10 (2022) 100131
A ailable online 23 Ma ch 2022
2590-1370/© 2022 The Au ho s. Published by Else ie B.V. This is an open access a icle unde he CC BY-NC-ND license (h p://c ea i ecommons.o g/licenses/by-
nc-nd/4.0/).
Recen ad ances in i us imp in ed polyme s
Ma iana S. Amo im, M. Go e i F. Sales
**
, Manuela F. F asco
*
BioMa k@UC/CEB - LABBELS, Depa men o Chemical Enginee ing, Facul y o Sciences and Technology, Uni e si y o Coimb a, Coimb a, Po ugal
ARTICLE INFO
Keywo ds:
Molecula imp in ing echnology
Vi uses
Biosenso s
Disease de ec ion
ABSTRACT
Molecula imp in ing is a ma u e and appealing echnology o ob ain highly selec i e ecogni ion si es c ea ed
wi hin a polyme ic ne wo k, ha ing o he impo an ea u es such as obus ness and cos e ec i eness. The
applica ions o molecula ly imp in ed polyme s (MIPs) a e as , bu he e is a con inuous pa icula in e es in
hei in eg a ion in o sensing de ices. MIPs possess ema kable p ope ies in e ms o selec i i y and low-cos o
ailo ed biomime ic ecogni ion elemen s. The need o imp o e disease diagnosis and p e en ion by de ec ing
pa hogenic i uses and o he biological a ge s a he nanoscale has led o g ea ad ances in MIPs. Despi e s ill
acing many challenges, imp in ing app oaches can p o ide apid and accu a e i us ecogni ion and hus be
applied as new sensing ma e ials. Following a gene al o e iew o MIP echnology, key examples o i us
imp in ed polyme s, he applica ion o MIP-based ma e ials o o he nanoscale a ge s, and hei de ec ion a e
p esen ed he e. Pe spec i es and challenges a e also highligh ed o eseeing new u u e s a egies and MIP
designs.
1. In oduc ion
The abili y o c ea e biomime ic ma e ials co e s se e al ields o
esea ch. One o he mos p oli ic examples is he a i icial ligand-
binding si es c ea ed wi hin a polyme ic ma ix, known as molecula ly
imp in ed polyme s (MIPs). The inspi a ion comes om he na u al
molecula in e ac ions esul ing in e y s able binding phenomena ha
occu in he biological sys ems. Thus, MIPs a e desc ibed as plas ic an-
ibodies, and a e obus and highly selec i e ma e ials. The his o y o
MIPs is linked o sepa a ion p ocesses, i.e., he necessi y o en ichmen
o ex ac ion o small molecules (G ebe and Fla , 2019). None heless,
he use o his echnology has apidly e ol ed and is used o he
de ec ion o a ange o molecules, om small o la ge a ge s (Re aa
e al., 2019). No su p isingly, MIPs ha e now been success ully applied
o la ge and complex i uses, bac e ia, and cells (Liu e al., 2020; Ren
and Za e, 2012). Rega ding i uses, he challenges posed by ecu en
ou b eaks o pa hogenic i uses, and hei impac on heal h and he
economy, make hem appealing a ge s o con inuous imp o emen o
de ec ion me hods.
In his e iew, a gene al o e iew o he MIP echnology is i s
ou lined o hen highligh he mos signi ican examples o MIP-based
ma e ials de eloped o ecognize i uses and biological a ge s a he
nanoscale. MIPs de eloped using whole o in ac i uses can be mo e
challenging, bu a he same ime o e g ea e insigh on he syne gy
be ween he es ablished mul iple in e ac ions and he shape-
complemen a y ca i ies. Thus, i us imp in ed polyme s will be he
ocus o his e iew, hoping he examples o biomime ic ma e ials p e-
sen ed will inspi e he de elopmen o new s a egies and MIP designs.
2. Molecula imp in ing echnology
MIPs a e ailo -made syn he ic ma e ials wi h a i icially c ea ed
ecogni ion si es ha a e able o selec i e ebind a a ge compound,
ins ead o closely ela ed compounds (Tu iel and Es eban, 2019). MIPs
a e gene a ed by polyme izing unc ional and c osslinking monome s
a ound a empla e molecule, hus ob aining a c osslinked
h ee-dimensional ne wo k polyme (Tu iel and Es eban, 2019).
Compa ed o o he ecogni ion sys ems, MIPs possess h ee majo
unique ea u es: s uc u e eliabili y, ecogni ion speci ici y and uni-
e sal applica ion. Thus, hei use is e y signi ican in many ields,
anging om pu i ica ion and sepa a ion, ca alysis, chemo/biosensing,
and d ug deli e y. The ea u es o MIPs ega ding high chemical and
physical s abili y, s aigh o wa d p epa a ion, ema kable obus ness
and low-cos a e e y a ac i e in hese esea ch ields (Ca doso e al.,
2018; Chen e al., 2016). Among he many applica ions, he in eg a ion
o MIPs in o biosenso s is one o he mos c ea i e due o he need o
* Co esponding au ho .
** Co esponding au ho .
E-mail add esses: [email p o ec ed], [email p o ec ed] (M.G.F. Sales), [email p o ec ed] (M.F. F asco).
Con en s lis s a ailable a ScienceDi ec
Biosenso s and Bioelec onics: X
jou nal homepage: www.jou nals.else ie .com/biosenso s-and-bioelec onics-x
h ps://doi.o g/10.1016/j.biosx.2022.100131
Recei ed 31 Decembe 2021; Recei ed in e ised o m 15 Ma ch 2022; Accep ed 18 Ma ch 2022
Biosenso s and Bioelec onics: X 10 (2022) 100131
2
imp o e he selec i i y and sensi i i y o de ec ion o diagnos ic
me hods h ough cos -e ec i e and obus biomime ic s a egies
(Fig. 1).
2.1. MIP componen s
The e ec i eness, a ini y, and selec i i y o he ecogni ion si es o a
MIP a e g ea ly in luenced by i s componen s ( empla e, monome s,
c osslinke , polyme iza ion ini ia o and sol en ) and by he quali y o
he in e ac ions be ween hem. Thus, when designing he MIP, he se-
lec ion o i s componen s is a c i ical s ep o achie e he desi ed p op-
e ies o a ce ain applica ion (Su yana e al., 2021; Tu iel and Es eban,
2019).
