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Advances in
Nanosensors for
Biological and
Environmental
Analysis
This page intentionally left blank
 Advances in
Nanosensors for
Biological and
Environmental
Analysis
EDITED BY

AKASH DEEP, MSC, MPHIL, PHD

Nanotechnology Lab
CSIR-Central Scientific Instrument Organisation (CSIR-CSIO)
Chandigarh, India
Academy of Scientific and Innovative Research (AcSIR-CSIO)
Chandigarh, India

SANDEEP KUMAR, PHD

Department of Bio and Nano Technology
Guru Jambheshwar University of Science and Technology
Hisar, India
Department of Civil Engineering
University of Nebraska Lincoln
Lincoln, United States

]
ADVANCES IN NANOSENSORS FOR BIOLOGICAL AND ISBN: 978-0-12-817456-2
ENVIRONMENTAL ANALYSIS
Copyright Ó 2019 Elsevier Inc. All rights reserved.

No part of this publication may be reproduced or transmitted in any form or by any means, electronic or
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This book and the individual contributions contained in it are protected under copyright by the Publisher (other
than as may be noted herein).

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List of Contributors

Valerio F. Annese Girish Chandra Mohanta

Electronics and Nanoscale Engineering CSIR-Central Scientific Instrument Organisation
School of Engineering (CSIR-CSIO)
University of Glasgow Chandigarh, India
Glasgow, United Kingdom Academy of Scientific and Innovative Research
(AcSIR-CSIO)
Gaurav Bhanjana Chandigarh, India
Department of Chemistry and Center of Advanced
Studies in Chemistry Ganga Ram Chaudhary
Punjab University Department of Chemistry and Center of Advanced
Chandigarh, India Studies in Chemistry
Panjab University
Neha Bhardwaj Chandigarh, India
Nanotechnology Lab
CSIR-Central Scientific Instrument Organisation Moondeep Chauhan
(CSIR-CSIO) Department of Environment Studies
Chandigarh, India Punjab University
Academy of Scientific and Innovative Research Chandigarh, India
(AcSIR-CSIO) Department of Chemistry and Center of Advanced
Chandigarh, India Studies in Chemistry
Panjab University
Sanjeev Kumar Bhardwaj Chandigarh, India
Nanotechnology Lab
Department of Bio and Nano Technology
CSIR-Central Scientific Instrument Organisation
Guru Jambheshwar University of Science and
(CSIR-CSIO)
Technology
Chandigarh, India
Hisar, India
Academy of Scientific and Innovative Research
(AcSIR-CSIO) David R.S. Cumming
Chandigarh, India Electronics and Nanoscale Engineering
School of Engineering
Deepanshu Bhatt University of Glasgow
Nanotechnology Lab Glasgow, United Kingdom
CSIR-Central Scientific Instrument Organisation
(CSIR-CSIO) Akash Deep
Chandigarh, India Nanotechnology Lab
Academy of Scientific and Innovative Research CSIR-Central Scientific Instrument Organisation
(AcSIR-CSIO) (CSIR-CSIO)
Chandigarh, India Chandigarh, India
Academy of Scientific and Innovative Research
(AcSIR-CSIO)
Chandigarh, India

v
vi LIST OF CONTRIBUTORS

Neeraj Dilbaghi Sandeep Kumar

Department of Bio and Nano Technology Department of Bio and Nano Technology
Guru Jambheshwar University of Science and Guru Jambheshwar University of Science and
Technology Technology
Hisar, India Hisar, India
Department of Civil Engineering
Ashraf Aly Hassan University of Nebraska Lincoln
Department of Civil & Environmental Engineering Lincoln, NE, United States
United Arab Emirates University
Al Ain, United Arab Emirates Poonma Malik
CSIR-Central Scientific Instrument Organisation
Sven Ingebrandt (CSIR-CSIO), Colony
IWE1eInstitut für Werkstoffe der Elektrotechnik1 Chandigarh, India
RWTH Aachen University
Germany Ruchi Mutreja
Chemical Biology Lab
Manpreet Kaur Biotechnology Department
Graduate Student IIT Roorkee
Indian Institute of Science Education and Research Roorkee, India
Mohali (IISER Mohali)
Mohali, India Suresh Neethirajan
BioNanoLab
Rajnish Kaur University of Guelph
Department of Physics Canada
Panjab University
Chandigarh, India Monika Nehra
Ubiquitous Analytical Techniques Department of Electronics and Communication
CSIR-Central Scientific Instruments Organisation Engineering
(CSIR-CSIO) Guru Jambheshwar University of Science and
Chandigarh, India Technology
Hisar, India
Madhu Khatri Department of Bio and Nano Technology
Faculty Guru Jambheshwar University of Science and
University Institute of Engineering and Technology Technology
(UIET) Hisar, India
Punjab University (PU)
Chandigarh, India Satish Kumar Pandey
CSIR-Central Scientific Instrument Organisation
Parveen Kumar (CSIR-CSIO)
CSIR-Central Scientific Instrument Organisation Chandigarh, India
(CSIR-CSIO)
Chandigarh, India Samadhan B. Patil
Electronics and Nanoscale Engineering
Rajeev Kumar School of Engineering
Department of Environment Studies University of Glasgow
Panjab University Glasgow, United Kingdom
Chandigarh, India
 LIST OF CONTRIBUTORS vii

Sandeep Kumar Sharma S.K. Tripathy

Department of Applied Science Department of Physics
Desh Bhagat University Panjab University
Mandi Gobindgarh, India Chandigarh, India

