Surface plasmon resonance assay for chloramphenicol

Article abstract of DOI:10.1021/ac801301p

We report a rapid and ultrasensitive surface plasmon resonance (SPR) assay of chloramphenicol (CAP) by using large gold nanoparticles (40 nm) for signal enhancement on a mixed self-assembled monolayer (mSAM) sensor surface. After immobilization of the target antibiotic CAP through its ovalbumin (OVA) conjugates with an oligoethylene glycol (OEG) linker on the mSAM surface, sequential binding of anti-CAP antibody and IgG/nanogold (40 nm) onto the sensor surface afforded a rapid (< 10 min) and ultrasensitive assay format for CAP. A limit of detection (LOD) for CAP as low as 0.74 fg/mL was achieved in aqueous buffer, and the linear working range was between 1-1000 fg/mL. While the LOD of CAP in a honey spiked-specimen is 17.5 fg/mL, the detection range is 80-5000 fg/mL. The mSAM sensor surface was also shown to be highly stable with over 400 binding/regeneration cycles performed.

Full text of DOI:10.1021/ac801301p

Anal. Chem. 2008, 80, 8329–8333
Surface Plasmon Resonance Assay for
Chloramphenicol
Jing Yuan,†,‡ Richard Oliver,† Marie-Isabel Aguilar,‡ and Yinqiu Wu*,†
Biosensors and Biomeasurement, The Horticulture and Food Research Institute of New Zealand, HortResearch
Ruakura, Private Bag 3123, Waikato Mail Centre, Hamilton, New Zealand, and Department of Biochemistry and
Molecular Biology, Monash University, Clayton, Australia
We report a rapid and ultrasensitive surface plasmon
resonance (SPR) assay of chloramphenicol (CAP) by using
large gold nanoparticles (40 nm) for signal enhancement
on a mixed self-assembled monolayer (mSAM) sensor
surface. After immobilization of the target antibiotic CAP
through its ovalbumin (OVA) conjugates with an oligoet-
hylene glycol (OEG) linker on the mSAM surface, sequen-
tial binding of anti-CAP antibody and IgG/nanogold (40
nm) onto the sensor surface afforded a rapid (<10 min)
and ultrasensitive assay format for CAP. A limit of detec-
tion (LOD) for CAP as low as 0.74 fg/mL was achieved
in aqueous buffer, and the linear working range was
between 1-1000 fg/mL. While the LOD of CAP in a
honey spiked-specimen is 17.5 fg/mL, the detection range
is 80-5000 fg/mL. The mSAM sensor surface was also
shown to be highly stable with over 400 binding/
regeneration cycles performed.
these ultrasensitive assays, rapid diffusion immunoassay in a
T-sensor3 or in a microchannel using a concentration gradient
format4 can measure small molecules in less than 1 or 10 min
but the assay sensitivity can only reach the subnanomolar level,
which is far less sensitive than an LOD value possible with the
LPCR or a noncompetitive immuno-enzymometric assay.1,2 It has
therefore been a significant challenge to simultaneously achieve
both ultrasensitive and rapid immunoassays of small molecules.
Here, we report a surface plasmon resonance (SPR) assay,
using large gold nanoparticles (40 nm), which by their proximity
to the sensor surface resulted in a significant enhancement of the
SPR signal to create a fast (<10 min) but extremely sensitive (LOD
of 0.74 fg/mL), yet simple, cheap, and stable SPR assay format
for chloramphenicol (CAP), a model analyte of small molecules.
EXPERIMENTAL SECTION
Reagents and Instrumentation. Mouse antichloramphenicol
antibody (mAb) was supplied by Biodesign. IgG/nanogold (40
nm) was obtained from British Biocell International (BBI). IgG/
nanogold (10 nm), chloramphenicol, and its succinate sodium salt,
ovalbumin (OVA), bovine serum albumin (BSA), succinic anhy-
dride, 11-mercaptoundecanol (11-MUOH), 16-mercaptohexade-
canoic acid (16-MHA), triethylamine, and trioxadecanediamine
were purchased from Sigma (St. Louis, MO). N, N′-Dicyclohexyl-
carbodiimide (DCC) was from Lancaster (Morecambe, England).
N-Hydroxy-succinamide (NHS) was supplied by ICN (Aurora,
OH). Polyethylene glycol (PEG)-400 was from Prolabo (Founteray
sous bois, France). Anhydrous magnesium sulfate, phosphate-
buffered saline with 0.05% Tween 20 (PBS/T, pH7.4) was from
Fluka (Buchs, Germany).