Fi s , i is c ucial o conside he molecula s uc u e o he empla e
ha will de e mine he choice o he unc ional monome (s). The
s uc u e and he numbe o binding si es in he imp in ed polyme , i.e.,
he molecula ecogni ion, depends on he chemical bonds es ablished
be ween hem. Fo his eason, he empla e and unc ional monome (s)
mus complemen each o he o maximize he imp in ing p ocess (Co -
mack and Elo za, 2004; Su yana e al., 2021). Mo eo e , he empla e
should no possess unc ionali ies esponsible o po en ially inhibi ing
o e a ding he polyme iza ion and should be s able unde he syn hesis
condi ions (Co mack and Elo za, 2004). The c osslinke has also o be
conside ed as a undamen al elemen o a MIP. Se e al c ucial s eps
depend on a p ope c osslinking o he polyme , such as: (a) o ma ion o
he imp in ed pocke by eezing he empla e-monome complex upon
polyme iza ion; (b) s abilizing he imp in ing binding si e; (c) ensu ing
polyme mechanical s abili y; (d) con olling he mo phology o he
polyme ma ix. Thus, he eac i i y o he c osslinke and he a io
monome o c osslinke a e e y impo an o ob ain unc ionalized si es
ha a e sp ead uni o mly h oughou he polyme ic ne wo k and o
ha e p ope ly spaced ca i ies wi h no cons ains o he di usion o he
empla e (Be gmann and Peppas, 2008).
The polyme iza ion begins wi h ini ia o s as sou ce o chemical
species ha eac wi h he monome s o o m an in e media e compound
capable o linking successi ely wi h o he monome s in a chain-g ow h
polyme iza ion un il a polyme ic compound is eached. They a e o en
he mo-, pho o- o edox-ini ia o s, which ha e been ex ensi ely used in
commonly ee adical polyme iza ions. The sol en is he medium
whe e he polyme iza ion occu s and i s molecules occupy space in he
polyme ic ma ix, hus c ea ing po es and being named as po ogen. In
his sense, he na u e and olume o he sol en is also impo an o
de elop a o able po es in he ma ix ha will con ibu e o a p ope
di usion o he empla e ou o he ne wo k and i s subsequen di usion
back in o he polyme du ing ecogni ion (Be gmann and Peppas, 2008;
Vasapollo e al., 2011).
2.2. MIP syn hesis and p epa a i e app oaches
The s uc u e o he polyme ma ix is mac opo ous wi h nano-
ca i ies complemen a y o ha o he empla e molecule (Włoch and
Da a, 2019). To achie e his o ganized ne wo k, he molecula
imp in ing p ocess has se e al s eps (Fig. 1): (1) dissolu ion o empla e,
unc ional monome s, c osslinke , and ini ia o in he sol en ; (2)
in e ac ion o he unc ional monome s wi h he empla e molecule,
which esul s in he o ma ion o a s able empla e-monome complex;
(3) ixing he unc ional monome s posi ioned a ound he empla e by
copolyme iza ion wi h c osslinke s; (4) emo al o he empla e
Fig. 1. Schema ic ep esen a ion o biosenso s, alongside he scheme o p oducing MIPs.
M.S. Amo im e al.
Biosenso s and Bioelec onics: X 10 (2022) 100131
3
molecules om he ma ix, o example by ex ac ion wi h a sol en ; (5)
he polyme ma ix is le wi h binding si es complemen a y in shape,
size and unc ionali ies o he a ge compound (Tu iel and Es eban,
2019). The e o e, he ob ained polyme ma ix ecognizes and binds
selec i ely he empla e molecules (Włoch and Da a, 2019).
The esul ing imp in ed polyme s a e s able, obus , and esis an o
a wide ange o pH, sol en s, and empe a u e. The e o e, he beha iou
o MIPs mimics he in e ac ions es ablished by na u al ecep o s o
selec i ely e ain a a ge molecule bu wi hou he associa ed s abili y
limi a ions. Besides, he syn hesis o MIPs is ela i ely simple and
inexpensi e, p o iding a clea al e na i e o he use o na u al ecep o s
(Tu iel and Es eban, 2019).
Based on he ypes o in e ac ions be ween he a ge molecule and
he unc ional monome s, h ee gene al app oaches ha e been desc ibed
o he syn hesis o MIPs, namely, co alen , non-co alen , and semi-
co alen app oaches. The co alen app oach in ol es he o ma ion o
e e sible co alen bonds be ween he empla e molecule and mono-
me s be o e polyme iza ion. Then, he empla e is emo ed om he
polyme by clea age o he co esponding co alen bonds, which a e
e o med upon ebinding o he a ge compound (Tu iel and Es eban,
2019). Since co alen imp in ing is s oichiome ic, unc ional monome
esidues exis only in he imp in ed ca i ies. Thus, i minimizes he
p esence o nonspeci ic si es due o he high s abili y o
empla e-monome in e ac ions (Chen e al., 2016; Tu iel and Es eban,
2019). Howe e , he co alen app oach is less lexible ela ed o limi ed
bond o ma ion and clea age eadily e e sible eac ions unde mild
condi ions and i is di icul o each he modynamic equilib ium due o
slow binding and dissocia ion esul ing om s ong co alen in-
e ac ions (Chen e al., 2016; Tu iel and Es eban, 2019). The
non-co alen app oach is based on es ablishing non-co alen in-
e ac ions be ween he empla e and he monome s, such as hyd ogen
bonds, ionic in e ac ions, an de Waals o ces and
π
−
π
in e ac ions.
Usually, he majo in e ac ion is hyd ogen bonding, and his app oach is
by a he mos common because he p epa a ion o MIPs is simple, he e
a e many a ailable monome s, and he binding and emo al s eps a e
apid (Chen e al., 2016; Tu iel and Es eban, 2019). None heless, he
non-co alen imp in ing is less obus because he in e ac ions holding
he complex oge he can be mo e easily dis up ed (Chen e al., 2016).
An in e media e al e na i e is o combine he s abili y o co alen
imp in ing and he as a ge up ake o non-co alen imp in ing, by
ollowing a semi-co alen imp in ing (Chen e al., 2016; Tu iel and
Es eban, 2019). In his case, he empla e is co alen ly bound o a
unc ional monome , bu he ebinding is based only on non-co alen
in e ac ions (Chen e al., 2016).
The e a e also a a ie y o me hods a ailable o p epa ing he
imp in ed polyme s, esul ing in di e en o ma s, and he choice is
usually in insic o he inal applica ion. MIPs can be p epa ed as bulk
polyme monoli hs, equi ing subsequen mechanical g iding o be
ob ained al eady in mic osizes using me hods such as p ecipi a ion and
emulsion polyme iza ions o sol-gel p ocesses. O he e y in e es ing
app oaches ha e eme ged wi h he possibili y o su ace imp in ing,
su ace g a ing, solid-phase syn hesis, click-chemis y, elec o-
polyme iza ion, among plen y examples ha can be ound in he li e -
a u e (Chen e al., 2016; Re aa e al., 2019; Vanecko a e al., 2020).
