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United S ta tes Pa tent [ 1 9 1

| I 1 | | | l l l l l l l l l l l l l l l l l l l l l I l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l
US 0051 65407A
Pa tent Number:
[ 1 1 1 5, 1 65, 407
W il s o n et a l . [ 45] D a te o f Pa tent: No v . 2 4, 1 9 9 2
[ 54] I MPLANTABLE GLUCOS E S ENS OR 8 561 3 41 3 / 1 9 8 7 j a p a n . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 742 54 / l 9 8 7 [ 751 I nv ento rs Geo rg e 8 - w il s o n; D il bir 8 - 8 mm; 1 2 9 69 1 3 3 / 1 9 8 7 3 . 1 1 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . - 9 9 9 / 1 1 1 9
Bria n S . H il l , a l l o f La w rence, K a ns . ; '
D a niel R. Th ev eno t, Pa ris Cedex , OTH ER PUBLI CATI ONS
F ra nce; Ro bert S temberg Th ia is Up dik e et a l . , Th e Enzyme El ectro de , Na ture, v o l .
F ra nce; Gera rd Rea ch , Pa ris Cedex , 2 1 4 J un 1 9 67, p p . 9 3 6_ 9 8 8 _
Emma ; Ya mm Z h a ng La w rence S a l k ind et a l . , I mp ro v ing . . . S ta bil ity , Med. I ns t, v o l .
1 1 1 5- 1 5, No . 2 , Ma n- Ap r. 1 9 2 1 , 1 1 1 9 . 1 2 6- 1 2 7.
[ 73 ] As s ig nee: Th e Univ ers ity o f K a ns a s , La w rence, F ri- ma n, Ex a miner_ _ Lee S . Co h en
K a ns ' Atto rney, Ag ent, o r F irm~ - H o v ey, W il l ia ms , Timmo ns &
[ 2 1 ] Ap p l . No . : 68 2 , 560 Co l l ins
[ 2 2 ] F il ed: Ap r. 9 , 1 9 9 1 [ 57] ABS TRACT
I mp l a nta bl e enzyma tic s ens o rs ( 2 5, 43 , 44) f o r bio ch emi
Rel a ted { 1 5- Ap p l iw tio n D a ta ca l s s uch a s g l uco s e a re p ro v ided h a v ing a n idea l s ize
[ 63 ] Co ntinua tio n- imp a rt o f S er. No . 51 1 , 049 , Ap r. 1 9 , a nd g eo metry f o r o p tio na l l o ng tel ' mil ' l ' l p l a ntmiq 1 a nd
1 9 9 0, a ba ndo ned. ' l inea r res p o ns es o v er th e co ncentra tio n ra ng es o f inter
5 es t. Th e s ens o rs ( 2 5, 43 , 44) incl ude a n el o ng a ted bo dy
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ( 1 0, 2 6, 46) s up p o rting a n indica ting el ectro de S ectio n
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . , 2 04/ 41 5, h a v ing a n a p p ro p ria te enzyme immo bil ized th ereo n to
. _ p res ent a n enzyma tic indica ting s urf a ce ( 2 1 , 3 3 , 54) . A
[ 58 ] F iel d o f S ea rch . . . . . . . . . . . . . . . . . 1 2 8 / 63 5, 2 04/ 403 , 41 5 p ermea bl e s ynth etic p o l ymer membra ne ( 2 4, 42 , 56) is
[ 56] Ref erences Cited a p p l ied o v er th e s ens o r bo dy ( 1 0, 2 6, 46) to p ro tect th e
U. S . PATENT D OCUMENTS
3 , 2 49 , 1 03 5/ 1 9 66 W o o dh o us e . . . . . . . . . . . . . . . . . . . . . . . . 1 2 8 / 63 5
3 , 72 6, 777 4/ 1 9 73 Ma cur . . . . . . . . . . . . . 1 2 8 / 63 5 X
4, 545, 3 8 2 1 0/ 1 9 8 5 H ig g ins et a 1 . 1 2 8 / 63 5
4, 671 , 2 8 8 6/ 1 9 8 7 Go ug h . . . . . . . . . . . . . . . . 1 2 8 / 63 5
F OREI GN PATENT D OCUMENTS
2 5
03 2 01 09 6/ 1 9 8 9 Euro p ea n Pa t. Of f . . . . . . . . . . . . . 1 2 8 / 63 5
01 69 668 1 0/ 1 9 8 2 J a p a n . . . . . . . . . . . . . . . . . . . . . . . . . . 60- 3 3 644 2 / 1 9 8 6 J a p a n . . . . . . . . . . . . . . . . . . , . . . . . . . . . . . . . . . . 1 2 8 / 63 5
enzyme a nd reg ul a te dif f us io n o f a na l yte th ereth ro ug h ,
to ens ure l inea rity o f s ens o r res p o ns e. Th e s ens o rs ( 2 5,
43 ) a re o f ? ex ibl e des ig n a nd ca n be imp l a nted us ing a
ca th eter. Al terna tel y, th e s ens o r ( 44) incl udes a n inter
na l indica ting el ectro de bo dy ( 46) h o us ed w ith in a n
a p ertured, h o l l o w needl e ( 48 ) . A h o l der ( 66) a f f ix ed to
th e needl e ( 48 ) a l l o w s f o r ea s y ma nip ul a tio n a nd im
p l a nta tio n o f th e s ens o r ( 44) .