The amine coupling kit (0.1 M NHS, 0.4 M EDC, and 1 M
ethanolamine), HBS buffer, SIA kit Au, and the Biacore Q were
supplied by Biacore AB (Uppsala, Sweden). The NMR spectra
were produced on a 400 MHz Bruker Avance DRX400 NMR
(Rheinstetten, Germany). Electrospray mass spectra (ES-MS)
were generated using a VG Platform II instrument (Beverley).
Spectra of matrix-assisted laser desorption/ionization mass spec-
trometry with time-of-flight (MALDI-TOF) were recorded on a
Bruker Daltonics Autoflex spectrometer.
The ability to both rapidly and sensitively detect trace amount
of small molecules (<1000 Da) is important in the area of food
safety screening for drug residues, detection of biological toxins,
and environmental applications. For biomedical analysis, it is also
critical in patient monitoring for those small molecular weight
drugs with rapid pharmacokinetics and narrow therapeutic ranges.
Recently, several novel bioanalytical methods have been
developed for ultrasensitive detection of small molecules such as
a liposome-polymerase chain reaction (LPCR) assay using lipo-
somes with encapsulated DNA reporters as a detecting reagent.1
Sensitivity of the LPCR assay for detection of a biotoxin or cholera
toxin ꢀ subunit (CTBS) was achieved to a limit of detection (LOD)
of 0.02 fg/mL.1 Another highly sensitive approach is a noncom-
petitive immunoenzymometric assay that can detect small mol-
ecules such as 11-deoxycortisol with attomole-range sensitivity
by employing single-chain Fv fragments of antibody (scFv)-enzyme
fusion protein and anti-idiotype antibodies.2 Although these two
novel methods provided extraordinarily low detection limits for
small molecules, the assay procedures were slow and complicated
with many reagents and multiple steps involved. In contrast to
* Corresponding author. Dr. Yinqiu Wu, Phone: +64-7-959 4498. Fax: +64-
7-959 4431. E-mail: ywu@hortresearch.co.nz.
† The Horticulture and Food Research Institute of New Zealand.
‡ Monash University.
(1) Mason, J. T.; Xu, L.; Sheng, Z.; O’Leary, T. J. Nat. Biotechnol. 2006, 24,
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Goto, J. Anal. Chem. 2006, 78, 2244–2253
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Weigl, B. H.; Yager, P. Nat. Biotechnol. 2001, 19, 461–465
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10.1021/ac801301p CCC: $40.75 2008 American Chemical Society
Published on Web 10/07/2008
Analytical Chemistry, Vol. 80, No. 21, November 1, 2008 8329

Scheme 1. Synthesis of
Chloramphenicol-Olbumin Conjugate
(CAP-OEG-OVA)a
a Conditions: (a) DCC/NHS/DMF, hetero-bifunctional OEG
linker 2, room temperature, overnight; (b) DCC/NHS/DMF, OVA
in phosphate buffer, 4 °C, overnight.
Preparation of CAP-OEG-OVA Conjugate (Scheme 1).
CAP-OEG-OVA 4 was prepared according to a previous method
with some modifications.5,6 Chloramphenicol succinate sodium
salt (200 mg) was dissolved in 5 mL of deionized water.
Chloroform (5 mL) was added, and the aqueous phase was
acidified with hydrochloric acid (2 M) while stirring. The aqueous
phase was then removed, and methanol (1 mL) was added to aid
dissolution. The organic phase was dried by passage through
anhydrous magnesium sulfate to yield chloramphenicol succinic
acid 1 (177 mg, 93%).
Figure 1. Rapid SPR assay of CAP (i). On CAP-OEG-OVA/mSAM
surface (a), mAb binding to the sensor surface (ii). IgG/nanogold (10
or 40 nm) binding on the sensor surface for signal enhancement (b
and iii). Au nanoparticles removed from surface (b) to return to (a)
after regeneration.