3. Applica ions
The i s and mos success ul applica ion ield o MIPs is in sepa a ion
echnologies. The MIPs a e used as so ben s in solid-phase ex ac ion
due o hei s uc u al p edic abili y, he mal and chemical s abili y.
These ea u es a e essen ial so ha di e en ypes o composi es can be
o med, and hus applied o selec i ely concen a e samples in biolog-
ical, pha maceu ical, and en i onmen al ields. This p ocedu e is
di ec ly coupled wi h speci ic analy ical sys ems, such as high-
pe o mance liquid ch oma og aphy, minimizing sample manipula-
ion, educing he loss o analy es, he isk o con amina ion and he ime
o p e- ea men o he samples. The aim is o inno a e he solid-phase
ex ac ion me hod owa ds ewe s eps, ha ul ima ely lead o
simplici y, au oma ion, and minia u iza ion. Addi ionally, when he
sample is complex and he p esence o in e e en s may p e en quan-
i ica ion by s anda d ch oma og aphic echniques, a cus omized sample
ea men s ep can be made wi h his echnology p io o he inal
de e mina ion (Gao e al., 2020; Qiao e al., 2006; Tamayo e al., 2007;
Vasapollo e al., 2011).
Ca alysis is ano he in e es ing esea ch ield wi h many po en ials
o MIPs. Enzymes, na u ally and e icien ly, ca alyze a la ge a ie y o
chemical eac ions, due o he speci ic in e ac ions in he ac i e si e
be ween key amino acid esidues and subs a es. Howe e , he e a e
some limi a ions o hei wide applica ions usually connec ed o hei
low s abili y in o ganic sol en s, as well as in ex eme empe a u es and
pH. Fo his eason, when sea ching o a i icial mimics ha could
o e come hese cons ain s, bu a he same ime main ain he expec ed
selec i i y and speci ici y p o ided by na u al enzymes, MIPs cons i u e
an e icien echnology. Wi h MIPs, i is possible o ob ain polyme s ha
display enzyme-like ca aly ic ac i i y by imp in ing analogues o sub-
s a es o ansi ion s a es. The e o e, hese syn he ic polyme s a e
ce ainly o g ea in e es when de eloping enzyme mimics, elying on
hei he mal and chemical s abili y, being easily p oduced o indus ial
applica ions. None heless, he posi i e p ope ies o MIPs as ca alys s,
migh also ep esen a disad an age. Namely, he igh subs a e binding
and he igidi y o he ecogni ion pocke may esul in low eac i i y
and p oduc inhibi ion (D amou and Ta annum, 2016; Mi a a and
Resmini, 2015; Vasapollo e al., 2011; Wul , 2001).
In ecen yea s, he applica ion o MIPs in d ug deli e y sys ems has
been g owing apidly. The esea ch on s imuli- esponsi e MIPs and he
use o biocompa ible polyme ic ma e ials has many en husias ic
Fig. 2. Illus a ion o he nume ous empla es ha ha e been used o de elop MIPs. Copy igh 2020, Rep oduced unde he e ms and condi ions o he C ea i e
Commons A ibu ion 4.0 License (El-Schich e al., 2020).
M.S. Amo im e al.
Biosenso s and Bioelec onics: X 10 (2022) 100131
4
p ospec s in hyb id biomedical in i o app oaches (Chen e al., 2015;
El-Schich e al., 2020; Haup , 2001; Iskie ko e al., 2016; Saylan e al.,
2017; Xu e al., 2011). The memo y nanoca i ies ha could be loaded
wi h a my iad o molecula compounds and be o -loaded h ough
a ious ou es o adminis a ion (e.g., ocula , de mal, in a enous, e c.)
ha e eno mous po en ial bo h o imaging and he apy (Vanecko a
e al., 2020).
MIP-based biosenso s, o sensi i e, apid, low-cos poin -o -ca e
diagnos ics, is also a huge ield o in e es , as MIPs can be combined wi h
se e al di e en ansduce app oaches in a wide a ie y o senso
pla o ms. Biosenso s combine a bio- ecogni ion elemen wi h a signal
p ocesso and a sui able ansduce app oach, which can be o elec o-
chemical, mass, o op ical na u e, among many o he s, o de ec ing
subs ances and moni o ing biological in e ac ions (Fig. 1) (Ca doso
e al., 2018; Chen e al., 2020). The bio-based ecogni ion elemen in-
cludes a cap u e compound ha binds o/in e ac selec i ely wi h he
a ge analy e. In his sense, MIPs can be used as excellen ecogni ion
elemen s due o hei high selec i i y, sensi i i y, long- e m s abili y,
and chemical ine ness (Vasapollo e al., 2011).
4. Vi us imp in ed polyme s
In ec ious diseases widely dissemina ed by pa hogenic mic oo gan-
isms, namely i uses, a e o pa icula conce n. Howe e , despi e huge
p og ess in diagnos ics, ea men and p e en ion, hese diseases a e s ill
a se ious global heal h isk. Fo his eason, ea ly diagnosis o i uses is
essen ial o clinical and poin -o -ca e applica ions. The ools used o
de ec ion o i uses a e mainly based on enzyme-linked immunoso ben
assay and polyme ase chain eac ion (PCR) ampli ica ion (Boonham
e al., 2014). These me hods a e ecognized by hei high sensi i i y, bu
hey a e s ill ime-consuming, wi h high p oduc ion cos s, and equi e
ope a ion by a specialis (Boonham e al., 2014; Cui e al., 2020).
Molecula imp in ing can be applied o a wide ange o a ge mol-
ecules, which is one o he many a ac i e ea u es o his echnique
(Fig. 2). The imp in ing o low molecula weigh compounds as a ge
analy es is well es ablished and highly success ul. When i comes o
p epa e selec i e ecogni ion ecep o s o la ge biological a ge s,
using mac omolecula empla es, he e a e inhe en challenges such as:
(a) he bulkiness o he empla e molecules is connec ed o slow di u-
sion o he molecula ca i ies, making he esponse o he MIP senso
undesi ably long; (b) cha ged unc ional monome s may be less e icien
in he imp in ing o high molecula weigh compounds; (c) he s ep o
empla e emo al is ha de , as i canno be emo ed smoo hly om he
imp in ed ca i ies, hus lowe ing he deso p ion e iciency o he em-
pla e (El-Schich e al., 2020; Haup , 2001; Iskie ko e al., 2016; Saylan
e al., 2017).