9 Cl a ims , 3 D ra w ing S h eets
US . Pa tent No v . 2 4, 1 9 9 2 S h eet 1 o f 3 5, 1 65, 407
mi. \\
2 4 ) 1 0
E 2
40
3 3 3 6 3 47 g 3 8
' 2 8
I I I / I / I / I / I / I / I / I / I / l i
3 2 43 / 2 3 ' 3 o
US . Pa tent No v . 2 4, 1 9 9 2 S h eet 2 o f 3 5, 1 65, 407
01 O
|
43 O
I
04 O
I
' 8
I
S ENS OR RES PONS E ( nA/ mmZ )
6
1 o 2 0 s o _
GLUCOS E CONCENTRATI ON ( mM)
F I G. 5.
F I G. 4.
O
O
S ENS OR RES PONS E ( nA/ mM. mmz)
o I in o
o I _ 1 0 2 0 50 4o
S TORAGE PERI OD ( EN/ s )
O
b.
US . Pa tent No v . 2 4, 1 9 9 2 S h eet 3 q r 3 5, 1 65, 407
' 1 l l l l l il l ul l ul - l l l l
Q . . . . . I . . I . . . . . . . " . . . . . . . . .
5, 1 65, 407
1
I MPLANTABLE GLUCOS E S ENS OR
Th is is a co ntinua tio n- in- p a rt o f a p p l ica tio n S er. No .
07/ 51 1 , 049 , ? l ed Ap r. 1 9 , 1 9 9 0, no w a ba ndo ned.
BACK GROUND OF TH E I NVENTI ON
1 . F iel d o f th e I nv entio n
Th e p res ent inv entio n is bro a dl y co ncerned w ith a
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s ubcuta neo us l y imp l a nta bl e enzyma tic s ens o r ch a ra c- 1
terized by s ma l l s ize, o p timum g eo metry a nd l inea rity o f
s ens o r res p o ns e o v er th e co ncentra tio n ra ng e o f inter
es t. Mo re p a rticul a rl y, it is p ref era bl y co ncerned w ith
a n imp l a nta bl e g l uco s e s ens o r o f th is typ e des ig ned to
p ro v ide, in co nj unctio n w ith a s uita bl e s ig na l p ro ces s ing
unit, a current w h ich is p ro p o rtio na l to s ubcuta neo us
g l uco s e co ncentra tio n. I n p ref erred f o rms , g l uco s e s en
s o rs o f th e inv entio n a re ba s ed o n th e enzyme- ca ta l yzed
o x ida tio n o f g l uco s e to g l uco nic a cid a nd h ydro g en
p ero x ide, th e l a tter being mo nito red a mp ero metrica l l y
by th e s ens o rs . -
2 . D es crip tio n o f th e Prio r Art
Th ere h a v e been a g rea t ma ny a ttemp ts in th e p a s t to
dev el o p v ia bl e imp l a nta bl e s ens o rs f o r co ntinuo us in
v iv o mea s urements o f bio ch emica l s . F o r ex a mp l e, co n
s idera bl e ef f o rt h a s been ma de to dev is e rel ia bl e im
p l a nta bl e s ens o rs f o r mo nito ring g l uco s e co ncentra tio ns
in bl o o d. S uch determina tio ns a re us ef ul in a v a riety o f
a p p l ica tio ns , e. g . , in th e trea tment o f dia betics . One
dif f icul ty in p ro v iding a rel ia bl e imp l a nta bl e g l uco s e
s ens o r is th a t g l uco s e l ev el s in th e bl o o ds trea m o f a
p a tient v a ry o n a time ba s is a nd a re no rma l l y dep endent
up o n th e p h ys ica l a ctiv ity o f th e indiv idua l , h is f o o d,
bev era g e a nd s ug a r inta k e, h is meta bo l ic ra te, a nd o th er
indiv idua l ized f a cto rs . F urth ermo re, th e g eo metry o f
th e s ens o r mus t be s uch a s _ to a da p t to imp l a nta tio n in a
l iv ing p a tient. .
Gl uco s e s ens o rs h a v e been p ro p o s ed in th e p a s t
w h ich rel y up o n th e w el l - es ta bl is h ed enzyme- ca ta l yzed
o x ida tio n o f g l uco s e w h erein g l uco s e a nd o x yg en f unc
tio n a s s ubs tra tes f o r th e enzyme g l uco s e o x ida s e in th e
p ro ductio n o f g l uco nic a cid a nd h ydro g en p ero x ide, th e
l a tter being mo nito red a mp ero metrica l l y. S ee, f o r ex
a mp l e, U. S . Pa t. No s . 3 , 53 9 , 455 to Cl a rk a nd 4, 671 , 2 8 8
to Go ug h .