The chloramphenicol succinic acid 1 (0.095 mmol) was
dissolved in DMF (0.5 mL). A solution of N, N′-dicyclohexyl-
carbodiimide (0.143 mmol) and N-hydroxy-succinamide (0.143
mmol) in DMF (0.8 mL) was added. The reaction mixture was
stirred at room temperature for 7 h, followed by the addition of a
heterobifunctional OEG linker 26 (0.12 mmol) in chloroform (1
mL) and triethylamine (0.5 mL). After stirring the reaction
overnight, the solvent was removed and the sample redissolved
in methanol (1 mL) and water (10 mL). The resulting solution
was then acidified with dilute hydrochloric acid and extracted with
chloroform (30 mL). After removing solvents, the residue was
chromatographed on silica gel using CH2Cl2/CH3OH (8:2) to yield
CAP-OEG-CO2H 3 (35 mg, 50%). 1H NMR (400 MHz, CD3OD) δ
(ppm): 1.79 (t, J ≈ 6 Hz, 4H, 2OCH2CH2CH2NHCO), 2.53 and
2.65 (2m, 8H, 2OCCH2CH2CO), 3.32 (m, 4H, 2OCNHCH2), 3.58
(t, J ≈ 6 Hz, 4H, 2OCNHCH2CH2CH2O), 3.65 and 3.69 (2m, 8H,
2OCH2CH2O), 4.27 and 4.45 (2m, 3H, OCNHCHCH2OCO), 5.14
(d, 1H, HOCH), 6.30 (s, 1H, CHCl2), 7.71 (d, J ≈ 8.4 Hz, 2H,
aromatic), 8.22 (d, J ≈ 8.4 Hz, 2H, aromatic). ESI-MS (negative
mode, m/z) for CAP-OEG-CO2H 3: 723 [M-H]+.
CAP-OEG-CO2H 3 (0.048 mmol) was dissolved in a solution
of DCC (0.1625 mmol) and NHS (0.1625 mmol) in DMF (0.384
mL), and the reaction was stirred at room temperature for 3 h
until a white precipitate was formed. The solution was then added
to OVA (0.96 µmol) in phosphate buffer (3.84 mL, 0.2 M, pH 7.4)
and stirred at 4 °C overnight. The conjugate was dialyzed against
deionized water for 2 days (3 changes per day) and PBS/T for
8 h (2 changes) at 4 °C. Finally, the conjugate 4 (2.5 mL) was
further purified by passing it through a PD-10 column using
PBS/T as the eluent, and the purified conjugate (3.5 mL) was
collected. The protein concentration of CAP-OEG-OVA 4 was
determined by a bicinchonimic acid protein assay, and its
conjugation degree was measured by MALDI-TOF.
Immobilization of CAP-OEG-OVA onto mSAM Surface
(Figure 1). The preparation of the mSAM and the immobilized
procedures have been described previously.6 A mixture of 11-
MUOH and 16-MHA (10 mmol, 9:1) was deposited on a bare gold
chip (SIA kit Au), and then, the CAP-OEG-OVA 4 (500 µg/mL in
10 mM sodium acetate pH 4.0) was immobilized on the mSAM
surface. The immobilization degree of ligands in a Biacore SPR
system is reported as refractive units (RU). One RU corresponds
to a shift in the resonance angle of approximately 0.1 mdeg and
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2007, 23, 144–148
.
.
8330 Analytical Chemistry, Vol. 80, No. 21, November 1, 2008

the adsorption of 1 pg of material/m2.7 A reference flow cell was
constructed by immobilization of progesterone-OEG-OVA.6
Binding Signal Enhancements and Competitive Assay of
CAP (Figure 1). Signal enhancement by IgG/nanogold particles
was determined by injecting a fixed amount of mAb solution (60
µL, 20 µL/min) over the immobilized surface, following sequential
injection of IgG/nanogold (40 µL, 20 µL/min) at various dilution
ratios (see Supporting Information). The IgG/nanogold enhance-
ment curves were produced by varying the concentration of mAb
in HBS (0-1 µg/mL) but keeping the Au nanoparticles diluted
to a ratio of 1:2 (10 nm IgG/nanogold), and 1:10 (40 nm IgG/
nanogold). Regeneration was performed with two pulses of 10 mM
glycine (pH 2.0, 20 µL/min).