Sharvan Sehrawat Satish K. Tuteja

Faculty IWE1eInstitut für Werkstoffe der Elektrotechnik1
Indian Institute of Science Education and Research RWTH Aachen University
Mohali (IISER Mohali) Germany
Mohali, India
This page intentionally left blank
Preface
An ever-increasing demand of routine tests of various
biological and environmental parameters is driving the
development of portable and user-friendly diagnostic
tools. Many of such tools are nowadays available in
different forms including papers, electrodes, chips, kits,
etc. A growing significance of portable detection tools
has placed them as useful alternatives to conventional
analytical techniques like spectroscopy and biochemical
tests. In order to attain sufficient competitiveness, the
modern-day portable detection tools and diagnostic
devices are being integrated with nanotechnology. The
application of nanomaterials and nanosurfaces has
enabled the researchers to design highly sensitive nano-
sensors that can match the performance of laboratory-
based instruments. Owing to the large surface areas
and various other material properties, the nanosensors
offer many significant advantages, such as high sensi-
tivity, robust attachment of analyte specific receptors,
label-free sensing, signal reproducibility, etc. Recent
decades have witnessed the emergence of carbon nano-
tubes, graphene, metal nanoparticles, quantum dots,
and transition metal dichalcogenides as transducer sur-
faces of choice in the development of nanosensors for
both environmentally and clinically important parame-
ters. This book provides a comprehensive overview on
the most important types of nanosensor platforms
explored and developed in the recent years for efficient
detection of environmental/clinical analytes. It covers
the information in such a way that the readers could
grasp knowledge on important aspects of nanosensor
development, e.g., material synthesis, their functionali-
zation with different receptors, and techniques to record
signals. A commercial perspective of the nanosensor
market is also discussed.
Chapter 1 of the book elaborates the applications of
carbon-based nanomaterials in the development of
various nanosensors. Carbon nanomaterials have
become immensely popular among the researchers
worldwide for designing sensitive sensors on account of
their different useful properties. Carbon nanomaterials
can be synthesized in different forms such as nanosheets,
nanotubes, fullerene, and quantum dots. Consequently,
these nanomaterials have opened up numerous oppor-
tunities in order to seek the realization of nanosensors
for diverse environmental and clinical parameters. As
Chapter 2 discusses, 2-dimensional transition metal
dichalcogenide (2D TMD) nanomaterials have been
touted as promising alternatives to carbon nano-
materials in recent years. Molybdenum- and tungsten-
based disulfides are two important examples in this
category. Atomically thin 2D TMDs possessing direct
band gap property provide the development of plat-
forms for the sensing of pH, gases, heavy metals, pro-
teins, etc. Along with carbon nanomaterials, 2D TMDs
are suitable to be integrated as lab-on-chip type of
devices.
The utility of conducting polymers in the develop-
ment of sensing systems is widely recognized. Chapter 3
elaborates information on the synthesis and sensing
applications of various conducting polymers. In addi-
tion to this, the characteristics and applications of con-
ducting polymer composites with other nanomaterials
have also been outlined. Metaleorganic frameworks
(MOFs) are a class of coordination polymers which are
attracting significante attention of researchers world-
wide. Chapter 3 also focuses on this trending topic of
developing MOF-based sensors. Some extraordinary
material characteristics of MOFs (e.g., their high surface
area, tunable porosity, formation in different forms) are
the factors inspiring scientists and researchers to explore
them in the sensing of gases, proteins, explosives, anions,
cations, antigens, etc. Chapter 4 is dedicated to discuss
different biorecognition molecules and methods of their
isolation and production. Biomolecules are an impor-
tant component of many sensors wherein they help to
attain the desired selectivity during analysis. Nucleic
acids, enzymes, aptamers, and antibodies are the main
categories of biomolecules used in the development of
selective sensors. Nowadays, specific biomolecules are
available in the market to target the selective recognition
of analytes such as antigens, pesticides, heavy metals, etc.
Chapter 5 provides information about the techniques
to modify the nanosurfaces with biomolecules. The
contents of this chapter would help the readers to
ix
x PREFACE
understand different covalent and physical
attachmentebased chemistries in order to immobilize
biomolecules over surfaces of nanomaterials. A robust
immobilization of biomolecules is immensely critical to
define the stability, accuracy, and shelf life of nano-
biosensors. The utilization of nanomaterials in the
fabrication of easy-to-use paper and lateral flow sensors
is described in Chapter 6. These simple sensing devices
find important application in soil, air and, water quality
monitoring due to their low cost and convenient
handling. The advantages and existing limitations/chal-
lenges of nanosensors have been discussed in Chapter 7.
Finally, commercial aspects and an overview of currently
available sensors for various analytes are mentioned in
Chapter 8.
The nanosensor market is growing at an exponential
pace. Important features like fast response, portable
instrumentation, and cost-effectiveness are major factors
behind the escalating demand of these point-of-care
devices. They not only match the detection limits of
laboratory-based instruments but also serve the purpose
utilizing much smaller sample volumes. Nanosensors
are being accepted as preferred tools by clinical labora-
tories, regulatory agencies, and common users. Their
integration with or into smartphones and wearable de-
vices is one of trending technologies of the 21st century.
This book provides information on the most useful
nanosurfaces that are being employed for the develop-
ment of nanosensor. Critical aspects, such as the type of
nanomaterials, their conjugation with biomolecules,
methods of signal collections, applications, challenges,
advantages and limitations, and commercial aspects
have been covered. The book is written for readers from
different related backgrounds including nanotech-
nology, biochemistry, environmental science, chemistry,
material science, and biomedical engineering.
Contents
1 Carbon-Based Nanomaterials for the
Development of Sensitive Nanosensor
Platforms, 1
Monika Nehra, Neeraj Dilbaghi,
Ashraf Aly Hassan and Sandeep Kumar
2 Advances in the Synthesis and
Development of Two-Dimensional
Transition-Metal Dichalcogenide-Based
Nanosensor Platforms, 27
Sanjeev Kumar Bhardwaj, Neha Bhardwaj,
Deepanshu Bhatt, Poonma Malik and
Akash Deep
3 Conducting Polymers and
Metal-Organic Frameworks as
Advanced Materials for Development
of Nanosensors, 43
Moondeep Chauhan, Sanjeev Kumar Bhardwaj,
Gaurav Bhanjana, Rajeev Kumar,
Neeraj Dilbaghi, Sandeep Kumar and
Ganga Ram Chaudhary
4 Synthesis and Production of Different
Biomolecules for Application in the
Sensing of Environmental Pollutants, 63
Manpreet Kaur, Madhu Khatri and
Sharvan Sehrawat
5 Bioconjugation of Different
Nanosurfaces With Biorecognition
Molecules for the Development
of Selective Nanosensor Platforms, 79
Satish K. Tuteja, Ruchi Mutreja,
Suresh Neethirajan and
Sven Ingebrandt
6 Development of Disposable Sensor
Strips for Point-of-Care Testing of
Environmental Pollutants, 95
Satish Kumar Pandey, Girish Chandra Mohanta
and Parveen Kumar
7 Advantages and Limitations
of Environmental Nanosensors, 119
Rajnish Kaur, Sandeep Kumar Sharma and
S.K. Tripathy
8 Commercial Aspects of Biosensors
for Diagnostics and Environmental
Monitoring, 133
Samadhan B. Patil, Valerio F. Annese
and David R.S. Cumming
Index, 143
xi
This page intentionally left blank
CHAPTER 1
Carbon-Based Nanomaterials for the
Development of Sensitive Nanosensor
Platforms
MONIKA NEHRA • NEERAJ DILBAGHI • ASHRAF ALY HASSAN •
SANDEEP KUMAR
INTRODUCTION
Carbon is one of the most commonly found ele-
ments in nature and its understanding has reached
new levels, from macroscopic to nanoscale, with
continuous advancement in nanotechnology.
The nanostructures of carbon in their different
forms have been applied in diverse fields such as
field emission displays, nanoelectronics, energy
conversion and storage, biological and chemical
sensors, and theranostics. In the 21st century,
owing to their extraordinary properties in terms
of structural perfection, carbon nanomaterials
have in fact led general science to many advanced
avenues. The structural properties of carbon mate-
rials have inspired the synthesis of novel nanoma-
terials with similar symmetries and structures, e.g.,
fullerenes, nanotubes, nanodots, and graphene.
Carbon atoms have the ability to form robust
mutual covalent bonds in different hybridization
states such as sp, sp2, and sp3. Carbon atoms also
interact with nonmetallic elements leading to
the formation of a wide range of structures from
small molecules to long chains. The major classi-
fication of carbon materials (such as carbon, dia-
mond, and graphite) is based on the way of
interconnection between carbon atoms, e.g., tetra-
hedral sp3 atom configuration in case of diamond
and hexagonal sp2 carbon atom configuration in
case of graphene monolayers. However, mixed
states also exist and lay the basis for nanocrystal-
line diamond, diamond-like carbon, and
Advances in Nanosensors for Biological and Environmental Analysis. https://doi.org/10.1016/B978-0-12-817456-2.00001-2
Copyright © 2019 Elsevier Inc. All rights reserved.
amorphous carbon. The carbon nanoallotropes
belonging to same group or same arrangement
of carbon atoms have common properties; how-
ever, there are significant differences because of
their different sizes and shapes.
A summary of carbon nanomaterials with
different morphologies but unique chemical
properties is shown in Fig. 1.1 (Yan et al., 2016).
They can be classified according to their structural
dimensionality as, e.g., (1) 0D nanostructures
(fullerenes, carbon dots, nanodiamonds, etc.),
(2) 1D nanostructures (carbon nanotubes
[CNTs]/carbon nanofibers [CNFs], etc.), and (3)
2D nanostructures (graphene, graphene nanorib-
bons, etc.). Diamond, a metastable state of car-
bon, consists of a 3D cubic lattice with 3.57 Å
lattice constant along a CeC bond length of
1.54 Å (Sque et al., 2006). In contrast, graphite
possesses a 2D layered structure having a CeC
bond length of 1.42 Å (Baughman et al., 1987).
In graphene, the layers are single atom thick and
interact through van der Waals forces having
3.35 Å interlayer spacing. Graphite corresponds
to the most thermodynamically stable form of car-
bon at room temperature. Graphene is commonly
referred as a 2D building block of sp2 hybridized
carbonaceous nanomaterials; it can be rolled
and/or distorted in order to form CNTs and fuller-
enes. The first successfully synthesized carbon
nanomaterial was C60 (also known as buckmin-
sterfullerene), through laser ablation of graphite
1
2 Advances in Nanosensors for Biological and Environmental Analysis
FIG. 1.1 Broad family of carbon nanomaterials. (Adapted from Yan, Q.L., Gozin, M., Zhao, F.Q.,
Cohen, A., Pang, S.P., 2016. Nanoscale 8, 4799e4851. Reprinted with permission from RSC.)
under helium flow (Kroto et al., 1985). However,
some reports also exist on the development of
even-numbered carbonaceous clusters before the
development of C60, but these clusters were un-
suitable for characterization because of their large
size distributions (Rohlfing et al., 1984). Fuller-
enes are viewed as the 0D form of graphitic carbon
and also referred as irregular sheets of graphene
that are curled in the form of a sphere via penta-
gons incorporation in the structure. Furthermore,
CNTs were isolated as an offshoot during the syn-
thesis of fullerene. The elongation of fullerene in
one dimension assumes the structure of CNTs
with high aspect ratios (e.g., from 102 to 107).
Different synthesis methods have been devel-
oped for the production of both single-walled
CNTs (SWCNTs) and multiwalled CNTs
(MWCNTs). These include arc discharge (Arora
and Sharma, 2014), chemical vapor deposition
(CVD) (Kumar et al., 2017a), and high-pressure
carbon monoxide method (Liu et al., 2011a).
The initial development of graphene was done
by its growth on insulating substrates by Geim
and coworkers (Novoselov et al., 2004). In the
current perspective, graphene can be referred as
the mother of all graphitic carbon. Graphene has
further been investigated as monolayer to a few-
layered nanomaterial depending on the thickness
requirement in a particular application (Liu et al.,
2018).
Carbon nanomaterials can possibly cover the
characteristics of different substances on the earth,
such as hardest to softest materials, insulators to
semiconductors and further to superconductors,
and fully light-absorbing to completely trans-
parent materials. The superiority of carbon nano-
materials is basically due to their hardness,
radiation characteristics, optical properties, elec-
tric conductivity, chemical resistance, heat resis-
tance, electric insulation, and surface/interface
properties in comparison to many other materials.
CARBON-BASED NANOMATERIALS FOR
NANOSENSOR DEVELOPMENT
Nanosensors are becoming a crucial part of mod-
ern lifestyle, particularly in healthcare due to the
demand of point-of-care devices, personalized
medicine, and cheaper and reliable diagnostic
tools. Carbon nanomaterials have motivated re-
searchers to implement them as ideal transduc-
tion materials mainly because of their geometry,
fast electron transfer kinetics, wide potential win-
dow, low residual current, fluorescent properties,
and readily renewable surfaces (Jariwala et al.,
2013). Voluminous research efforts have been
dedicated to employ carbon nanomaterials in
the development of highly sensitive and selective
nanosensors.
Fabrication of Sensing Platforms
The device architecture of electrochemical nano-
sensors can be broadly classified into two main
categories: (1) the conventional three-electrode
 CHAPTER 1 Carbon-Based Nanomaterials for Nanosensor Development 3