Al th o ug h th e idea o f a n imp l a nta bl e enzyma tic g l u
co s e s ens o r is no t p er s e new , co ns idera bl e dif ? cul ty h a s
been enco untered in p ro ducing rel ia bl e, co s t- ef ? cient
dev ices o f th is ch a ra cter. F o r ex a mp l e, ma ny p ro p o s ed
s ens o r g eo metries a re s imp l y no t rea l is tica l l y imp l a nt
a bl e, a t l ea s t f o r th e p erio ds o f time req uired f o r a de
q ua te cl inica l g l uco s e mo nito ring . Th us , th e dev ices
p ro p o s ed in th e 2 8 8 Go ug h Pa tent, beca us e o f a re
q uirement o f mul tip l e el ectro des ca rried ' w ith in a tubu
l a r needl e, inev ita bl y a re o f s uch dia meter a s to be un
co mf o rta bl e to th e us er a nd no t p ra ctica l f o r ex tended
imp l a nta tio n. F urth ermo re, ma ny p rio r s ens o rs do no t
ex h ibit a s ta bl e a nd l inea r res p o ns e, p a rticul a rl y o v er
ex tended times o f imp l a nta tio n, a nd do no t g iv e a ccu
ra te a nd rel ia bl e res ul ts . F ina l l y, f a brica tio n o f p rio r
g l uco s e s ens o rs h a s p res ented f o rmida bl e dif f icul ties , to
th e ex tent th a t o nl y a bo ut o ne in ? v e s ens o rs p ro duced
by co nv entio na l tech niq ues a re deemed us a bl e. Th is
o bv io us l y rep res ents a co ns idera bl e inef f iciency, to th e
p o int th a t no trul y s ucces s f ul imp l a nta bl e g l uco s e s en
s o r h a s h ereto f o re been p ro duced o n a l a rg e s ca l e.
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3 0
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S UMMARY OF TH E I NVENTI ON
Th e p res ent inv entio n o v erco mes th e p ro bl ems o ut
l ined a bo v e, a nd p ro v ides a g rea tl y imp ro v ed enzyma tic
s ens o r s p eci? ca l l y des ig ned f o r l o ng - term imp l a nta tio n
in a p a tient. Th e s ens o r is a da p ted f o r p o s itio ning in a n
env iro nment ch a ra cterized by th e p res ence o f bio l o g i
ca l mo l ecul es w h ich a re s ubs tra tes f o r o r p ro ducts p ro
duced by enzymes , in o rder to determine th e p res ence
o f s uch bio l o g ica l mo l ecul es . W h il e th e p rincip l es o f th e
inv entio n ma y be us ed in th e f a brica tio n o f g l uco s e
s ens o rs , th e inv entio n is no t s o l imited. I ndeed, th e s en
s o rs in a cco rda nce w ith th e inv entio n ma y be p ro duced
us ing a w ide v a riety o f immo bil ized enzymes , f o r th e
detectio n o f a n eq ua l l y l a rg e number o f a na l ytes . Ex em
p l a ry enzymes a nd th eir co rres p o nding s ubs tra tes a re
g iv en in US . Pa t. No . 4, 72 1 , 677 to Cl a rk , a nd th is p a
tent is inco rp o ra ted by ref erence h erein.
I n a ny ev ent, th e enzyma tic s ens o rs in a cco rda nce
w ith th e inv entio n p ref era bl y a re in th e f o rm o f a n
el o ng a ted bo dy s up p o rting a t l ea s t a n indica ting el ec
tro de, w ith th e indica ting el ectro de p res enting a s ectio n
a da p ted f o r ex p o s ure to th e bio l o g ica l env iro nment.
Th e indica ting el ectro de s ectio n h a s a n enzyme o p era
bl y immo bil ized th ereo n to p res ent a n enzyma tic indi
ca ting s urf a ce. A number o f v a ria nts a re p o s s ibl e f o r th e
ref erence el ectro de. F o r ex a mp l e, us e ma y be ma de o f
a n ex terna l l y a p p l ied el ectro ca rdio g ra m s k in el ectro de
( a n 8 mm dis k co v ered w ith s il v er ch l o ride a nd a v a il
a bl e a s Mo del E- 2 43 f ro m th e Ph ymep Co mp a ny, 2 1
Rue Ca mp o f o rmio , Pa ris , F ra nce) , o r a ref erence el ec
tro de w h ich is imp l a nted w ith th e indica ting el ectro de.
I n o ne s p eci? c embo diment emp l o ying a n imp l a nted
ref erence el ectro de, th e indica ting s urf a ce o f th e indi
ca ting el ectro de a nd th e ref erence el ectro de a re l a ter
a l l y s p a ced a p a rt a l o ng th e l eng th o f th e bo dy a nd ea ch
s ubs ta ntia l l y circums cribes th e l a tter a nd is s ubs ta ntia l l y
ex p o s ed to th e bio l o g ica l env iro nment w h en th e s ens o r
is p l a ced th erein. Us e o f s uch circumf erentia l l y ex tend
ing enzyma tic indica ting s urf a ces a nd ref erence el ec
tro des s ectio ns is bel iev ed to be a n imp o rta nt a s p ect o f
th is embo diment. Al terna tel y, th e ref erence el ectro de
s ectio n ma y co mp ris e a co nductiv e s a l t bridg e circum
s cribing th e bo dy a nd l ying in a p l a ne tra ns v ers e to th e
l o ng itudina l a x is o f th e bo dy; in th is ca s e, a ref erence
el ectro de is p l a ced in el ectrica l co nta ct w ith th e s a l t
bridg e, th ro ug h us e o f a buf f ered el ectro l yte. I n a no th er
embo diment, th e ref erence el ectro de is s imp l y p l a ced
a dj a cent th e indica ting el ectro de a s a p a rt o f th e o v era l l
s ens o r.