Honey spiked-CAP samples were prepared according to a
literature reference8 with some modification. In briefly, 5.0 ± 0.02 g
of New Zealand honey was weighed into a plastic centrifuge tube
and mixed with 9 mL of PBS/T buffer (pH 7.4) until it went into
solution. Then the samples were spiked with 1 mL of the CAP
working standard solution or HBS buffer for the blank and
incubated 5 min at room temperature. A volume of 12 mL of ethyl
acetate was added, and the sample was mixed for 30 s with a
vortex and then shaken for 30 min on a rolling mixer. Following
centrifugation at 3200 rpm for 15 min, 5 mL of the organic layer
was transferred to a test tube and evaporated to dryness under
nitrogen. The dried sample residue was reconstituted in 1 mL of
HBS buffer containing 0.5% methanol and sonicated until it
dissolved completely. The samples were ready for the following
competitive assays.
Competitive assays were determined by mixing mAb (1 µg/
mL for 10 nm nanogold and 0.2 µg/mL for 40 nm nanogold
enhancement assays) with a series of standard CAP solutions in
HBS buffer or honey spiked-samples (1:1 v/v). After incubation
for 30 min, the mixture was then injected (60 µL, 20 µL/min)
into the sensor surface, followed by sequential injection of IgG/
nanogold particles (40 µL for 2 min). Final regeneration was
achieved using the same procedure as above. The results were
analyzed statistically using Sigma Plot version 8.0 (SPSS, Chicago,
IL). All assay standard curves were fitted to a four-parameter
logistic plot.
Determination of Antibody Cross-Reactivity. The cross-
reactivity profile of the antibody with thiamphenicol (TAP) and
chloramphenicol-base (CAP-base) was determined in buffer using
the same assay conditions as described in the competitive assay
of CAP. The cross-reactivity value for each drug was calculated
by comparing the IC50 for CAP with the IC50 for TAP and CAP-
base according to the following equation:9
RESULTS AND DISCUSSION
Immobilization of CAP-OEG-OVA Conjugate onto mSAM
Surface. Previous work on the design of protein conjugates has
shown that the insertion of a water-soluble, 23-atom long OEG
linker10 between the small molecule and the protein can signifi-
cantly increase the degree of hapten density in the protein
conjugate.6 In the present work, the CAP-OEG-OVA conjugate 4
was prepared by the same procedure6 and a similar hapten (CAP)
number of 11 was achieved (see Supporting Information). There
are several advantages in attaching protein conjugates onto the
mSAM surface rather than direct attachment of small molecules.
These include (1) the surface density of the small molecule can
be increased by attachment of small molecules through multiple
sites of amino acid residues on the three-dimensional surface
structure of the protein and (2) protection and stabilization of the
potentially vulnerable mSAM surface by covering the mSAM with
the proteins and making the hydrophobic mSAM surface hydro-
philic, which also prevents nonspecific binding. In the present
study, we have demonstrated an additional benefit whereby the
protein conjugates can increase the stability of the chemical bonds
between the antigen and the surface. In our work, the CAP is
covalently linked to the OEG via an ester linkage, which is easily
hydrolyzed under acidic or basic conditions (Scheme 1). However,
this vulnerable ester linkage is stable in the CAP-OEG-OVA
conjugate 4 on the mSAM surface, on which over 400 regenera-
tions were performed under acidic conditions (pH 2.0) without
damaging the antigen surface (Figure 1). Such chemical stability
of the protein conjugate is important since many small molecules
have no primary amine or carboxylic groups appropriate for a
stable amide linked immobilization. Therefore, protein conjugation
is an excellent alternative approach in immobilizing these small
molecules without the need for a stable amide linkage.
Au Nanoparticle Enhanced Bindings and Sensitivity
Improvement. SPR assay has been widely employed for rapid
biomolecular interaction studies including detection of small
molecules.11 The label-free format of SPR, however, has limited
applications for ultrasensitive bioanalysis because of limited
surface refractive index changes for detection particularly for small
molecules. For example, current SPR inhibition assays for small
molecules have only achieved an LOD of 0.1 ng/mL12-14 with
monoclonal antibodies, or an LOD of 20 pg/mL15 with a polyclonal
antibody, as binding agents. To address this limitation, we have
recently developed an SPR assay of small molecules by using 10
nm Au nanoparticles for signal enhancement on a mSAM surface
immobilized with analyte-protein conjugates.6 The assay achieved
the lowest reported LOD of 4.9 pg/mL for progesterone and also
created a stable mSAM sensor surface as demonstrated by the
performance of over 400 binding/regenerations. However, it is
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IC50 for CAP
IC50 for TAP or CAP-base
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B 2005, 107, 193–201
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Analytical Chemistry, Vol. 80, No. 21, November 1, 2008 8331

still a significant challenge to further enhance the sensitivity of
SPR analysis of small molecules from an LOD of 4.9 pg/mL to
the ultrasensitive level of femtograms per milliter possible with
LPCR1 or the attomole-range achievable with noncompetitive
immunoenzymometric assays.2
SPR is a quantum optical-electrical phenomenon whereby the
plasmon produces an electronic field approximately 300 nm out
from the sensor surface. Within this 300 nm range, any SPR
response or enhancement effect by metal nanoparticles is col-
lectively a function of substrate metal,16,17 particle size,18 and
distance19 between the metallic nanoparticles and metal surface.