FIG. 1.2 (A and B) A schematic representation of functionalized carbon nanomaterialebased

electrochemical biosensors. (Adapted from Yang, Y., Yang, X., Yang, Y., Yuan, Q., 2018a.
Carbon 129, 380e395. Reprinted with permission from Elsevier.)

setup and (2) the chemiresistive/field-effect tran-
sistor (FET) setup. The three-electrode setup is
commonly used for potentiometric, ampero-
metric, and impedimetric nanosensors that
comprise working, counter, and reference elec-
trodes for detection of different analytes
(Fig. 1.2) (Yang et al., 2018a). Carbon nanomate-
rials have been popularly used to modify the sur-
face of glassy carbon electrode to develop them as
a working electrode. As such, the electrochemical
sensing techniques are particularly helpful in un-
derstanding the behavior of analytes and their
associated electrochemical reaction mechanisms.
The analyte species that can undergo redox transi-
tions on glassy carbon electrodes can be detected
via different categories of electrochemical tech-
niques, e.g., amperometry and potentiometry.
The transfer of ions takes place through a conduc-
tive electrolyte medium. Carbon nanomateriale
based electrodes offer several benefits in terms of
wide potential window, good electrocatalytic ac-
tivities, and chemical inertness during redox reac-
tions. In electrochemical biosensors, carbon
nanomaterials also serve as a platform for biomol-
ecule immobilization, thereby improving electro-
chemical transduction (Hu et al., 2016).
Chemiresistive/FET sensors have attracted
considerable attention because of their enormous
benefits in terms of their fast response time, seam-
less integration with electronic manufacturing
processes, potential for miniaturization, and
parallel sensing (Bandodkar et al., 2016; Barbaro
et al., 2012). Carbon nanostructures, such as
CNTs, have been reported as excellent electrode
material for gas sensors in order to detect the gas
molecules, with high sensitivity, low device cost,
and fast response time even at room temperature
(Dube et al., 2015). In a similar manner, the
detection of gas molecules through graphene is
based on the change in their electric conductivity
as a result of the formation of surface adsorbates.
These surface adsorbates can work as either do-
nors or acceptors depending on their chemical na-
ture, preferential adsorption sites, and the
surrounding environment (Varghese et al., 2015).
The different formats of FET sensors may be
listed as ion-sensitive FET, unmodified comple-
mentary metal-oxide semiconductor, extended-
gate FET, floating-gate FET, and dual-gate FET
(Ramnani et al., 2016). Carbon nanomaterials
have been found very useful to function as the
functional channel in chemiresistor/FET nanosen-
sor configuration; for example, both the tubular
geometry of CNTs and the planar geometry of gra-
phene ensure maximum exposure of surface
atoms for the binding of target analyte molecules
to the electrode material. The Debye length (lD) is
a measure of field penetration into bulk materials
and also causes significant modulation in the elec-
tronic properties of electrode materials upon
exposure to the analytes. The lD is comparable
to the dimensions of the carbon nanostructures,
4 Advances in Nanosensors for Biological and Environmental Analysis
which ensures label-free sensing of analytes with
relatively low limits of detection and high sensi-
tivities. Carbon nanomaterials in an FET configu-
ration can detect multiple analytes concurrently
(Cullen et al., 1990). The fabrication of
graphene-FET devices requires the bulk produc-
tion of graphene films via different synthesis tech-
niques such as the CVD method, which offers
control over crystallinity, grain size, and number
of layers over the desired substrate (Srivastava
et al., 2010). CNTs can be synthesized as cylindri-
cal tubes by rolling up single and/or multiple gra-
phene sheets. The nature of CNTs, either metallic
or semiconducting, depends on their chirality
(armchair, zigzag, or chiral) and the diameter of
the tubes (Dresselhaus et al., 2004). For instance,
armchair SWCNTs are metallic in nature and,
therefore, these cannot be used for fabrication of
FET nanosensors.
Surface Functionalization of
Nanocarbon Electrodes
The surface chemistry of electrode materials has
significant importance with respect to their inter-
action with analytes present in complex solu-
tions/matrices. The surface functionalization of
carbon nanomaterials has been reported through
covalent/noncovalent interactions and/or decora-
tion with some inorganic nanomaterials (Liu
et al., 2015; Karimi et al., 2015; Balasubramanian
and Burghard, 2005). Carbon nanomaterials with
multiple functional groups at their surface/edges
can satisfy the specific requirements of different
kinds of sensors with specific intermolecular inter-
actions. For instance, graphene plays a significant
role in electrochemical sensing application
because of its high carrier mobility, exceptional
electrochemical properties (electron transfer
rates), optical properties, and structural character-
istics. Furthermore, its properties can be
controlled/modulated through adoption of suit-
able preparation methods and or functionaliza-
tion to generate specific target-sensing properties
(Kybert et al., 2014). Likewise, the presence of
functional groups such as carboxyl and amine
makes CNTs compatible in their conjugation
with biomolecules as well as other materials
such as metallic nanoparticles (Kong et al., 2001;
Gao et al., 2012). For covalent functionalization,
carboxylic (eCOOH) groups can be introduced
on the edge planes and sidewalls of the carbon
nanomaterials through oxidation. Some forms of
carbon such as reduced graphene oxide have
readily available eCOOH groups. As a matter of
concern, the covalent functionalization can deteri-
orate the sp2 structure of the honeycomb lattice of
carbon while also inducing some defects on their
surface (such as disruption in p-electronic
network), ultimately resulting in deflation of elec-
tronic properties of the materials. Noncovalent
functionalization can be a solution to this limita-
tion, which does not influence the intrinsic struc-
ture of carbon nanomaterials and also maintains
their electronic and mechanical properties
(Georgakilas et al., 2012).
The functionalization of carbon nanomaterials
helps in improving their dispersibility, biocom-
patibility, and sensing properties. In literature,
carbon nanomaterials have been used in two ma-
jor ways to modify the biosensing electrodes: (1)
modification of the bulk material, for example,
mixing the electrode material with carbon paste
and (2) modification of the electrode surface, for
example, preparation of films on premade elec-
trode through CVD or any other method. The
incorporation of carbon paste in electrode mate-
rials (e.g., other nanomaterials) enhances the
overall electrocatalytic properties and supports
improved signal transduction (Fig. 1.3) (Huang
et al., 2006). Furthermore, the application of car-
bon paste in immobilizing the enzymes can offer
protection of these protein molecules from the
external environment, while also rendering better
stability and durability in comparison to the sys-
tem having enzyme immobilized over the bare
electrode surface (Akyilmaz et al., 2017).
APPLICATIONS OF NANOCARBON
ELECTRODES FOR SENSING OF
DIFFERENT ANALYTES
In the recent years, the nanosensor platforms have
gained significant research interest for sensitive
detection of a wide variety of analytes including
both chemical (such as dopamine, ascorbic acid,
uric acid, and norepinephrine) and biological
 CHAPTER 1 Carbon-Based Nanomaterials for Nanosensor Development 5

FIG. 1.3 A schematic representation of the different strategies used to design functionalized
carbon nanotube electrodes. SWCNTs, single-walled carbon nanotubes. (Adapted from Huang,
X.J., Im, H.S., Yarimaga, O., Kim, J.H., Jang, D.Y., Lee, D.H., Kim, H.S., Choi, Y.K., 2006. Journal of
Electroanalytical Chemistry 594, 27e34. Reprinted with permission from Elsevier.)