These di icul ies a e some imes su passed by epi ope imp in ing, i.
e., a small agmen o he mac omolecule is imp in ed aiming o c ea e
s onge in e ac ions. None heless, his app oach also has in insic lim-
i a ions ha may lead o less speci ic in e ac ions, e.g., unsui able se-
lec ion o epi ope leng h and s uc u e (Die l e al., 2021; Zhang e al.,
2021). A emp ing o g asp he complexi y o mac omolecules in a
polyme ic image o he whole, p ese ed in he imp in ed subs a es,
can be pa icula ly ele an o imp o e he ecogni ion o i us and
o he biological a ge s wi h sizes a he nanoscale.
Pa icula ly o he de ec ion o in ac i uses, he applica ion o
MIPs can ul ima ely be a way o o e come he limi a ions o cu en
me hodology, and in e es in his ield o esea ch has been h i ing
(A zal e al., 2017; Al in as, 2016; Cui e al., 2020; Jamalipou Sou i
e al., 2021). In e es ing examples o MIPs de eloped by empla ing
whole i uses and in eg a ion o hose ma e ials wi hin di e se ans-
duce schemes a e discussed in he ollowing sec ions.
4.1. Tobacco mosaic i us
Tobacco mosaic i us (TMV) is he i s obse ed and epo ed i us
disease, which has been well-s udied and cha ac e ized. This i us has
he po en ial o damage he lea es o di e en plan s, pa icula ly o-
bacco, being esponsible o obacco mosaic. TMV is s able in a ious
condi ions o pH, empe a u e, sol en s, and educing agen s. The TMV
i ion is a odlike i us wi h a leng h o 300 nm and diame e o 18 nm,
composed o 2130 p o ein subuni s ha o m a helical s uc u e (Hema
e al., 2019; Soui i e al., 2019).
The e a e se e al wo ks o TMV de ec ion based on di e en
imp in ing me hods. Fo example, a MIP app oach elied on p oducing a
lexible non-co alen TMV imp in ed polyme hyd ogel o polyallyl-
amine c osslinked wi h e hylene glycol diglycidyl e he using TMV as
empla e (Bolisay and Ko inas, 2010). None heless, mos s udies ound
in he li e a u e a e based on su ace imp in ing. One MIP has been in-
eg a ed on a mic o luidic biochip and he de ec ion elied on
con ac -less bioimpedance spec oscopy (Bi nbaume e al., 2009). The
MIP was assembled by su ace imp in ing using a i us s amp ha was
p essed in o a co-polyme o me hac ylic acid and N- inylpy olidone,
spin-coa ed on he de ice. One o he ad an ages o combining mic o-
luidics wi h he MIP had o do wi h he p ecise con ol o e luid dy-
namic shea o ces. Thus, i was possible o s udy i al binding a ini y
and dissocia ion kine ics, and he de eloped chip p esen ed as
esponse imes and eusabili y (Bi nbaume e al., 2009). In e es ingly,
such s amping echnique has been explo ed in o he wo ks oge he wi h
mass-sensi i e measu emen s using qua z c ys al mic obalances (QCM)
(Dicke e al., 2003, 2004, 2003; Hayden e al., 2003, 2006). In such
s amping me hod, he s amp and he polyme coa ing a e p epa ed
sepa a ely, and he inal ma e ial is ob ained by mechanically p essing
he wo subs a es oge he (Hayden e al., 2006). TMV was success ully
s amped on p e-polyme ized mix u es o me hac ylic acid, s y ene and
di inylbenzene, ollowed by UV cu ing and emo al o i uses. The
ca i ies on he imp in ed polyme su ace ecognized he i us and he
senso e ec s we e obse ed on QCM (Dicke e al., 2003). The
esul ing sensi i e laye con aining he su ace pa e ning was gene a ed
di ec ly on gold elec odes. Using his ype o senso s and polyme s
based on ac ylic acid and e hylene glycol dime hac yla e (EGDMA),
TMV could be de ec ed in a ange o concen a ions om 100 ng mL
−1
o
1 mg mL
−1
, wi hin minu es (Dicke e al., 2004). When pho o-
imp in ing TMV on he su ace o an azopolyme , he immobiliza ion
a e pho oi adi ion was s udied by a omic o ce mic oscopy (AFM) and
by an immunological enzyme luminescence me hod (Ikawa e al., 2010).
The AFM analysis e ealed ha TMV g adually embeds in o he azo-
polyme as a g oo e benea h he i us inc eases du ing i adia ion, i.e.,
as he azopolyme su ace de o ms complemen a y o he shape o TMV.
Also, he luminescence assay e ealed ha he immobiliza ion e iciency
inc eased p opo ionally o he pho oi adia ion ime. Thus, he pho o-
physical induced change in he su ace shape and he isome iza ion o
he azo-dyes enables o imp in bo h he opog aphical ea u e and he
su ace cha ac e is ics o i us like TMV (Ikawa e al., 2010).
4.2. Tobacco nec osis i us
Tobacco nec osis i us (TNV) is a 26 nm diame e nec o i us, which
is no longe associa ed wi h diseases o obacco, bu wi h bean s ipple
s eak and ulip nec osis diseases. This wo ldwide sp ead i us is
ansmi ed by he aqua ic ungus Olpidium b assicae and o en occu s in
i iga ed soils and g eenhouses (Palukai is, 2017; Tolin, 2008).
Wanka e al. (2016) o med molecula imp in s o he TNV wi hin
poly hiophene nano ilms o app oxima e 200 nm hickness, which ha e
been elec ochemically deposi ed on o conduc ing gold su aces. Upon
ebinding, he TNV poly hiophene complex changes he luo escence
in ensi y o he nano ilm p opo ionally o he concen a ion o TNV. I
was shown ha he nano ilm esponds o TNV wi hin 2 min in he
0.1–10 ng L
−1
ange and wi h a limi o de ec ion (LOD) o 2.29 ng L
−1
.
Mo eo e , he selec i i y was es ed using TMV, which is od-shaped and
bigge han TNV, and he senso esponse showed o be selec i e o TNV.
This wo k has demons a ed he po en ial o luo escence o speci ic,
M.S. Amo im e al.