I n p ref erred p ra ctice, th e s ens o r bo dy a dv a nta
g eo us l y co mp ris es a n el ectrica l l y co nductiv e no bl e
meta l ( e. g . p l a tinum o r p l a tinum- iridium) el ectro de co v
ered w ith el ectrica l l y ins ul a tiv e ma teria l , w ith a p o rtio n
o f th is ma teria l remo v ed f ro m th e el ectro de to de? ne a n
enzyme- receiv ing zo ne. Th us , a s h o rt l eng th o f Te? o n
( p o l ytetra tl uo ro eth yl ene) co a ted p l a tinum~ iridium w ire
ma y be p ro v ided, w ith a s h o rt s ectio n o f th e ins ul a tio n
remo v ed intermedia te th e ends o f th e w ire, s o th a t re
s p ectiv e s eg ments o f th e ins ul a ting ma teria l a re o n o p
p o s ite s ides o f a nd de? ne a reces s ed enzyme- receiv ing
circumf erentia l zo ne. Al terna tel y, th e endmo s t p o rtio n
o f th e Te? o n ma y be remo v ed, l ea v ing a p ro truding
ex p o s ed s tretch o f w ire w h ich de? nes th e enzyme
receiv ing zo ne. An enzyme is o p era bl y immo bil ized o n
th e ex p o s ed s ectio n o f th e p l a tinum- iridium w ire, by
k no w n mea ns s uch a s a ds o rp tio n o f th e enzyme o n a
5, 1 65, 407
3
cel l ul o s e a ceta te o r Na ? o n l a yer ( 1 - 3 micro ns th ick
nes s ) , f o l l o w ed by cro s s l ink ing w ith g l uta ra l deh yde.
Ano th er imp o rta nt a s p ect o f th e p res ent inv entio n
res ides in th e p ref erred us e o f a s ynth etic p o l ymer mem
bra ne dis p o s ed o v er th e enzyma tic indica ting s urf a ce to
s erv e a s a p ermea bl e p ro tectiv e l a yer. I n p a rticul a r, a
l a yer o f p o l yureth a ne is a dv a nta g eo us l y a p p l ied a s a
th in co a ting o v er a t l ea s t th e indica ting s urf a ce ( a nd
p ref era bl y th e entire indica ting el ectro de) in o rder to
p ro tect th e enzyma tic rea ctio n s urf a ce f ro m th e bio l o g i
ca l env iro nment. Mo reo v er, th is l a yer p ro v ides a dif f u
s io na l ba rrier f o r g l uco s e w h ich s l o w s do w n th e ? o w o f
g l uco s e a nd crea tes a l inea r s ens o r res p o ns e o v er th e
co ncentra tio n ra ng es o f interes t. I n p a rticul a r, in o rder
to a ch iev e th e des ired l inea r res p o ns e, us e is ma de o f a n
a ctiv e enzyme l a yer a nd a rel a tiv el y th in p ro tectiv e
membra ne. I t is imp o rta nt th a t th e membra ne reg ul a te
th e p a s s a g e o f mo l ecul es th ereth ro ug h to a n ex tent th a t
th e enzyma tic rea ctio n betw een th e indica ting s urf a ce
a nd th es e mo l ecul es is determined by th e ra te o f dif f u
s io n th ro ug h th e membra ne, a nd no t th e enzyma tic
rea ctio n k inetics . I n p ra ctice us ing th e meth o ds o f s en
s o r co ns tructio n h erein des cribed, a n o p tima l ba l a nce
betw een th e co mp eting g o a l s o f l inea r res p o ns e a nd
s ens itiv ity a nd res p o ns e times ma y be a ch iev ed.
Th e us e o f a n a dditio na l , neg a tiv el y ch a rg ed inner
membra ne l a yer immedia tel y a dj a cent th e Pt- I r w ire
a l s o reta rds th e dif f us io n o f neg a tiv el y ch a rg ed s p ecies
( e. g . a s co rba te a nd ura te) in th e bio l o g ica l env iro nment
w h ich a re interf ering s p ecies . Of co urs e, th is inner
membra ne do es no t s ig ni? ca ntl y ex cl ude h ydro g en
p ero x ide, a nd el ectrica l l y neutra l s p ecies .
Al th o ug h th e th ick nes s o f th e o utermo s t p o l yure
th a ne membra ne h a s no t been s p eci? ca l l y a s certa ined, it
is es tima ted th a t th e membra ne h a s a th ick nes s o f f ro m
a bo ut 5 to 1 0 micro ns in th e p ref erred g l uco s e s ens o rs
h ereo f .