(such as airborne bacteria, folic and pantothenic
acid, protein, and mycotoxins). The use of carbon
nanomaterials as the functional electrode surface
of nanosensor platform offers several advantages
in terms of good electrocatalytic activity, enhanced
interfacial adsorption properties, fast electron
transfer kinetics, and high biocompatibility in
comparison to traditional materials. There are
different strategies for incorporation of these
nanomaterials into electrochemical sensors, e.g.,
drop casting (Kaniyoor et al., 2009), direct growth
on a substrate (Wang et al., 2009), polymer-based
coatings (Barsan et al., 2015), use of binders such
as Nafion or dihexadecyl hydrogen phosphate
(Liao et al., 2015), and screen printing (Chen
et al., 2016). In comparison to drop casting or
dip coating, the direct growth of the carbon nano-
materials over electrode surface provides more ho-
mogeneous coating as well as supports the batch
fabrication of nanosensors (Gooding, 2005).
Furthermore, polymer-based coatings can aid in
the physical and chemical properties of the carbon
nanomaterials as well as their dispersion for depo-
sition. Additional incorporation of metallic nano-
particles in polymer matrix is sometimes desired
to maintain the requisite level of electrode conduc-
tivity (Chun et al., 2010).
Detection of Heavy Metal Ions
The quantification/detection of heavy metal ions
(including Hg2þ, Cd2þ, Cu2þ, Pb2þ, As3þ, etc.) is
a major concern among chemists, environmental-
ists, and biologists due to their toxic nature as well
as prolonged persistence in the biosphere. The
heavy metal ion contamination has mainly been
caused by the fast industrial development and
related activities such as electroplating, battery
manufacturing, mining, and smelting. The metal-
lurgical industries release various harmful toxins
into our environment, which include nonbiode-
gradable chemicals and heavy metals. The exis-
tence of these contaminants, especially heavy
metal ions in water bodies, has direct influence
over the health of living systems (Bhanjana
et al., 2015, 2017). The conventional methods
for the analysis of heavy metal contamination
are based on different techniques such as atomic
absorption spectroscopy (AAS) (Luo et al.,
2015), atomic emission spectroscopy (Zhang
et al., 2014), inductively coupled plasma mass
spectrometry (ICP-MS) (Li et al., 2015), and
X-ray fluorescence spectrometry (Sitko et al.,
2015). These techniques are very costly and not
appropriate for on-site analysis. Moreover, these
approaches can only quantify the total amount
of heavy metals and it is not possible to analyze
the bioavailable concentrations that are accessible
to the living bodies. Electrochemical detection us-
ing nanosensors is among the simple, accurate,
and sensitive methods for the detection of metal
contamination in food and environment. The
electrochemical sensing techniques offer several
6 Advances in Nanosensors for Biological and Environmental Analysis
benefits in terms of low cost, portability, high
sensitivity, short analytical time, and easy adapt-
ability for in situ detections (Bhanjana et al.,
2016). The working electrode of the three-
electrode electrochemical system can be modified
with nanomaterials for improving the sensitivity
of the nanosensors (Cheng et al., 2018). Such
nanosensors are then operated by recording
changes in their potential, current, electrochemi-
cal impedance, and electroluminescence upon
the recognition of an analyte (Simpson et al.,
2018).
Among nanomaterials, carbon nanomaterials
are most interesting materials as adsorbents/
preconcentrator agents or transducer materials
used in the development of nanosensors. The car-
bon nanomaterials can respond to both organic
and inorganic analytes. The functionalization of
carbon nanomaterials with biological recognition
elements (e.g., enzymes, antibodies, DNA, or mi-
croorganisms) enables highly specific and sensi-
tive sensing of metal ions (Wanekaya et al.,
2008). In particular, DNA-based nanosensors
have gained much research interest in the recent
years for the detection of heavy metal ions owing
to their stability in biological pH environments
(Primo et al., 2015). Wen et al. (2018) have re-
ported the application of DNA-modified gra-
phene oxide/Prussian blue nanoparticles for
arsenite detection. The interaction of graphene ox-
ide with 50 -thiolate-labeled (GT)21-ssDNA facili-
tated the generation of Prussian blue
nanoparticles on gold electrode surface. The
(GT)21-ssDNAecontaining arsenite recognition
sequence offered excellent specificity for arsenite
detection (detection limit down to 0.058 ppb)
in real water samples. DNAzyme-functionalized
carbon nanostructureebased biosensors have
also been developed for some other metal ions
including Pb2þ, Hg2þ, and Cu2þ (Zhou et al.,
2016).
The utility of carbon nanomaterials has also
been demonstrated in the development of sensi-
tive chemosensors. For example, Lu et al. (2018)
proposed a novel 3D honeycomb structure of
N-doped carbon nanosheet framework decorated
with bismuth nanoparticles (Bi-NCNF) for selec-
tive and sensitive electrochemical sensing of
Pb2þ and Cd2þ (Fig. 1.4). The proposed system
offered a detection limit of 0.04 and 0.02 mg L1
for Pb2þ and Cd2þ, respectively.
In many reported cases, the performance of car-
bon nanomaterialebased nanosensors for detect-
ing heavy metal ions has been successfully
validated with standard techniques such as AAS
and ICP-MS. A summary of the recently developed
glassy carbonebased nanosensors (electrode sur-
face modified with different carbon nanostruc-
tures) for heavy metal detection is provided in
Table 1.1, which is intended to acquaint the
readers about the main design and performance
parameters of these related technologies. The
coupling of nanosensitive platform with carbon
nanomaterials has resulted in high sensitivity,
fast response, multianalyte detection, and low
detection limits. In spite of numerous benefits,
there are a few challenges associated in this area:
(1) accurate detection of metal ions in biological
samples (involving blood, saliva, urine, etc.), (2)
issue of false-positive signal interference and
chemical fouling, (3) detection of heavy metal
ions in their complex form, and (4) continuous
monitoring of water resources for detection of
metal ions (Gumpu et al., 2015). As per the
commercialization perspective, the research ef-
forts should be directed toward reusability, mass
production, and system integration.
Detection of Food Additives and
Pesticide Residues
The innovation in food industry is increasing at an
immense speed in terms of development and
application of pesticides, food additives, and ma-
terials for food protection/processing/coating/
packaging. Pesticides are excessively used in agri-
cultural activities to enhance the production yield
by controlling pests, insects, weeds, etc. Food ad-
ditives are introduced to deliver functional attri-
butes in order to improve food life/safety.
Undesirably high concentrations of food additives
and accidental contamination of veterinary drug
residues and pesticides in foodstuff and water re-
sources have become issues of major health
concern. The pesticide residues are very toxic sub-
stances and can lead to several health issues (such
as cholinergic dysfunction) in both humans and
 CHAPTER 1 Carbon-Based Nanomaterials for Nanosensor Development 7

FIG. 1.4 A schematic layout for fabrication of 3D N-doped carbon nanosheet framework
decorated with bismuth nanoparticles (Bi-NCNF) followed by their application for electrochemical
detection of Pb2þ and Cd2þ. EG, ethylene glycol; GCE, glassy carbon electrode; NPs, nanoparticles;
PVP, polyvinylpyrrolidone. (Adapted from Lu, Z., Dai, W., Lin, X., Liu, B., Zhang, J., Ye, J., Ye, J.,
2018. Electrochimica Acta 266, 94e102. Reprinted with permission from Elsevier.)