Biosenso s and Bioelec onics: X 10 (2022) 100131
5
label- ee and apid de ec ion o TNV in wa e esou ces using nano ilm
senso s (Wanka e al., 2016).
4.3. Adeno i us
Adeno i us (AdV) i ion has a unique icosahed al shape wi h abou
90 nm in diame e . AdVs cause espi a o y, ocula , u ina y ac and
gas oin es inal, ansmi able, and some imes highly con agious, in-
ec ions. These in ec ions a e gene ally sel -limi ing, bu o unknown
easons, hey can lead o local epidemics. Howe e , he e a e cu en ly
no a ailable he apies p o ed e ec i e an i-AdV. Fo his eason, he e is
an u gency o ha e apid de ec ion and ea ly diagnosis o his i us
(G ebe and Fla , 2019).
Al in as e al. (2015) epo ed a no el MIP echnology o speci ic
and sensi i e ecogni ion o AdVs based on MIP nanopa icles coupled o
su ace plasmon esonance de ec ion. In his wo k, MIP nanopa icles
we e p oduced by a solid-phase syn hesis me hod whe e glass beads
we e used as solid suppo o he immobiliza ion o he a ge AdVs,
while a mix u e o monome s, namely N-isop opylac ylamide, ac ylic
acid, N,N′-me hylenebisac ylamide, N- e -bu ylac ylamide, and
N-(3-aminop opyl) me hac ylamide hyd ochlo ide, was used o poly-
me iza ion. The ecogni ion o AdVs was s udied in a concen a ion
ange o 0.01–20 pmol L
−1
, and a LOD o 0.02 pmol L
−1
was ob ained
(Al in as e al., 2015). A di e en syn he ic s a egy o AdVs cap u e
was p oposed by using su ace-imp in ed co e-shell pa icles. The ma-
e ial was ob ained by immobilizing he i uses on he su ace o
mic ome e silica pa icles, ollowing by co-polyme iza ion o selec ed
o ganosilanes and emo al o he empla e i uses (Gas e al., 2018)
(Fig. 3). To p e en unspeci ic binding, a p o ein, bo ine se um albumin,
was used as blocking agen . This sol-gel imp in ing me hod yielded
excellen binding a ini y, selec i i y, and egene a ion abili y (Gas
e al., 2018). Mo eo e , he amoun o bound i us was de e mined by
quan i a i e PCR, bo h du ing i us imp in ing and ebinding expe i-
men s, e ealing i s alue conside ing he absence o any p io DNA
isola ion s eps (Gas e al., 2018). A la e expansion o his MIP me h-
odology was p esen ed by Gas e al. (2020) by combining i wi h
luo escence labeling. In his wo k, i was possible o isualize indi idual
i uses a ached o he de eloped imp in ed pa icles by
supe - esolu ion mic oscopies (Gas e al., 2020).
4.4. Japanese encephali is i us
Japanese encephali is i us (JEV) is a posi i e sense single-s and
RNA mosqui o-bo ne la i i us co e ed wi h a i al capsid. This i us
causes Japanese encephali is, a i al encephali is ha a ec s housands
o people e e y yea , mainly in he Asia Paci ic egion. Thus, he e is a
g ea need o quick and low-cos de ec ion me hods o his i us
(Ganeshpu ka e al., 2018; S ikas e al., 2018).
In he wo k by Feng e al. (2018), luo escence de ec ion o JEV was
demons a ed by eso ing o su ace molecula imp in ing on silica
mic osphe es modi ied wi h a luo escen dye, dansyl chlo ide. The
de eloped MIPs, p epa ed wi h (3-aminop opyl) ie hoxysilane
(APTES) and e ae hyl o hosilica e (TEOS), showed o selec i ely
ecognize JEV, by luo escence quenching, in he p esence o hepa i is A
i us (HAV), simian i us 40 and abies i us. Mo eo e , he me hod
p o ed o be sensi i e, gi ing a esponse wi hin 55 min, and wi h a LOD
Fig. 3. Schema ic illus a ion o su ace-imp in ing co e-shell pa icles o AdV ecogni ion (A), and images o scanning elec on mic oscopy o ba e silica pa icles
(a), imp in ed pa icles ob ained by he co-polyme iza ion o o ganosilanes (b) and a e lysis (c), non-imp in ed pa icles (d), and pa icles a e i us ebound (e),
wi h schema ic inse s o AdV (e, ) (B). Copy igh 2018, Ame ican Chemical Socie y, Rep oduced wi h pe mission (Gas e al., 2018).
M.S. Amo im e al.

Biosenso s and Bioelec onics: X 10 (2022) 100131
6
in he picomola ange (Feng e al., 2018). Simila ly, Liang e al. (2016)
de eloped a luo escen senso based on a MIP laye ancho ed on he
su ace o luo escen silica mic osphe es (Liang e al., 2016). In his
s udy, JEV was de ec ed based on luo escence esonance ene gy
ans e be ween he i us as ene gy dono and he luo escen dye
(py ene-1-ca boxaldehyde) as ene gy accep o . Thus, an enhancemen
o luo escence in ensi y occu ed p opo ionally o he concen a ion o
he i us in he ange o 24–960 pmol L
−1
. The LOD was de e mined o
be 9.6 pmol L
−1
, and he selec i i y was demons a ed when es ing
o he i uses, namely HAV, lep osy i us and abies i us (Liang e al.,
2016). Taking ad an age o silica mic opa icles wi h a magne ic co e,
Luo e al. (2019a) de eloped a magne ic su ace molecula ly
imp in ed- esonance ligh sca e ing (RLS) senso o apid and highly
sensi i e de ec ion o JEV (Luo e al., 2019a). The cap u e o JEV by he
imp in ed Fe
3
O
4
@SiO
2
mic osphe es esul ed in an inc ease o he RLS
in ensi y, wi h a esponse ime wi hin 20 min, and LOD o 1.3 pmol L
−1
,
allowing apid and sensi i e de ec ion o JEV in p ac ical applica ions.
Rega ding he selec i i y, he senso demons a ed a selec i e esponse
o JEV when o he i uses we e e alua ed (HAV, dimensionally di e en
abies i us, simian acuola ing i us 40) (Luo e al., 2019a). A di e en
luo escen senso based on a me al-o ganic amewo k (MOF) has also
been p oposed (Yang e al., 2020). The MOF ma e ial (MIL-101) was
coa ed wi h silica and u he inyl- unc ionalized o enable he
imp in ing o JEV using zinc ac yla e as unc ional monome and
EGDMA as c osslinke (Fig. 4). Mo eo e , polye hylene glycol (PEG) was
used as blocking agen . In he p esence o JEV he in ensi y o he
luo escence signal inc eased linea ly in a wide ange o concen a ions
(50 pmol L
−1
o 1400 pmol L
−1
) wi hin 20 min, also p esen ing a good
selec i i y and a low LOD o 13 pmol L
−1
. In addi ion, he MIP pa icles
we e selec i e o JEV when HAV, abies, and lep osy i uses we e es ed
(Yang e al., 2020).
