Given ~
.....
the thickness of the monolayer in the condensed phase from Vi
Cl 0
synchrotron x-ray experiments as d, = 2.25 nrn (Ref. 20) '-
'E
a
we obtain for its index of refraction n 1 = 1.51. The thickness u

of the fluid/amorphous areas are less well defined in the x- -20

ray reflectivity measurement on the water surface but may
be estimated to be d 2 = 1.65 nm. 21 We then obtain for the
index of refraction of the expanded areas n 2 = 1.304. This is
a surprisingly low value but may be compared with a similar-
ly low value of the electron density for the expanded phase
found by x-ray studies. 21 It may point, on the other hand, toa
slight expansion of these areas during the transfer of the
monolayer to the solid support.
In passing, we note that a recently developed technique
for studying monolayers on a thin (15 nm) water film pre-
pared on a solid substrate22 can be combined with SPM to
allow for a microscopic characterization oflipid monolayers
at various degrees of condensation, over the full coexistence
range of the phase transition.
As a quantitative measure of the contrast that is achie-
vable we have plotted the difference of the two Fresnel
curves in Fig. 6(b). As expected for an asymmetric PSP
45 50 55
external ang le /deg
FIG. 6. (a) Reflected intensity vs external angle of incidence obtained by
quantitative analysis of SPM pictures as outlined in Fig. 5 for amorphous
( - 0 - ) and crystalline (----8---) areas of a DMP A monolayer. Solid lines
are Fresnel fits with the substrate parameters taken from Fig. 3 and
d, = 2.25 nrn, n , ,~ 1.51 for the crystalline and d 2 = 1.65 nm, n 2 = 1.304
for the amorphous regions, respectively. (b) Contrast [""difference oftbe
twoi'it curves shown in (a) 1achievable in SPM for the imaging of the coex-
istence of amorphous and crystalline domains of aD MP A monolayer in the
phase transition range.
resonance curve like for Cr/Au substrates, the contrast is
higher on the low-angle side (cf. also Fig. 4). Although the
absolute numbers for this contrast are relatively low because
we had increased the loss (i.e., the width) of the PSP reso-
nance in order to optimize the lateral resolution, one never-
theless can see that even a 5% contrast yields decent pic-
tures. (We should note that no electronic contrast
enhancement was applied to the pictures in Fig. 4).

FIG. 5. One the quantitative
evaluation of the reflected in-
tensity from areas of order
20 X 20 p.m 2 at a given cou-
pling anle 8, The SPM pic-
tUfC taken at 46.9' (b) and
(c), (ef Fig. 4) was locally
analyzed by calculating the
pixel intensity histograms of
preset frames. (a) fEsto-
grams obtained from the rec-
tangular frame (27X 18
p.m') placed in the (dark)
amorphous regions of the
imaged monolayer [see (b) 1;
(d) Same as (a) butthistime
a quadratic frame (20X 20
ftm 2 ) was placed into a crys-
talline domain ! sec (c) J.
IV. CONCLUSIONS
In conclusion, we have shown that SPM can be used to
characterize heterogeneous thin-film coatings with an areal
resolution of about 5 X 5 pm 2 • This was demonstrated for a
lipid monolayer whose coexisting condensed and expanded
domains were imaged and simultaneously analyzed.
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3574 J. Appl. Phys., Vol. 67, No.8, 15 April 1990 W. Hickel a.nd W. Knoll 3574

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3575 J. AppL Phys., Vol. 67, No.8, 15 April 1990 W. Hickel and W. Knoll 3575

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