animals. The analysis of various types of contam-
inations (i.e., toxic food additives, veterinary drug
residues, and pesticides) that can affect our food
samples is usually carried out using methods
such as high-performance liquid chromatography
(HPLC) (Wahed et al., 2016), HPLC-mass spec-
trometry (Hoffmann et al., 2017), gas
chromatography-mass spectrometry (Jiménez-
Salcedo and Tena, 2017), and capillary electro-
phoresis (Omar et al., 2017). The development
of nanosensors has been reported based on elec-
trochemical biosensing of contaminants in food-
stuff and water. The electrochemical biosensors
offer fast operation and high selectivity, sensi-
tivity, and reproducibility (Rotariu et al., 2016;
Herzog et al., 2008).
The incorporation of carbon nanostructures en-
hances the loading of bioreceptors on the elec-
trode surface, apart from providing high
stability. Carbon nanoparticles also serve as a
relay for transfer of electrons between biomole-
cules and the electrode. Elyasi et al. (2013) re-
ported a Pt/CNT nanocompositeemodified
ionic liquid carbon electrode for specific determi-
nation of Sudan I (a coloring agent) with an excel-
lent limit of detection (0.003 mM). The
immobilization of an enzyme (from Inga edulis)
onto a carbon paste electrode containing
MWCNTs and Nafion has been reported to pro-
vide a highly selective and sensitive detection of
tert-butylhydroquinone (TBHQ, an antioxidant),
with quantification and detection limits of 1.25
and 0.41 mg L1, respectively (de Oliveira et al.,
2014). This sensing method can offer determina-
tion of TBHQ in commercial salad dressing sam-
ples with an acceptable level of accuracy (e.g.,
the relative error limiting to 5.4%). Likewise, a
gold-modified carbon paste electrode has been
demonstrated for electrochemical sensing of syn-
thetic dyes (e.g., Sunset Yellow and tartrazine) in
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Lord Streatley but also of the different sergeants he had had during the war,
who, however unlike each other to look at, were identical to listen to,
thought he must be one of Lady Laura’s friends after all, and began to open
the door again; and Mr. Thorpe advancing, damning as he went and saying
things about flunkeys that were new to the footman, entered that marble hall
which had struck such a chill into Sally’s unaspiring soul.
The butler appeared. The butler was suave where the footman had been
haughty. He had heard some of the things Mr. Thorpe was saying as he
hurried from his private sitting-room into the echoing hall, and had no
doubt that he was a friend of the family’s.
Lady Laura had been in to lunch, but had gone out again; Mrs. Luke was
motoring with Lord Charles—who the devil was he, Mr. Thorpe wondered
—down to Crippenham, where she was going to stay the night. Her
ladyship had had a telegram from his lordship to that effect, and she herself
was going down the following morning.
‘Where’s Crippenham?’ asked Mr. Thorpe.
The butler was surprised. Up to that moment he had taken Mr. Thorpe
for a friend, if an infrequent one, of Lady Laura’s.
‘His Grace’s Cambridgeshire seat,’ he said, in his turn with hauteur. ‘His
Grace is at present in residence.’
‘Crikey!’ thought Mr. Thorpe. ‘Got right in with the Duke himself, has
she?’ And he felt fonder of Sally than ever.

§
At this point Mr. Thorpe, who had been behaving so well, began to
behave less well. The minute the pressure of anxiety was relaxed, the
minute, that is, that he no longer suffered, he became callous to the
sufferings of the Lukes; and instead of at once letting them know what he
had discovered he kept it to himself, he hugged his secret, and deferred
sending till some hours later a telegram to each of them saying, ‘Hot on her
tracks.’
Quite enough, thought Mr. Thorpe, as jolly again as a sand-boy, and
immediately unable to imagine the world other than populated by sand-boys
equally jolly,—quite enough that would be to go on with, quite enough to
make them both feel better. If he told them more, they’d get rushing off to
Crippenham and disturbing the Duke’s house-party. The whole thing should
now be allowed to simmer, said Mr. Thorpe to himself. Sally should be
given a fair field with her duke, and not have relations coming barging in
and interrupting.
But what a girl, thought Mr. Thorpe, slapping his knee—he was in his
car, on the way to his club—what a girl. She only had to meet dukes for
them to go down like ninepins at her feet. Apart from her beauty, what
spirit, what daring, what initiative, what resource! It had been worth all the
anxiety, this magnificent dénouement. Safe, and sounder than ever. A
glorious girl; and he too had at once seen how glorious she was, and at
once, like the Duke, fallen at her feet. That girl, thought Mr. Thorpe, who
began to believe she would rise triumphant even over a handicap like
Jocelyn, might do anything, might do any mortal thing,—no end at all, there
wasn’t, to what that girl couldn’t do. And, glowing, he telephoned to
Scotland Yard, and later on, after having had his tea and played a rubber of
bridge, sent his telegrams.
Then he went quietly home. Things should simmer. Things must now be
left to themselves a little. He went quietly home to Abergeldie, and didn’t
let Mrs. Luke know he was there. Her feelings, he considered, were
sufficiently relieved for the present by his telegram; things must now be
allowed to simmer. And he took a little walk in his shrubbery, and then had
a hot bath, and dressed, and dined, ordering up a pint of the 1911 Cordon
Rouge, and sat down afterwards with a great sigh of satisfaction by his
library fire.
He smoked, and he thought; and the only thing he regretted in the whole
business was the rude name he had called Lady Laura Moulsford to that
fool Pinner. But, long as he smoked and thought, it never occurred to him to
resent, or even to criticise, the conduct of the Moulsford family. Strange as
it may seem, considering that family’s black behaviour, Mr. Thorpe dwelt
on it in his mind with nothing but complacency.
 XV