4.5. In luenza i us
In luenza i us belongs o he O homyxo i idae amily and can be
di ided in h ee sub ypes A, B and C, which ha e simila s uc u e bu
di e en an igenic p ope ies. In luenza i uses ha e oughly a sphe ical
shape wi h a size o abou 80–120 nm and con ain a single-s anded
nega i e-sense segmen ed RNA genome. The in ec ion by in luenza A
i us (IAV), which is gene ally ound in humans, is mos common and
se e e. This is a highly con agious ai bo ne disease, and he symp oms
ange om mild a igue o espi a o y ailu e and dea h. This disease
sp eads apidly and g ea ly a ec s he human popula ion globally
wi hin a sho pe iod o ime (Dangi and Jain, 2012).
In he wo k by Rand ian sile isoa e al. (2020), he op ical p ope ies
o gold nanopa icles and he high swelling capaci y o polyol-based
hyd ogels we e used o o m a nanocomposi e o bo h ha changes i s
colou and sh inkage in he p esence o IAV (Rand ian sile isoa e al.,
2020). The hyd ogel was o med by click chemis y using unc ional
dend i ic polyglyce ol cyclooc yne and polye hylene glycol diazide
while sialic acids p o ided he speci ic and high binding a ini y o he
hemagglu inin on IAV. Thus, he empla e IAV (s ain H3N2) mixed wi h
gold nanopa icles unc ionalized wi h sialic acids we e added o he
hyd ogel p ecu so mix u e. The esponsi e imp in ed hyd ogel p o-
duced an op ical and mechanical esponse upon emo al and ebound o
he IAV (Rand ian sile isoa e al., 2020). Di e en MIPs, based on
Fig. 4. Scheme o JEV imp in ing on he su ace o a MOF ma e ial, using zinc ac yla e as unc ional monome and PEG as passi a ing agen , and he de ec ion
p inciple. Copy igh 2020, Else ie , Rep oduced wi h pe mission (Yang e al., 2020).
M.S. Amo im e al.
Biosenso s and Bioelec onics: X 10 (2022) 100131
7
ac ylamide, me hyl me hac yla e, me hac ylic acid and N- inyl-
py olidone, de eloped o he pa hogenic s ain H5N1 ha e been
s udied and op imized bo h by bulk imp in ing suspension copolyme -
iza ion o o m polyme beads and by su ace imp in ing using a i us
s amping and o ming a hin- ilm. These MIPs ecognize IAV, being, in
his way, a iable me hod o he de ec ion o he i us (Sangma e al.,
2017). Ano he in e es ing wo k was de eloped by Wangcha eansak
e al. (2013), which applied a molecula imp in ing s a egy as a
sc eening p o ocol o di e en in luenza A sub ypes (H5N1, H5N3,
H1N1, H1N3 and H6N1), c ea ing MIPs o each sub ype and e alua ing
senso cha ac e is ics on a QCM (Wangcha eansak e al., 2013). The
senso s we e p epa ed on gold elec odes o a dual-elec ode QCM by
spin-coa ing he p e-polyme , consis ing o ac ylamide, me hac ylic
acid, me hyl me hac yla e and N- inylpy olidone, and polyme izing
he mix u e in he p esence o a s amp coa ed wi h he empla e i us.
The senso showed o be sensi i e, leading o LODs as low as 10
5
pa -
icles mL
−1
, and selec i e, allowing o i us sub ype cha ac e iza ion
and apid sc eening (Wangcha eansak e al., 2013). This imp in ing
me hodology was la e used o sc een molecula p obes, o di e en size,
shape and binding a ini ies, which could bind o he i us (H5N1) and
induce a con o ma ional change (Wangcha eansak e al., 2014). The
me hod was success ul in di e en ia ing be ween induced con o ma-
ional e ec s a ising om high and low a ini y ligands because he MIP
binding was p opo ionally a ec ed in compa ison o he ecogni ion o
unmodi ied i us. These esul s sugges a e y in e es ing applica ion o
MIPs o s udy no el inhibi o s and hei mode o ac ion (Wangcha -
eansak e al., 2014). Despi e su ace imp in ing by s amping coupled o
QCM has been highly explo ed o a ious IAV sub ypes (Liebe zei
e al., 2011; Wangcha eansak e al., 2013), ano he s a egy has been
also ad anced h ough he syn hesis o g anula MIPs by p ecipi a ion
polyme iza ion (Sukjee e al., 2017). Fou monome s, namely ac yl-
amide, me hac ylic acid, me hyl me hac yla e and N- inylpy olidone,
we e used o p epa e he MIP. The me hod was conside ed low-cos and
easy as he MIPs can be p oduced in la ge quan i ies, while he ecog-
ni ion abili y o he MIP was es ima ed based on indi ec agglu ina ion
es and also on QCM, he la es demons a ing a be e analy ical pe -
o mance (Sukjee e al., 2017).
4.6. Hepa i is i us
HAV is a non-en eloped single-s anded posi i e-sense RNA i us,
wi h a size o 7.5 kb and a diame e o 27 nm, a membe o he Pico -
na i idae amily ha is sp ead by he aecal-o al ou e, bu he si e o
i us eplica ion is he li e . HAV causes he hepa i is A disease, which is
an acu e in lamma o y condi ion and is cha ac e ized by se e al
symp oms, including ano exia, a igue, weigh loss and jaundice.
Fig. 5. Schema ic illus a ion o MIPs luo escence
senso (A), he me al chela ion and six-membe ed
ing o med be ween he empla e and zinc ac yla e
(B), e ec o indica ed concen a ions o HAV and
HBV on luo escence in ensi y o MIPs (a) wi h an
inse o luo escence images o hyb id MIPs solu ions
unde a 365 nm UV lamp, and he espec i e e ec on
he luo escence in ensi y o NIPs (b) (C). Copy igh
2019, Ame ican Chemical Socie y, Rep oduced wi h
pe mission (Luo e al., 2019b).
M.S. Amo im e al.