Th e s ens o rs des cribed a bo v e a re, by v irtue o f th eir
co ns tructio n, rel a tiv el y ? ex ibl e a nd th eref o re co mf o rt
a bl e in us e. H o w ev er, th is s a me ch a ra cteris tic ? ex ibil ity
ma k es it neces s a ry to emp l o y a ca th eter to imp l a nt th e
s ens o rs . I n a n a l terna tiv e embo diment, s ens o rs ma y be
p ro v ided w h ich ca n be rea dil y imp l a nted w ith o ut th e
need o f a ca th eter, ev en by th e p a tient h ims el f . I n s uch
embo diments , us e is ma de o f a n el o ng a ted, tubul a r,
meta l l ic h o us ing , typ ica l l y a co nv entio na l h yp o dermic
needl e; th e s ens o r a p p a ra tus is ins erted w ith in th e nee
dl e, a nd incl udes a n indica ting el ectro de h a v ing a s ec
tio n th ereo f p ro v ided w ith immo bil ized enzyme. I n
o rder to ex p o s e th e enzyme to th e bio l o g ica l env iro n
ment, th e needl e s idew a l l is a p ertured in reg is try w ith
th e enzyme. A h o l der is a l s o p ro v ided a dj a cent th e
rea rw a rd end o f th e needl e bo dy in o rder to f a cil ita te
ma nip ul a tio n a nd ins ertio n o f th e s ens o r. Th is h o l der
a dv a nta g eo us l y is in th e f o rm o f a tra ns v ers el y ex tend
ing ? a g - l ik e p l a s tic bo dy s ecured to th e needl e h o us ing .
Th e inv entio n a l s o co mp reh ends a no v el meth o d o f
a p p l ying th e p o l yureth a ne membra ne des cribed p rev i
o us l y. Th a t is to s a y, a rea l dif f icul ty in th e p ro ductio n
o f enzyma tic s ens o rs s tems f ro m th e dif ? cul ty o f a p p l y
ing v a rio us ma teria l s unif o rml y to a v ery s ma l l , imp l a nt
a bl e dev ice. Th is dif ? cul ty h a s been o v erco me in th e
co ntex t o f th e p res ent inv entio n, by a p p l ying to th e
s ens o r s urf a ce a w el l - de? ned v o l ume o f a p o l ymer dis
s o l v ed in a n o rg a nic s o l v ent s uch th a t th e ? l m is uni
f o rml y dis tributed a cro s s th e s urf a ce, I n p ra ctice, th is
meth o d is ca rried o ut by p ro v iding a w ire l o o p , a nd
h o l ding th e co a ting l iq uid in th e l o o p by s urf a ce tens io n
1 0
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to f o rm th e des ired p o l ymer s o l utio n dro p l et, f o l l o w ed
by p a s s ing th e el ectro des th ro ug h th e l o o p to a ch iev e
unif o rm co a ting a l o ng th e l eng th o f th e s ens o r bo dy.
Th e enzyma tic s ens o rs o f th e inv entio n h a v e a n idea l
g eo metry f o r imp l a nta tio n. Genera l l y s p ea k ing , th e
? ex ibl e units no t h o us ed w ith in a needl e a re eq uiv a l ent
in s ize a nd s h a p e to a 2 6- g a ug e needl e ( i. e. , a bo ut 0. 45
mm. o uts ide dia meter) . Mo reo v er, th eir g eo metry p er
mits th e rep ro ducibl e dep o s itio n o f ? l ms a nd ma teria l s
a nd a l l o w s ca ref ul co ntro l o f th e a mo unt a nd o rienta
tio n o f th e enzyme o nto th e indica ting el ectro de. F i
na l l y, th e p ref erred s ens o rs a re ef f ectiv el y ca p p ed
w ith ins ul a tio n ( T ef l o n) w h ich p rev ents th e s ens o rs
f ro m p enetra ting f urth er into th e tis s ue th a n is req uired.
Th us , th e ins ertio n o f th e s ens o r ca us es minima l tra uma
to th e tis s ue a nd to th e s ens o r its el f . Th e s ens o r ca n ? ex
l a tera l l y, a nd th is a g a in minimizes tis s ue da ma g e ca us ed
by mo v ement o f th e p a tient.
I n th e ca s e o f imp l a nta bl e g l uco s e s ens o rs , res p o ns e
times o f l es s th a n tw o minutes a nd l inea rities o v er g l u
co s e' co ncentra tio ns o f 0- 2 5 mM ca n be a ch iev ed. At
th e s a me time, th ro ug h us e o f th e f a brica tio n tech niq ues
o f th e inv entio n, th e rej ectio n ra te up o n initia l ma nuf a c
ture is dra s tica l l y reduced.
I n th e ca s e o f s ens o rs receiv ed w ith in a needl e h o us
ing , s uch ca n be rea dil y ma nip ul a ted by th e p a tient f o r
imp l a nta tio n p urp o s es . Th es e s ens o rs typ ica l l y h a v e a
s l ig h tl y l a rg er dia meter th a n th e ? ex ibl e s ens o rs de
s cribed p rev io us l y, but a re no t s o l a rg e a s to ca us e s ig
ni? ca nt dis co mf o rt. Th is rel a tiv el y s ma l l s ize is a s s ured
beca us e o f th e s ens o r co ns tructio n, ma k ing us e o f a
s ma l l Te? o n- co a ted Pt- I r w ire a nd immo bo l ized en
zyme.