The electromagnetic coupling between the nanoparticles and the
surface, light scattering of the nanoparticles, and energy field
modulation by the dielectric spacer layer can all influence the SPR
responses. Despite the potential of large Au nanoparticles in SPR
signal enhancement for the determination of extremely low
quantities of analyte, it has been generally considered that large
Au particles are not ideal for SPR analysis because of slow diffusion
kinetics and steric hindrance especially for the analysis of small
molecules. The large steric bulk of the nanoparticles can block
the approach of the antigen to the antibody thereby preventing
the approach of the gold nanoparticles sufficiently close to the
gold binding surface to allow plasmon resonance in the nanopar-
ticles to occur and the signal to be enhanced. The preparation of
a stable surface has also been difficult but is essential to allow
repeated surface regenerations to remove large bound nanopar-
ticles. So far, there have been no successes in the literature5 that
use large Au nanoparticles (g40 nm) for SPR assays of small
molecules.
Figure 2. Binding responses in HBS buffer for mAb only (b), IgG/
nanogold (10 nm) enhancement (9), and IgG/nanogold (40 nm)
enhancement (2).
In the present work, we have for the first time to successfully
use large Au nanoparticles (40 nm) as an enhancement tag for
achieving a SPR assay of small molecules (CAP) at a level of
femtogram per milliliter (fg/mL) (Figure 1). Transmission electron
microscopy (TEM) results showed that both 40 and 10 nm Au
nanoparticles are almost spherical and well separated from each
other (see Supporting Information), and therefore the 40 and 10
nm Au nanoparticles for SPR signal enhancement can be com-
pared on their size differences only since both Au nanoparticles-
enhanced assays have the same substrate metal and separating
distance between the nanoparticles and the surface.
After anti-CAP antibody (mAb) binding on the surface, the
enhancement performance of the Au nanoparticle/IgG (Figure
2) clearly showed that the large Au nanoparticles (40 nm)
dramatically increased the signal of antibody binding by 21.5-fold,
while the 10 nm Au nanoparticles only gave 2.3-fold signal
amplification. Both signal enhancements lead to a large reduction
in the mAb concentration required in a competitive assay of CAP
from 6 µg/mL (nonenhanced assay) to 1 (10 nm nanogold
enhanced assay) or 0.2 µg/mL (40 nm nanogold enhanced assays)
separately. The competition assays were evaluated with a nonen-
hanced and either a 10 or 40 nm nanogold-enhanced CAP assay
Figure 3. Comparison of four assay standard curves of CAP on
CAP-OEG-OVA/mSAM sensor surface with mAb only (b), IgG/
nanogold enhancement of 10 nm in HBS buffer (O), and 40 nm in
HBS buffer (1) or for honey spiked-samples (3).
(Figure 3). Different ranges of free CAP detection were deter-
mined by mixing mAb with a series of CAP standard solutions in
HBS buffer or honey spiked-samples (1:1 v/v). After incubation,
the mixture was injected over the sensor surface, followed by
sequential injection of 10 or 40 nm nanogold. The linear range of
detection was found to be 25-800 pg/mL (IC50 of 124.2 pg/mL
in HBS buffer) for the 10 nm nanogold enhancement and 1-1000
fg/mL (IC50 of 303.6 fg/mL in HBS buffer) or 80-5000 fg/mL
(IC50 of 664.6 fg/mL in honey spiked-specimen) for the 40 nm
nanogold enhancement (Figure 3). The sensitivity was increased
by 2.5 × 106-fold after 40 nm Au particle enhancement, achieving
an LOD as low as 0.74 fg/mL in HBS buffer or 17.5 fg/mL in the
honey spiked-specimen, while 10 nm gold particles only gave 10-
fold decrease with an LOD of 34.1 pg/mL (Table 1). The results
of 40 nm nanogold enhancements in HBS buffer and honey spiked
samples have clearly demonstrated some influence of matrix
effects on the CAP assay performance characteristics as shown
by different IC50 and LOD values in different types of assay matrix.