§
At Crippenham next morning it was very fine. London and South Winch
were in a mist, but the sun shone brightly in Cambridgeshire, and the Duke
woke up with a curiously youthful feeling of eagerness to get up quickly
and go downstairs. He knew he couldn’t do anything quickly, but the odd
thing was that for years and years he hadn’t wanted to, and that now
suddenly he did want to; and just to want to was both pleasant and
remarkable.
He had been thinking in the night,—or, rather, Charles’s thoughts, placed
so insistently before him, had sunk in and become indistinguishable from
his own; and he had thought so much that he hadn’t gone to sleep till nearly
five. But then he slept soundly, and woke up to find his room flooded with
sunshine, and to feel this curiously agreeable eagerness to be up and doing.
The evening before, when Charles came in from the garden and packed
his bewitching guest off to bed, he had been very cross, and had listened
peevishly to all his son was explaining and pointing out; not because he
wasn’t interested, or because he resented the suggestions being made, but
simply because the moment that girl left the room it was as if the light had
gone out,—the light, and the fire. She needn’t have obeyed Charles. Why
should she obey Charles? She might have stayed with him a little longer,
warming him by the sight of her beauty and her youth. The instant she went
he felt old and cold; back again in the condition he was in before she
arrived, dropped back again into age and listlessness, and, however stoutly
he pretended it wasn’t so, into a deathly chill.
Now that, thought the Duke, himself surprised at the difference his
guest’s not being in the room made, was what had happened to David too
towards the end. They didn’t read it in the Lessons in church on Sundays,
but he nevertheless quite well remembered, from his private inquisitive
study of the Bible in his boyhood, how they covered David when he was
old with clothes but he got no heat, and only a young person called the
Shunammite was able, by her near presence, to warm him. The Duke didn’t
ask such nearness as had been the Shunammite’s to David, for he, perhaps
because he was less old, found all he needed of renewed life by merely
looking at Sally; but he did, remembering David while Charles talked, feel
aggrieved that so little as this, so little as merely wishing to look at her,
should be taken from him, and she sent to bed at ten o’clock.
So he was cross, and pretended not to understand, and anyhow not to be
interested. But he had understood very well, and in the watches of the night
had come to his decision. At his age it wouldn’t do to be too long coming to
decisions; if he wished to secure the beautiful young creature—Charles said
help, but does not helping, by means of the resultant obligations, also
secure?—he must be quick.
He rang for his servant half an hour before the usual time. He wanted to
get up, to go to her again, to look at her, to sit near her and have her
fragrant, lovely youth flowing round him. The mere thought of Sally made
him feel happier and more awake than he had felt for years. Better than the
fortnight’s cure of silence and diet at Crippenham was one look at Sally,
one minute spent with Sally. And she was so kind and intelligent, as well as
so beautiful—listening to every word he said with the most obvious
interest, and not once fidgeting or getting sleepy, as people nowadays
seemed to have got into the habit of doing. It was like sitting in the sun to
be with her; like sitting in the sun on a warm spring morning, and freshness
everywhere, and flowers, and hope.
Naturally, having found this draught of new life the Duke wasn’t going
to let it go. On the contrary, it was his firm intention, with all the strength
and obstinacy still in him, to stick to Sally. How fortunate that she was poor,
and he could be the one to help her. For she, owing all her happiness to him,
couldn’t but let him often be with her. Charles had said it would be both
new and desirable to do something in one’s life for nothing; but the Duke
doubted if it were ever possible, however much one wished to, to do
anything for nothing. In the case of Sally it was manifestly impossible.
Whatever he did, whatever he gave, he would be getting far more back; for
she by her friendship, and perhaps affection, and anyhow by her presence,
would be giving him life.
‘Come out into the garden, my dear,’ he said, when he had been safely
helped downstairs—the stairs were each time an adventure—putting his
shaking hand through her arm. ‘I want to see your hair in the sun, while I
talk to you.’
 And leading him carefully out, Sally thought, ‘Poor old gentleman,’ and
minded nothing at all that he said. Her hair, her eyes, all that Oh my ain’t
you beautiful business, of which she was otherwise both sick and afraid,
didn’t matter in him she called the Jewk. He was just a poor old gentleman,
an ancient and practically helpless baby, towards whom she felt like a
compassionate mother; and when he said, sitting in the sunny sheltered seat
she had lowered him on to and taking her hand and looking at her with his
watery old eyes, that he was going to give her Crippenham, and that the
only condition he made was that he might come and do a rest-cure there
rather often, she smiled and nodded as sweetly and kindly as she smiled and
nodded at everything else he said.
Like the croonings of a baby were the utterances of the Duke in Sally’s
ears; no more meaning in them, no more weight to be attached to them, than
that. Give her Crippenham? Poor old gentleman. Didn’t know what he was
talking about any more, poor old dear. She humoured him; she patted his
arm; and she wished to goodness Laura would be quick and come and take
her to her husband.
Sally now longed to get to Jocelyn as much as if she had passionately
loved him. He was her husband. He was the father of the little baby. Her
place was with him. She had had enough of this fleshpot business. She was
homesick for the things she knew,—plain things, simple things, duties she
understood. Kind, yes; kind as kind, the picks were, and they meant well;
but she had had enough. It wasn’t right it wasn’t, at least it wasn’t right for
her, to live so fat. What would her father have said if he had seen her in the
night in Laura’s bedroom, among all that lot of silver bottles and brushes
and laces and silks, and herself in a thin silk nightgown the colour of skin,
making her look stark naked? What would he have said if he had seen her
having her breakfast up there as though she were ill,—and such a breakfast,
too! Fleshpots, he’d have said; fleshpots. And he would have said, Sally,
strong if inaccurate in her Bible, was sure, that she had sold her husband for
a mess of fleshpots.
This was no life for her, this was no place for her, she thought, her head
bowed and the sun playing at games of miracles with her hair while the
Duke talked. She drew impatient patterns with the tip of her shoe on the
gravel. She hardly listened. Her ear was cocked for the first sounds of
Laura. She ached to have done with all this wasting of time, she ached to be
in her own home, getting on with her job of looking after her man and
preparing for her child. ‘Saturday today,’ she mused, such a lovely look
coming into her eyes that the Duke, watching her, was sure it was his
proposed gift making her divinely happy. ‘We’d be ’avin’ shepherd’s pie for
dinner—or p’raps a nice little bit of fish....’
And, coming out of that pleasant dream with a sigh, she thought,
‘Oughtn’t never to ’ave met none of these ’ere. All comes of runnin’ away
from dooty.’
Apologetically she turned her head and looked at the Duke, for she had
forgotten him for a moment, besides having been thinking on lines that
were hardly grateful. Poor old gentleman—still keeping on about giving her
Crippenham. Crippenham? She’d as soon have the cleaning of Buckingham
Palace while she was about it as of that great, frightening house—or, come
to that, of a prison.
But how like a bad dream it was, being kept there with the morning
slipping past, and she unable to reach him across the gulf of his deafness.
By eleven o’clock she was quite pale with unhappiness, she could hardly
bear it any longer. Would she have to give manners the go-by and take to
her heels once more? This time, though, there would be no kind father-in-
law to lend her a car; this time she would have to walk,—walk all the way,
and then when she got there find Jocelyn unaided. And the old gentleman
kept on and on about Crippenham being hers, and everything in it....
’E’s nothin’ but a nimage,’ she said to herself in despair. ‘Sits ’ere like a
old idol. Wot do ’e know about a married woman’s dooties?’
‘Where’s Charles?’ asked the Duke.
Sally shook her head. She hadn’t seen a sign of him that morning.
‘I want him to get my solicitor down—no time to lose,’ said the Duke.
‘You’re to have the place lock, stock and barrel, my dear, such as it is—
servants and all.’
Servants and all? Poor old gentleman. Why, she wouldn’t know which
end of a servant to start with. She with servants? And these ones here who,
however hard she tried up there in the bedroom, wouldn’t make friends.
They called her Madam. She Madam? Oh, my gracious, thought Sally,
shrinking in horror from such a dreadful picture.
‘It’s a hole of a place,’ went on the Duke, ‘and quite unworthy of you,
but we can have more bathrooms put in, and it’ll do till we find something
you like better. And Charles tells me you married rather suddenly, and
haven’t got anywhere to go to at present. He also says you have to live close
to Cambridge, because of your husband’s studies. And he also says, and I
entirely agree with him, my dear, that you oughtn’t to be in Cambridge
itself, but somewhere more secluded—somewhere where you won’t be seen
quite so much, somewhere hidden, in fact. Now I think, I really do think,
that Crippenham, in spite of all its disadvantages, does exactly fulfil these
requirements. And I want you to have it, my dear—to take it as my wedding
present to you, and to live in it very happily, and bless it and make it
beautiful by your presence.’
Thus the Duke.
‘ ’E don’t ’alf talk,’ thought Sally, quivering to be gone.

§
Charles, on being sent for by the Duke, was nowhere to be found. That
was because he was in South Winch. He had gone off at daybreak in his car,
and at the very moment his father woke up to the fact of his absence and
asked where he was, he was standing in the drawing-room at Almond Tree
Cottage, his eyes fixed eagerly on the door, waiting for Mrs. Luke.
He hadn’t been able to sleep for thinking of her. Somehow he had got it
into his head that she, more than her son, would suffer through Sally’s
disappearance, and be afraid. Because, thought Charles, she would feel that
it was from her the girl had run, and that any misfortune that might happen
to her would be, terribly, laid at her door. For two whole days and two
whole nights that unfortunate woman must have gone through torture. What
Charles couldn’t understand was why he hadn’t thought of this before.
Indeed his and Laura’s conduct had been utterly unpardonable. The least he
could now do, he thought, as he lay wide awake throughout the night, was
to get to South Winch without losing a minute, and put Mrs. Luke out of her
misery, and beg her forgiveness.
She was in the garden when he arrived. The little maid, staring at the
card he asked her to take to her mistress, said she would fetch her, and
ushered him into the drawing-room, where he waited with the books, the
bright cushions, the Tiepolo, and two withered tulips in a glass from which
nearly all the water had dried away; and while he waited he fought with a
feeling he considered most contemptible, in face of the facts, that he was
somehow on an errand of mercy, and arriving with healing in his wings,—
that he was somehow a benefactor.
Sternly he told himself he ought to feel nothing but shame; sternly he
tried to suppress his glow of misplaced self-satisfaction. There was nothing
good about him and Laura in this business. They had, the pair of them, been
criminally impulsive and selfish. He knew it; he acknowledged it. Yet here
he was, secretly glowing, his eyes watching the door, as much excited as if
he were going to bestow a most magnificently generous, unexpected
present.
Then it opened, and Mrs. Luke came in. He was sure it was Mrs. Luke,
for no one else could look so unhappy; and the glow utterly vanished, and
the feeling of shame and contrition became overwhelming.
‘She’s safe,’ said Charles quickly, eager to put a stop at once to the
expression in her eyes. ‘She’s at my father’s. She’s going to Cambridge
today to your son. She’s been with us the whole time——’
And he went to her, and took her hand and kissed it.
‘If it weren’t so ridiculous,’ he said, his face flushed with painful
contrition, still holding her hand and looking into her heavy, dark-ringed
eyes, ‘I’d very much like to go down on my knees to you, and beg your
pardon.’