BRI EF D ES CRI PTI ON OF TH E D RAW I NGS
F I G. 1 is a n enl a rg ed, s ectio na l v iew il l us tra ting a
g l uco s e s ens o r in a cco rda nce w ith th e inv entio n;
F I G. 2 is a n enl a rg ed, s ectio na l v iew o f a no th er g l u
co s e s ens o r in a cco rda nce w ith th e inv entio n;
F I G. 3 is a g ra p h s h o w ing th e l inea r s ens o r res p o ns e
o f th e F I G. 1 g l uco s e s ens o r o v er a g l uco s e co ncentra
tio n ra ng e o f O- 2 5 mM;
F I G. 4 is a g ra p h il l us tra ting th e s to ra g e s ta bil ity o f
th e F I G. 1 g l uco s e s ens o r;
F I G. 5 is a s ectio na l v iew dep icting a no th er s ens o r
embo diment w h erein th e indica ting el ectro de is h o us ed
w ith in a n imp l a nta bl e needl e;
F I G. 6 is a p ers p ectiv e v iew o f th e s ens o r il l us tra ted
in F I G. 5; a nd
F I G. 7 is a p ers p ectiv e v iew o f a n embo diment s imi
l a r to th a t o f F I G. 6, but dep icting th e us e o f a n imp l a nt
a bl e ref erence el ectro de.
D ES CRI PTI ON OF TH E PREF ERRED
EMBOD I MENTS
Th e f o l l o w ing ex a mp l es il l us tra te th e co ns tructio n o f
g l uco s e s ens o rs dep icted in F I GS . 1 a nd 2 , a nd a re
des cribed w ith p a rticul a r ref erence to th es e dra w ing s . I t
w il l be unders to o d, h o w ev er, th a t th e ex a mp l es a re
il l us tra tiv e o nl y, a nd no th ing th erein s h o ul d be ta k en a s
th e l imita tio n up o n th e o v era l l s co p e o f th e inv entio n.
EXAMPLE l - F I G. 1
One end o f a 1 0 cm s ectio n 1 0 o f Medw ire Co rp o ra
tio n Te? o n- Co a ted p l a tinum- iridium w ire is p ro v ided.
Th e s ectio n 1 0 incl udes a centra l p l a tinum- iridium w ire
1 2 ( 0. 1 8 mm ed) a nd a co a ting o f ins ul a tiv e Te? o n 1 4
( 0. 03 5 mm th ick nes s ) th erea ro und. Th e centra l w ire 1 2
5, 1 65, 407
5
f o rms th e indica ting el ectro de f ro m th e s ens o r. A ca v ity
1 6 ( 1 - 3 mm in l eng th ) is f o rmed in th e w ire 1 0 a s s h o w n
in F I G. 1 . Th is is a ch iev ed by ? rs t p utting a circul a r cut
o n th e Te? o n co a ting w ith a p a p er cutter a nd th en
p ul l ing th e Tef l o n o ut to crea te a ca v ity o f a bo ut 1
mil l imeter in l eng th , ex p o s ing a co rres p o nding s ectio n
o f th e w ire 1 2 . Th e ex ces s Te? o n ex tending beyo nd th e
l ef t end o f th e w ire 1 2 is th en trimmed o f f w ith th e
cutter. . .
Th e ref erence el ectro de 1 8 is f o rmed o n th e Te? o n
s urf a ce, a bo ut l . 5 mil l imeters to th e rig h t o f th e ex p o s ed
p l a tinum iridium s urf a ce a s v iew ed in F I G. 1 . A th in
s il v er w ire ( 0. 1 mm o . d. , 1 5 cm l eng th ) is tig h tl y
w ra p p ed a ro und th e TEF LON TM s urf a ce co v ering to
f o rm a co il 2 0 o f a bo ut 5 mil l imeters in l eng th . A w ire
w ra p p ing to o l ma y be util ized f o r th is p urp o s e. Th e
tra il ing p o rtio n o f th e w ire to th e rig h t o f co il 2 0 is
co v ered w ith a s ectio n 2 2 o f h ea t s h rink a bl e Te? o n
tubing ( 5 cm l o ng , l . 5 mm o . d. , Z eus s I ndus tria l Pro d
ucts I nc. ) , l ea v ing s ma l l l eng th s o f th e s il v er w ire a nd
p l a tinum iridium w ires unco v ered to s erv e a s el ectrica l
l ea ds . A h ea t g un o p era ting a t 600 C. is emp l o yed f o r
s h rink ing th e Te? o n tubing . A l a yer o f s il v er ch l o ride is
f o rmed o n th e co il 2 0 by p a s s ing current ( 0. 4 mA/ cm2 )
f o r 60 minutes th ro ug h th e w ra p p ed s il v er w ire w h il e it
is cl ip p ed in a s tirred 0. 1 N I - I CL s o l utio n. Th e ex p o s ed
p o rtio ns o f ref erence el ectro de 1 8 a re th en rins ed w ith
de- io nized w a ter f o r 6 h o urs . Th e ref erence el ectro des
p rep a red in th is ma nner s h o w a p o tentia l o f - 64i3
mV ( n= 1 0) v s . Ag / Ag Cl ( 3 M Na Cl ) in 0. 1 5M Na Cl a t
3 7 C.
I n o rder to immo bil ize g l uco s e o x ida s e ( GOx ,
E. C. l . l . 3 . 4) o n th e ex p o s ed p o rtio n o f w ire 1 2 , a n inner,
neg a tiv el y ch a rg ed membra ne is ? rs t a p p l ied to th e
ex p o s ed w ire s ectio n. Th erea f ter, a circumf erentia l l y
ex tending enzyma tic indica ting l a yer 2 1 is f o rmed
w ith in ca v ity 1 6. Tw o dif f erent a p p ro a ch es h a v e been
emp l o yed to a ch iev e th es e ends . .