Biosenso s and Bioelec onics: X 10 (2022) 100131
8
Howe e , he symp oms o he disease a e he e ogeneous depending on
he age o in ec ion, anging om silen in lec ions mainly in young
child en o classical hepa i is in olde age g oups, and a al cou ses o he
disease also occu (A e ho e al., 2015; Do zaue , 2008; Gup a, 2018).
The MIP echnology has also been p o i ing om he use o s imuli-
esponsi e polyme s. In a wo k by Liu e al. (2017), he use o a he -
mosensi i e polyme made o N-isop opylac ylamide enabled he
de elopmen o an imp in ed polyme on he su ace o silica pa icles as
suppo ma e ial o ecogni ion o HAV (Liu e al., 2017). The speci ic
cap u e o he i us occu ed a 40 ◦C and he elease a 20 ◦C, i.e., a he
lowe empe a u e he polyme swelled and a he highe one he
polyme sh inked. The senso pe o mance was ollowed by RLS in-
ensi y, which inc eased upon HAV de ec ion. The senso showed o be
selec i e when es ed in he p esence o in e e ing i uses, namely
hepa i is B i us (HBV), abies i us and JEV, and demons a ed a e y
good LOD o 1.1 pmol L
−1
(Liu e al., 2017). Likewise, Luo e al. (2020)
applied MIP nanop obes o he selec i e de e mina ion o HAV h ough
RLS echnique, ob aining a lowe LOD o 0.1 pmol L
−1
and a linea
concen a ion ange o 0.02–2.0 nmol L
−1
(Luo e al., 2020). In his
wo k, he pH-sensi i e imp in ed polyme dime hylaminoe hyl me h-
ac yla e was p epa ed (swelling wi h pH dec ease) on he su ace o a
MOF suppo . Mo eo e , he pH- esponsi e MOF nanocomposi e p e-
sen ed a selec i e esponse, which indica ed i s po en ial abili y o
de e mine HAV in eal applica ions (Luo e al., 2020). An enhancemen
o RLS in ensi y has been also employed in o he wo ks whe e MIPs we e
p oduced by su ace imp in ing o silica o magne ic pa icles (Yang
e al., 2017; Zhang e al., 2018). In bo h hese wo ks, he
sel -polyme iza ion abili y o dopamine, c ea ing a polydopamine-based
MIP, was s udied as a biomime ic mussel-inspi ed app oach, esul ing in
selec i e and sensi i e senso s wi h LODs in he picomola ange (Yang
e al., 2017; Zhang e al., 2018). In a di e en ype o s a egy, Luo e al.
(2018) c ea ed a luo escence MIP made om CdTe/CdS quan um do
(QD)-based silica nanopa icles using a sol-gel p ocess. The HAV was
selec i ely cap u ed by he imp in ed polyme laye , and he luo escen
quenching o he QDs was analysed wi hin 20 min. Mo eo e , a linea
ange be ween 0.2 and 1.4 nmol L
−1
and a LOD o 88 pmol L
−1
we e
ob ained (Luo e al., 2018).
Rega ding he HBV, i s genome is a pa ially double-s anded ci -
cula DNA o abou 3.2 kb pai s ha belong o he Hepadna i idae amily
(A e ho e al., 2015; Do zaue , 2008; Gup a, 2018). HBV is esponsible
o he po en ially li e- h ea ening li e in ec ion: hepa i is B. Hepa i is
B is a majo global heal h p oblem ha leads o a wide spec um o li e
diseases, anging om acu e o ch onic hepa i is, ci hosis, and hepa-
ocellula ca cinoma (li e cance ) (Liang, 2009).
Some s udies combine he use o MIPs wi h o he ecogni ion p obes.
An app oach de eloped o de ec HBV employed a dual- ecogni ion
me hod based on MIPs and ap ame s in a sandwich RLS senso (Chen
e al., 2021). The MIP a ge ing HBV was achie ed by su ace imp in ing
made o TEOS on ca bon sphe es as ca ie s, while he o he p obe was
ob ained by modi ica ion o silicon sphe es wi h ap ame s. The use o a
second p obe was mean o imp o e he speci ic ecogni ion o HBV and
o p o ide a second enhancemen o he RLS in ensi y. Wi h his
MIP-HBV-ap ame sandwich, he senso esponse demons a ed high
sensi i i y and good selec i i y, wi h a LOD o 0.011 nmol L
−1
(Chen
e al., 2021).
The e a e also a emp s o pu sue he simul aneous de ec ion o
mul iple i uses. In he wo k o Luo e al. (2019) he use o g een and ed
colou ed QDs enabled such mul iplex analysis o de ec bo h HAV and
HBV (Luo e al., 2019b). The MIPs we e syn hesized using zinc ac yla e
and N-isop opylac ylamide as monome s on he su ace o g een and ed
emi ing QDs coa ed wi h shells o silica (Fig. 5). The luo escence senso
enabled simul aneous de ec ion o HAV (LOD o 3.4 pmol L
−1
) and HBV
(LOD o 5.3 pmol L
−1
) by a dec ease in bo h emission peaks (Luo e al.,
2019b).
4.7. Zika i us
Zika i us (ZIKV) is a posi i e sense, single-s and RNA i us wi h a
genome size o app oxima ely 11 kb. ZIKV is an a h opod-bo n i us
(a bo i us) belonging o he Fla i i idae amily. The ansmission o
ZIKV ypically occu s h ough he bi e o an in ec ed emale mosqui o
du ing i s blood eeding, leading o he appea ance o he ZIKV disease.
I is es ima ed ha mos cases o his disease a e asymp oma ic. How-
e e , i has ecen ly caused ou b eaks and epidemics, being associa ed
Fig. 6. Schema ic ep esen a ion o he MIP p epa ed
o CSFV, using se e al monome s (ac ylamide –
AAM; me hac ylic acid – MAA; me hyl me hac yla e –
MMA; N- inylpy olidone – VP), he c osslinke
(dihyd oxye hylene-bisac ylamide – DHEBA) and he
ini ia o o he polyme iza ion (2,2
′-azobis(iso-
bu y oni ile) – AIBN). The su ace-imp in ed poly-
me enabled o de elop a QCM-based senso .
Copy igh 2020, Rep oduced unde he e ms and
condi ions o he C ea i e Commons A ibu ion BY-
NC-ND 4.0 License (Klangp apan e al., 2020).
M.S. Amo im e al.
Biosenso s and Bioelec onics: X 10 (2022) 100131
9
wi h se e e clinical mani es a ions and congeni al mal o ma ions
(Zanluca and dos San os, 2016).