§
And while Charles was in South Winch, Laura was in Cambridge,
dealing with Jocelyn. She, like Charles, had become conscious of the
sufferings of the Lukes, and, like him, was obsessed by them and lost in
astonishment that she hadn’t thought of them sooner; but for some obscure
reason, or instinct, her compunctions and her sympathies were for Jocelyn
rather than for his mother, and after a second sleepless night, during which
she was haunted by the image of the unfortunate young husband and greatly
tormented, she went down, much chastened, to Cambridge by the first
possible train, with only one desire now, to put him out of his misery and
beg his forgiveness.
So that Jocelyn, sitting doing nothing, his untouched breakfast still
littering the table, sitting bent forward in the basket-chair common to the
rooms of young men at Cambridge, his thin hands gripped so hard round his
knees that the knuckles showed white, his ears strained for the slightest
sound on the staircase, his eyes hollow from want of sleep, sitting as he had
sat all the previous afternoon after getting Mr. Thorpe’s telegram and most
of the night, sitting waiting, listening, and perhaps for the first time in his
life, for his mother had not included religious exercises in his early
education, doing something not unlike praying, did at last hear a woman’s
step crossing Austen’s Court, hesitating at what he felt sure was his corner,
then slowly coming up his staircase, and hesitating again at the first floor.
All the blood in his body seemed to rush to his head and throb there. His
heart thumped so loud that he could hardly hear the steps any more. He
struggled out of his low chair and stood listening, holding on to it to steady
himself. Would they come up higher? Yes—they were coming up. Yes—it
must be Sally. Sally—oh, oh, Sally!
He flew to the door, pulled it open, and saw—Laura.
‘It’s all right,’ she panted, for the stairs were steep and she was fat, ‘it is
—about Sally—don’t look so——’ she stopped to get her breath—‘so
dreadfully disappointed. She’s safe. If you’ll—oh, what stairs——’ she
pressed her hand to her heaving bosom—‘come with me, I’ll—take you—to
her——’
And having got to the top, she staggered past him into his room, and
dropped into the basket-chair, and for a minute or two did nothing but gasp.
But how difficult she found him. Jocelyn, whose reactions were always
violent, behaved very differently from the way his mother at that moment
was behaving, placed in the same situation of being asked forgiveness by a
Moulsford. Instead of forgiving, of being, as Laura had pictured, so much
delighted at the prospect of soon having Sally restored to him that he didn’t
mind anything, he appeared to mind very much, and quarrelled with her.
She, accustomed to have everything she did that was perhaps a little wrong
condoned and overlooked by all classes except her own, was astonished.
Here she was, doing a thing she had never done before, begging a young
man to forgive her, and he wouldn’t. On the contrary, he rated her. Rated
her! Her, Laura Moulsford. She knew that much is forgiven those above by
those below, and had frequently deplored the practice as one that has
sometimes held up progress, but now that the opposite was being done to
herself she didn’t like it at all.
 ‘Oh, what a nasty disposition you’ve got!’ she cried at last, when
Jocelyn had been telling her for ten impassioned minutes, leaning against
the chimney-piece and glowering down at her with eyes flashing with
indignation, what he thought of her. ‘I’m glad now, instead of sorry, for
what I did. At least Sally has had two days less of you.’
‘If you’re going to rag me as well——’ began Jocelyn, taking a quick
step forward as if to seize and shake this fat little incredibly officious
stranger,—so like him, his mother would have said, to waste time being
furious instead of at once making her take him to Sally.
But Laura, unacquainted with his ways, was astonished.
Then he pulled himself up. ‘It’s not you I’m cursing really at all,’ he
said. ‘It’s myself.’
‘Well, I don’t mind that,’ said Laura, smiling.
‘I’ve got the beastliest temper,’ said Jocelyn.
‘So I see,’ said Laura.
‘Do you think,’ he asked, for in spite of his anger he was all soft and
bruised underneath after his two days of fear, and when the fat stranger
smiled there was something very motherly about her, ‘I shall ever get over
it?’
‘Perhaps if you try—try hard.’
‘But—look here, I don’t care what you say—what business had you to
make away with my wife?’
‘Now you’re beginning all over again.’
‘Make away with my wife, smash up everything between me and my
mother——’
‘Oh, oh——’ interrupted Laura, stopping up her ears, and bowing her
head before the storm.

§
It was ten more minutes before she got him out of his rooms and into a
taxi.
‘We’ve lost twenty minutes,’ she said, looking at her watch. ‘You’ve lost
twenty kisses you might have had——’
 ‘For God’s sake don’t rag me!’ cried Jocelyn, gripping her by the arm
and bundling her into the taxi.
‘But what,’ asked Laura, who had tumbled in a heap on the seat, yet who
didn’t mind being thrown in because she knew she deserved worse than
that, ‘what else can one do with a creature like you?’
And she told him very seriously, as they heaved along towards
Crippenham, that the real mistake had been Sally’s marrying beneath her.
‘Beneath her?’ repeated Jocelyn, staring.
‘Isn’t it apparent?’ said Laura. ‘Angels should only marry other angels,
and not descend to entanglements with perfectly ordinary——’
‘No, I’m damned if I’m ordinary,’ thought Jocelyn. ‘And who the devil
is she, anyhow?’
‘Bad-tempered,’ continued Laura.
‘Yes, I’m beastly bad-tempered,’ he admitted.
‘Conceited——’
‘I swear I’m not conceited,’ he said.
‘Aren’t you?’ said Laura, turning her head and scrutinising him with
bright, mocking eyes.
And then, coming swift and silent as an arrow along the road towards
their taxi, she saw her father’s car.
‘Oh, stop!’ she cried, leaping to her feet and thrusting as much of herself
as would go through the window. ‘Here’s my father—yes, and Sally. Stop—
oh, stop!’ she cried, frantically waving her arms.

§
It had been decreed by Fate that Jocelyn should be reunited to Sally in
the middle of the road just beyond Waterbeach, at the point where the lane
to Lyddiatt’s Farm turns off; for such was the Duke’s desire to help his
lovely friend and such his infatuation, that he had actually broken his rule
of never emerging from Crippenham, once he got there, till the day
appointed for his departure, and was himself taking her to Ananias to hand
her over in person to her husband, afterwards lunching with the Master,—a
thing unheard of, this lunching, for the Duke disliked the Master’s politics
and the Master disliked the Duke’s, but what wouldn’t one do to further the
interests, by saying a good word for them, of the young couple?
This he had arranged that morning before coming downstairs, his
amazed servant telephoning the message and receiving the Master’s
hypocritical expressions of pleasure in return, for apart from the Duke’s
politics the Master was no fonder of a deaf guest than anybody else; and
just as Sally, on that garden seat, was coming to the end of her patience and
submissiveness and was seriously thinking of jumping up and taking to her
heels, the parlourmaid appeared on the path; and when she was quite close
she stood still, and opened her mouth very wide, and roared out that the car
was at the door; and the Duke, with a final pat of benediction, bade Sally
fetch her hat, and come with him to her husband.
So there it was that they met,—the taxi and the Rolls Royce, Laura and
Jocelyn, Sally and the Duke. And on the Swaffham Prior side of
Waterbeach, where the crooked signpost points to Lyddiatt’s Farm, the dull,
empty road was made radiant for a moment that day by happiness.
‘Stop! Stop!’ cried Laura, frantically waving.
‘Sally! Oh—oh, Sally!’ shouted Jocelyn, standing up too, and trying too,
behind Laura, to wave.
The chauffeur recognised Laura, and pulled up as soon as he could; the
taxi pulled up with a great grinding of its brakes; Jocelyn jumped out of one
door, and Laura of the other; and both ran.
‘Why,’ said Sally, who didn’t know what had happened, turning her head
and looking in astonishment at the two running figures coming along
behind, ‘why,’ she said, forgetting the Duke was deaf, ‘ ’ere is Mr. Luke
——’
And in another instant Jocelyn was there, up on the step of the car,
leaning over the side, dragging her to him with both arms, hugging her to
his heart, and kissing her as if there were no one in the world except
themselves.
‘Sally—oh, my darling! Oh, Sally—oh, oh, Sally!’ cried Jocelyn, raining
kisses on her between each word. ‘How could you—why did you—oh, yes
—I know, I know—I’ve been a beast to you—but I’m not going to be any
more—I swear, I swear——’
‘Now don’t, Mr Luke,’ Sally managed to say, stifled though she was,
‘don’t get swearin’ about it——’
 And pulling her head away from him she was able to attend to the
proprieties, and introduce him.
‘My ’usband,’ introduced Sally, looking over his arm, which was round
her neck, at the old man beside her. ‘The Jewk,’ she said, turning her face
back to Jocelyn, who took no notice of the introduction, who didn’t indeed
hear, because the moment she turned her face—oh, her divine, divine little
face!—back to him, he fell to kissing it again.
And Laura, coming panting up just then, got up on the step on the other
side of the car, and shouted in her father’s ear, who could always hear
everything she said, ‘This is Jocelyn Luke, Father—Sally’s husband.’
And the Duke said, ‘I thought it must be.’
 XVI