A. Atta ch ment o f GOx to Bo v ine S erum Al bumin
Co up l ed Cel l ul o s e Aceta te
Th e ex p o s ed p l a tinum iridium s urf a ce w ith in ca v ity
1 6 is deg rea s ed by w a s h ing w ith a ceto ne. I t is th en
rins ed w ith de- io nized w a ter a nd dried in co l d a ir
s trea m bef o re p o l ymer dep o s itio n.
Th e l ef t h a nd p a rt o f th e s ens o r ( p o rtio n to th e l ef t o f
th e ref erence el ectro de co il 2 0) is dip p ed into 5% cel l u
l o s e a ceta te ( 3 9 . 8 % a cetyl co ntent) in 50% a ceto ne a nd
50% eth a no l f o r 1 0 s eco nds a nd is w ith dra w n s l o w l y. I t
is th en ex p o s ed to th e v a p o r a bo v e th e cel l ul o s e a ceta te
s o l utio n f o r 5 s eco nds a nd is dip p ed a g a in into th e cel l u
l o s e a ceta te s o l utio n f o r 1 0 s eco nds . Th e s ens o r is th en
remo v ed a nd dried in a ir a t ro o m temp era ture ( 2 3 C. )
f o r o ne minute a nd p l a ced in deio nized w a ter f o r 6
h o urs to p ermit dis p l a cement by w a ter o f entra p p ed
s o l v ent in th e membra ne p o res . Th e cel l ul o s e a ceta te
membra ne p rep a red in th is f a s h io n s h o w s g o o d l o ng
term s ta bil ity a nd a l s o dis crimina tes w el l a g a ins t a s co r
ba te a nd ura te. Bo v ine s erum a l bumin ( BS A) is th en
co v a l entl y co up l ed to cel l ul o s e a ceta te a nd a s ubs e
q uent rea ctio n o f th e membra ne w ith 60x , w h ich h a s
p rev io us l y been a ctiv a ted w ith a n ex ces s o f p - benzo q ui
no ne, is ca rried o ut. Th e deta il ed p ro cedure f o r th is
rea ctio n is des cribed in th e l itera ture, S ternberg et. a l . ,
Ana l . Ch em. 1 9 8 8 , 60, 2 78 1 , w h ich is inco rp o ra ted
h erein by ref erence.
5
2 5
3 0
3 5
45
50
55.
65
6
B. Ph ys ica l Ads o rp tio n o f Enzyme o n Cel l ul o s e
Aceta te o r Na ? o n F o l l o w ed by Cro s s l ink ing w ith
Gl uta ra l deh yde
l . Th e s ens o r is co a ted w ith cel l ul o s e a ceta te in ex
a ctl y th e s a me ma nner a s des cribed a bo v e to crea te
membra ne. Th e GOx ( 2 70 U/ mg ) is p h ys ica l l y a d
s o rbed by dro p p ing 5 p l o f 60x s o l utio n ( 40 mg / Ml in
0. 1 M p h o s p h a te buf f ered s a l ine) o n th e indica ting el e
ment w ith in ca v ity 1 6, a nd is a l l o w ed to dry f o r 1 0
minutes a t ro o m temp era ture. To immo bil ize th e en
zyme a nd f o rm circumf erentia l s urf a ce 2 1 , th e s ens o r is
ex p o s ed to g l uta ra l deh ydev v a p o r g enera ted f ro m 2 5%
g l uta ra l deh yde s o l utio n p l a ced a t th e bo tto m o f a n en
cl o s ed g l a s s ch a mber f o r 1 2 h o urs a t ro o m temp era ture.
Th e s ens o rv is th en rins ed in de- io nized w a ter a nd dried
in a ir f o r 2 h o urs . Th e cro s s l ink ing w ith g l uta ra l deh yde
p ro tects th e enzyme f ro m h ea t deg ra da tio n, p ro teo l ytic
enzymes a nd h ydro l ys is , E. M. S a l o na , C. S a ro nio , a nd
S . Ga ra ttini ( eds ) , I ns o l ubil ized Enzymes . Ra v en,
New Yo rk , 1 9 74, inco rp o ra ted by ref erence h erein.
2 . Na ? o n ( Per? uo ro s ul f o nic a cid p o l ymer, o bta ined
f ro m E. I . D uPo nt de Nemo urs a nd Co . , ma y a l s o be
us ed a s a n a l terna te f o r cel l ul o s e a ceta te f o r th e inner
membra ne. Af ter cl ea ning th e s ens ing p o rtio n o f th e
s ens o r a s a bo v e, it is el ectro co a ted w ith Na ? o n us ing
th e meth o d des cribed by Ada ms et a 1 , Neuro s ci. Meth .
Vo l . 2 2 , 1 9 8 7, p p l 67- l 72 , inco rp o ra ted by ref erence
h erein. One dro p o f Na ? o n ( 5% s o l utio n, Al drich ) is
p l a ced in a 2 mm l o o p f o rmed a t o ne end o f a co p p er
w ire. A D C p o tentia l o f + 3 V is a p p l ied to th e w o rk ing
el ectro de w ith res p ect to th e l o o p f o r 1 0 s eco nds . Th e
s ens o r is p ul l ed o ut o f th e l o o p bef o re turning o f f th e
p o tentia l a nd is dried in a ir f o r 2 h o urs , a nd th e 60x
enzyme is a p p l ied a s des cribed a bo v e.