Rico a e al. (2019) de eloped a po en ial poin -o -ca e diagnos ic
sys em using a chip-based po en iome ic senso inco po a ing he mo-
lecula imp in ing echnology (Rico a e al., 2019). The imp in ing
p ocess was designed by he co-adso p ion o he ZIKV and a
sel -assembled monolaye (SAM) o hyd oxyl- e mina ed alkane hiols
on a gold-coa ed chip. This sys em was able o de ec 10
−1
PFU mL
−1
ZIKV in a bu e ed solu ion, and 10 PFU mL
−1
ZIKV in samples o human
sali a con aining clinical i al loads, hus wi h g ea p ospec s o apid
and accu a e sc eening o ZIKV (Rico a e al., 2019).
4.8. Polio i us
Polio i us is a non-en eloped, posi i e-sense, single-s anded RNA
i us, membe o he amily Pico na i idae, which causes poliomyeli is.
Polio i us is mainly ansmi ed by he aecal-o al ou e and eplica es
in he pha ynx and lowe in es inal ac . Polio i us is en i ely asymp-
oma ic in 90% o indi iduals, howe e , in ewe han 1% o cases he
i us en e s he cen al ne ous sys em, in ec ing and des oying mo o
neu ons, leading o muscle weakness and acu e laccid pa alysis (Cla -
wo hy, 2014; Ga g and Ka s , 2016; T oy and Maldonado, 2012).
Wang e al. (2010) applied su ace molecula imp in ing using SAMs
on a gold-coa ed silicon chip (Wang e al., 2010). The design consis ed in
co-adso bing he empla e wi h hyd oxyl- e mina ed alkane hiol mole-
cules on he me al su ace allowing hem o o m he SAM. A e em-
pla e emo al, he imp in ed ca i ies a e le behind on he senso
su ace. As demons a ed in a simila app oach o ZIKV (Rico a e al.,
2019), his s a egy was success ully applied in he po en iome ic
de ec ion o polio i us, wi h no c oss- eac i i y o AdV (Wang e al.,
2010).
4.9. Classical swine e e i us
Classical swine e e i us (CSFV) o 40-60 nm in diame e , a e
en eloped i us o icosahed al symme y o he amily Fla i i idae, and
he i al genome is a single-s anded posi i e-sense RNA o app oxi-
ma ely 12.3 kb. CSFV in ec ion leads o a b eakdown o he immune
sys em accompanied by a p o-in lamma o y esponse. This disease is
associa ed wi h many symp oms, including se e e lymphopenia and
lymphocy e apop osis, h ombocy openia, pla ele agg ega ion, bone
ma ow deple ion, hymus a ophy, and hymocy e apop osis. Since
classical swine e e is conside ed one o he mos ele an e-eme gen
a al i al diseases in swine, he e is an economic necessi y ha u ges
apid ea ly de ec ion o his i us (Ganges e al., 2020). A MIP ecog-
ni ion elemen has been de eloped o de ec his i us, elying on CSFV
s amping ha was p essed on a p e-polyme o ac ylamide, me hac ylic
acid, me hyl me hac yla e and N- inylpy olidone, spin-coa ed on QCM
gold elec odes (Fig. 6). This senso selec i ely binds CSFV, wi h a LOD
o 1.7
μ
g mL
−1
(Klangp apan e al., 2020).
4.10. Foo and mou h disease i us
Foo and mou h disease i us (FMDV) is a small, icosahed al, non-
en eloped, single-s anded, posi i e-sense RNA i us o app oxima ely
8.3 kb, ha belongs o he amily Pico na i idae. This highly in ec ious
pa hogen causes se ious debili a ing disease, he oo and mou h disease,
in ca le and o he li es ock and wildli e, and is hus conside ed a
ele an e e ina y pa hogen (Malik e al., 2017; Yan e al., 2017).
An elec ochemical polyme iza ion o he oxidized o-aminophenol
ilm wi h FMDV se o ype O on a gold sc een-p in ed elec ode deployed
a new ecogni ion sys em o his i us (Hussein e al., 2019). The
biosenso showed a high selec i i y o FMDV se o ype O in compa ison
o se o ype A, SAT-2, inac i a ed se o ype O, and lumpy skin disease
i us, and a LOD o a ound 2 ng mL
−1
. Besides, he as esponse (5 min)
and he eusabili y o he biosenso , p esen s a p omising, a o dable,
and po able ool ha could be used in he ield (Hussein e al., 2019).
4.11. O he i uses
Cumbo e al. (2013) desc ibed a syn he ic s a egy o p oduce
o ganic/ino ganic nanopa icula e hyb ids con aining i us imp in s on
he su ace. Fo ha , i us-imp in ed pa icles we e p oduced o plan
i uses as models, namely oma o bushy s un i us (TBSV) and u nip
yellow mosaic i us (TYMV) (Cumbo e al., 2013). The i uses we e i s
bound on he su ace o silica nanopa icles, ollowed by incuba ion wi h
a mix u e o o ganosilanes and subsequen polycondensa ion o g ow an
o ganosilica (silsesquioxane) ecogni ion laye , which a e i us
emo al display he ee imp in s. The TBSV and TYMV imp in ed silica
nanopa icles displayed a ema kable selec i i y and a ini y o bo h
i uses, ecognizing hem in wa e a concen a ions down o he pico-
mola ange (Cumbo e al., 2013). A simila app oach was pu sued bu
using i us-like pa icles as a sa e subs i u e o he imp in ing o human
pa hogenic No o i uses, and he MIP showed also an excellen a ini y in
he picomola ange (Syko a e al., 2015).
A no el double imp in ing me hod has been p oposed o he apple
s em pi ing i us (ASPV) (Bai and Spi ak, 2014). The
Fig. 7. Scheme o he bioimp in ing p ocess o ob ain i us esponsi e supe -
ap ame hyd ogels, using polyme izable speci ic ap ame s copolyme ized
wi h N-isop opylac ylamide – NIPAM and ac ylamide – AM as monome s, N,N
′-
me hylenebisac ylamide – MBAA as c osslinke , and ammonium pe sul a e –
APS and N,N,N′,N′- e ame hyle hylenediamine (TEMED) o ini ia ion o he
polyme iza ion in phospha e-bu e ed saline – PBS (A); op ical mic oscopy
images o he esponsi e MIP o ASPV and he co esponding lase di ac ion
pa e ns wi hou i us (a, b) and in he p esence o ASPV (c, d) (B). Copy igh
2014, Wiley-VCH, Rep oduced wi h pe mission (Bai and Spi ak 2014).
M.S. Amo im e al.