§
Now the end of this story, which is only the very beginning of Sally, the
merest introduction to her, for it isn’t to be supposed that nothing more
happened in her life,—the end of it is that she did as she was told about
Crippenham, and if the Duke had been less than ninety-three there would
have been a scandal.
But after ninety there is little scandal. The worst that was said of the
Lukes was that they had got hold of the old man, and nobody who saw
Sally believed that. Indeed, the instant anyone set eyes on her the Duke’s
behaviour was accounted for, and after five minutes in her company it
became crystal clear that she was incapable of getting hold of anybody. So
young, so shy, so acquiescent,—absurd to suppose she ever had such a thing
as an ulterior motive. And the husband, too; impossible to imagine that
silent scholar, also so young, and rather shy too, or else very sulky,—
impossible to imagine him plotting. On the contrary, he didn’t seem to like
what had happened to him much, and showed no signs whatever either of
pleasure or gratitude. But of Jocelyn no one thought long. He was without
interest for the great world. He was merely an obscure young man at
Cambridge, somebody the Duke’s amazing beauty had married.
Sally did, then, as she was told about Crippenham. It was given her, and
she took it; or rather, for her attitude was one of complete passivity, it
became hers. But she had an unsuspected simple tenacity of purpose, which
was later to develop disconcertingly, and she refused to live anywhere
except in the four-roomed cottage in the corner of the garden, built years
before as a playhouse for Laura and Charles.
On this one point she was like a rock; a polite rock, against which
persuasions, though received sweetly and amiably, should beat in vain. So
the Duke had the little house fitted up with every known labour-saving
appliance, none of which Sally would use because of having been brought
up to believe only in elbow-grease, and two bathrooms, one for her and one
for Jocelyn; and he attached such importance to these bathrooms, and he
insisted so obstinately on their being built, that Sally could only conclude
the picks must need a terrible lot of washing. Whited sepulchres they must
be, she secretly thought; looking as clean as clean outside, fit to eat one’s
dinner off if it came to that, but evidently nothing but show and take-in.
The Duke, much concerned at first, settled down to this determination of
Sally’s, and explained it to himself by remembering Marie-Antoinette. She
had her Trianon. She too had played, as Sally wished to play, at being
simple. He consoled himself by speaking of the cottage as Little Trianon; a
name Sally accepted with patience, though she told Jocelyn—who was so
much stunned at the strange turn his life had taken that she found she could
be quite chatty with him, and he never corrected, and never even said
anything back—she wouldn’t have thought of herself. Some day, the Duke
was sure, the marvellous child would grow up and get tired of her Trianon,
and then, when she wanted to move into the house, she should find
Versailles all ready for her, and very different from what it used to be.
So, on the excuse of seeing to the alterations, he was hardly ever away
from Crippenham, and if he had been less than ninety-three there would
certainly have been a scandal.
But Jocelyn, who woke up after the wild joy and relief of being reunited
to Sally to find himself the permanent guest of a duke, didn’t know whether
to be pleased or annoyed. The problems of his and Sally’s existence were
solved, it was true, but he wasn’t sure that he didn’t prefer the problems. He
rubbed his eyes. This was fantastic. It had no relation to real life, which was
the life of hard work and constant progress in his cloister at Ananias. Also,
its topsy-turviness bewildered him. Here was the Duke, convinced that
Sally had married beneath her, and so unshakably convinced that Jocelyn
had enormous difficulty in not beginning to believe it too. He couldn’t help
being impressed by the Duke. He had never met a duke before, never come
within miles of meeting one, and was impressed. That first afternoon, when
he had been carried off in the Rolls Royce to Crippenham, he had spent the
time between luncheon and tea shut up in the old man’s study being
upbraided for having taken advantage, as he was severely told, of Sally’s
youth and inexperience and motherlessness to persuade her into a marriage
which was obviously socially disastrous for her; and he couldn’t even if he
had wished to, which he certainly didn’t, tell him about Mr. Pinner, because
he couldn’t get through the barrier of his deafness. There the old man had
sat, with beetling brows and great stern voice, booming away at him hour
after hour, and there Jocelyn had sat, young, helpless, silent, his forehead
beaded with perspiration, listening to a description, among other things, of
the glories which would have been Sally’s if he hadn’t inveigled her into
marrying him. And so sure was the Duke of his facts, and so indignant, that
gradually Jocelyn began to think there was something in it, and every
moment felt more of a blackguard. In the old man’s eyes, he asked himself,
would there be much difference between him and Pinner? And was there, in
anybody’s eyes, much difference? More education; that was all. But of
family, in the Duke’s sense, he had as little as Pinner, and if Pinner had been
to a decent school, as Jocelyn had, and then gone to Cambridge—no,
Oxford for Pinner—he would probably have cut quite as good a figure, if
not in science then in something else; perhaps as a distinguished cleric.
He sat dumb and perspiring, feeling increasingly guilty; and if he could
have answered back he wouldn’t have, because the Duke made him feel
meek.
This meekness, however, didn’t last. It presently, after a period of
bewilderment, gave way to something very like resentment, which in its
turn developed into a growing conviction that he had become just a cat’s
paw,—he who, if left to himself, could have done almost anything.
Naturally he didn’t like this. But how, for the moment, could he help it?
Sally was going to have a baby. They had to live somewhere. It was really
heaven-sent, the whole thing. Yet—Sally, whom he had been going to
mould, was moulding him. Unconsciously; nothing to do with any intention
or desire of her own. And what she was moulding him into, thought
Jocelyn, as he drove himself backwards and forwards every day between
Crippenham and Cambridge, between his domestic life and his work,
between the strange mixture of emotions at the one end and the clear peace
and self-respect at the other, turning over in his mind with knitted brows, as
he drove, all that had happened to him in the brief weeks since he had
added Sally to his life—what she was moulding him into was a cat’s paw.
Yes. Just that.
Were all husbands cat’s paws?
Probably, thought Jocelyn.

§
 Mrs. Luke also reacted to the Moulsfords in terms of meekness. Hers,
however, lasted. She found them permanently dazzling. Besides, there was
nothing to be done. Jocelyn had gone; she had lost him for ever; he would
never come back, she very well knew, to the old life of dependence on her.
And if he must go, if she must lose him, there really was no one in the
world she would more willingly lose him to than the Duke of Goring. For
certainly it was a splendid, an exalted losing.
When she had had time to think after that visit from Lord Charles—he
had, she considered, a curious attractiveness—and was more herself again,
when she had recovered a little from the extreme misery she had gone
through and began not to feel quite so ill, she found it easy to forgive her
mauvais quart d’heure. The Moulsfords were heaping benefits on her boy.
They were settling all his difficulties. That morning when she was so
unhappy, Lord Charles had been most delightfully kind and sympathetic,
and had told her that the Duke, his father, intended to help the young
couple,—‘You know my son won this year’s Rutherford Prize,’ she had
said. ‘Indeed I do,’ he had answered in his charming, eager way, adding
how much interested his father was in the careers of brilliant young men,
especially at Cambridge, helping them in any way he could—and who
would not, in such circumstances, forgive?
Mrs. Luke forgave.
The fact, however, remained that she was now alone, and she couldn’t
think what her life was going to be without Jocelyn. For how, she
wondered, did one live without an object, with no raison d’être of any sort?
How did one live after one has left off being needed?
That year the spring was late and cold. The days dragged along, each one
emptier than the last. There was nothing in them at all; no reason, hardly,
why one should so much as get up every morning and dress for days like
that,—pithless, coreless, dead days. She tried to comfort herself by
remembering that at least she wasn’t any longer beaten down and
humiliated, that she could lift her head and look South Winch in the face,
and look it in the face more proudly than ever before; but even that seemed
to have lost its savour. Still, she mustn’t grumble. This happened to all
mothers sooner or later, this casting loose, this final separation, and to none,
she was sure, had it ever happened more magnificently. She mustn’t
grumble. She must be very thankful. She was very thankful. Like Toussaint
l’Ouverture—Wordsworth, again—she had, she said to herself, sitting
solitary through the chilly spring evenings by her fire after yet another
empty day, great allies; only fortunately of a different kind from poor
Toussaint’s, for however highly one might regard, theoretically, exultations
and agonies and love and man’s unconquerable mind, she, for her part,
preferred the Moulsfords.
But did she?
A bleak little doubt crept into her mind. As the weeks passed, the doubt
grew bleaker. Invisible Moulsfords; Moulsfords delightful and most
friendly when one met them, but whom one never did meet; Moulsfords full
of almost intimacies; Moulsfords who said they were coming to see one
again, and didn’t come; Moulsfords benignant, but somewhere else: were
these in the long run, except as subjects of carefully modest conversation in
South Winch—and South Winch, curiously, while it was plainly awe-struck
by what had happened to Jocelyn yet was also definitely less friendly than it
used to be—were these in the long run as life-giving, as satisfying, as
fundamentally filling as Toussaint’s exultations and agonies?
Ah, one had to feel; feel positively, feel acutely. Anything, anything, any
anger, any pain, any anxiety, any exasperation, anything at all that stabbed
one alive, was better than this awful numbness, this empty, deadly, settled,
stagnant, back-water calm....
And one evening, when it had been raining all day, after a period of
standing at the drawing-room window looking out at the dripping front
garden, where the almond-tree by the gate shivered in the grey twilight like
a frail, half-naked ghost, she turned and went to her writing-table, and sat
down and wrote a little note to Mr. Thorpe, and asked if he would not come
in after his dinner, and chat, and show that they could still be good friends
and neighbours; and when she had finished it, and signed herself Margery,
with no Luke, she rang for the little maid, and bade her take it round to
Abergeldie and bring back an answer.
‘For after all,’ she said to herself while she waited, standing by the fire
and slowly smoothing one cold hand with the other, ‘he has sterling
qualities.’
 THE END

Printed in Great Britain by R. & R. Clark, Limited, Edinburgh.

Typographical error corrected by the etext transcriber:
It it were a=> If it were a {pg 126}