However, such a huge difference between the 40 and 10 nm Au
nanoparticles is larger than would be anticipated on the basis of
particle mass or volume alone. According to the Mie theory on
the radiating plasmons (RPs) model,20 incident energy is dis-
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Actuators, B 2001, 105, 8–12
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11168
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8332 Analytical Chemistry, Vol. 80, No. 21, November 1, 2008

Table 1. Summary of Cap Immunoassay Parameters of LOD, IC50, Sensitivity and Signal Enhancement with Change
in Signal Enhancement Labeling and Primary Antibody Concentration
mAb concentration
signal enhancement
ratio
assay
sensitivity
assay format
(µg/mL)
LOD
IC50
mAb only (HBS buffer)
mAb + IgG/gold (10 nm)
(HBS buffer)
6
1
353.4 ± 66.9 (pg/mL)
34.1 ± 4.80 (pg/mL)
13.6 ± 2.7 (ng/mL)
n/a
2.3
2.5ng/mL
25pg/mL
124.2 ± 24.8 (pg/mL)
mAb + IgG/gold (40 nm)
(HBS buffer)
0.2
0.2
0.74 ± 0.12 (fg/mL)
17.50 ± 4.2 (fg/mL)
303.6 ± 30.3 (fg/mL)
664.6 ± 6.7 (fg/mL)
21.5
1 fg/mL
mAb + IgG/gold (40 nm)
(honey spiking)
80 fg/mL
sipated by absorption and far-field radiation is created by scatter-
ing. Therefore, the huge SPR enhancement effect by 40 nm Au
nanoparticles mainly resulted from scattering, which is dominant
over absorption for large colloids; while for small colloids (10 nm),
absorption dominates over scattering thus producing far less SPR
enhancement.
Similar scattering enhancement by large Au nanoparticles (50
nm) has also been recently reported for an ultrasensitive colori-
metric detection of methicillin-resistant Staphylococcus aureus
(MRSA) total genomic DNA (66 ng/mL).21 Moreover, for ultra-
sensitive SPR assay, the size of Au nanoparticles (40 nm) for small
molecules seems sufficiently large since further increases in the
particle size from 45 to 59 nm did not result in any significant
change in the SPR response.18 Such an ultrasensitive SPR assay
of small molecules (0.74 fg/mL) is also a result of the proximity
of large Au nanoparticles to the Au surface on the chip. For
example, the ideal distance, or Au particle-Au surface separation,
is 32 ± 5 nm19 to achieve the maximum SPR angle shift when
SiO2 was used as a dielectric spacer layer.
CONCLUSIONS
It has been demonstrated, for the first time, that the use of
large nanoparticles (40 nm) as a signal enhancement on a mSAM
sensor surface creates both a rapid and ultrasensitive SPR assay
of small molecules, demonstrated by a rapid CAP assay in less
than 10 min and ultrasensitivity (LOD ) 0.74 fg/mL). With control
of the proximity of the large Au nanoparticles to the mSAM surface
through protein conjugation with an optimal OEG linker,6 the
assay design provided a favorable binding and enhancing environ-
ment for large Au nanoparticles. Moreover our methods resulted
in the formation of a stable sensor surface on which over 400
binding/regeneration cycles were performed without damaging
the SAM attachment and immobilized CAP. The assay format is
generic and can be applied to many other small molecules for
rapid and ultrasensitive detection with various SPR technologies.
ACKNOWLEDGMENT
Thanks to the New Zealand Foundation for Research Science
and Technology (FRST) for financial support (Grant C06X0411).
SUPPORTING INFORMATION AVAILABLE
Additional information as noted in text. This material is
available free of charge via the Internet at http://pubs.acs.org.
Finally, the SPR assay showed a small degree of cross reactivity
for TAP (3.1%) and no cross reactivity for CAP-base, suggesting
that the antibody conjugate is specific for the CAP on the surface.
Received for review June 25, 2008. Accepted September
5, 2008.
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Biotechnol. 2004, 22, 883–887
.
AC801301P
Analytical Chemistry, Vol. 80, No. 21, November 1, 2008 8333