Al terna te p o l ymers ma y be us ed in l ieu o f o r in co m
bina tio n w ith cel l ul o s e a ceta te o r Na ? o n f o r co a ting o f
th e ex p o s ed Pt- l r w ire s urf a ce. F o r ex a mp l e, p o l ya ni
l ine a nd p o l yp h eno l deriv a tiv es ca n be el ectro ch emi
ca l l y dep o s ited o nto th e ex p o s ed indica ting el ectro de
s urf a ce. Ox ida tiv e el ectro p o l ymeriza tio n o f a nil ine a nd
p h eno l mo no mer yiel ds s ta bl e a nd a dh es iv e co a ting
o v er th e ex p o s ed w ire. Th es e ma teria l s mo reo v er h a v e
g o o d s ize s el ectiv ity w h ich ca n be util ized to f urth er
imp ro v e th e s ens o r s el ectiv ity a g a ins t el ectro ch emica l
interf erences in bio l o g ica l env iro nments . Th e co mbina
tio n o f a s ize s el ectiv e co a ting w ith a ch a rg e s el ectiv e
? l m ( e. g . cel l ul o s e a ceta te) ma y reduce th e in v iv o ba ck
g ro und current a nd th e ris k o f el ectro ch emica l interf er
ence. El ectro p o l ymeriza tio n o f a nil ine a nd p h eno l is
w el l k no w n, s ee f o r ex a mp l e Oh s a k a et a 1 . Ana l . Ch em.
1 9 8 7, 59 , l 758 - 6l , a nd Ma l ites ta et a l . Ana l . Ch em. 1 9 9 0,
62 , 2 73 5- 40, bo th o f w h ich a re inco rp o ra ted by ref er
ence h erein.
F ina l l y, Ea s tma n- K o da k AQ 2 9 - D p o l ymer ( p o l y( es
ter- s ul f o nic a cid) ) h a s bo th ch a rg e a nd s ize s el ectiv e
f ea tures , a nd ma y be a p p l ied to th e ex p o s ed indica ting
el ectro de w ire in l ieu o f Na ? o n. A co a ting o f th is typ e
a p p l ied to th e indica ting el ectro de w ith a cel l ul o s e a ce
ta te l a yer th ereo v er s h o ul d imp ro v e o v era l l s el ectiv ity.
Co mbined co a ting s ma de f ro m mix tures o f cel l ul o s e
a ceta te a nd th e AQ 2 9 - D p o l ymer s h o ul d a l s o p ro v ide
a dv a nta g es in terms o f s ens o r s el ectiv ity.
I n o rder to co mp l ete th e p rep a ra tio n o f th e s ens o r,
th e w h o l e a s s embl y, incl uding th e ref erence el ectro de,
is cl ip co a ted w ith 4% p o l yureth a ne ( Th ermedics , S G
8 5A) dis s o l v ed in 9 8 % tetra h ydro f ura n ( T H F ) a nd 2 %
dimeth yl f o rma mide ( D MF ) to f o rm a n o uter membra ne
5, 1 65, 407
7
2 4. Th e p o l yureth a ne s o l utio n ( 1 0 uL) is h el d in a w ire
l o o p ( 2 mm id. ) by s urf a ce tens io n a nd th e s ens o r is
p a s s ed th ro ug h th e l o o p . Th is l ea v es a unif o rm p o l ymer
? l m o n th e co mp l eted s ens o r 2 5 to th e a p p ro p ria te ex
tent dep icted in F I G. 1 . Th is meth o d p ro v ides a g o o d
co ntro l o v er th e a mo unt o f p o l ymer w h ich is a p p l ied to
th e s ens o r. Th e s ens o r is dried in a ir f o r 6 h o urs a t ro o m
temp era ture a nd th en l ef t in 0. 1 M p h o s p h a te buf f ered
s a l ine, p H =7. 4 f o r 72 h o urs f o r th e v a rio us o uter mem
bra nes to co nditio n f ul l y. I t is p o s s ibl e to reco a t th e
s ens o r w ith p o l yureth a ne if th e des ired l inea r ra ng e o f
g l uco s e s ens itiv ity is no t o bta ined a f ter th e ? rs t co a ting .
EXAMPLE 2 F I G. 2
One end o f a 1 0 cm s ectio n 2 6 o f Te? o n- co a ted p l a ti
h um- iridium w ire is p ro v ided h a v ing a 0. 1 8 mm o . d. , a
centra l Pt- I r w ire 2 8 a nd a te? o n s h ea th 3 0 ( 0. 03 5 mm
th ick nes s ) . Th e l ef t h a nd end o f th e w ire is s trip p ed to
f o rm a ca v ity 3 2 a s des cribed in Ex a mp l e 1 . Th e rig h t
h a nd end o f s ectio n 2 6 is th en ins erted into a 5 centime
ters l o ng p o l yeth yl ene tube 3 4 ( 0. 67 mm o . d. , 0. 3 0 mm
id) . Th e l ef t h a nd ex tremity o f th e p o l yeth yl ene tube is
s ea l ed by p utting a dro p o f 4% cel l ul o s e a ceta te s o l utio n
( in a ceto ne) into th e o p ening . Th e a ceto ne is a l l o w ed to
dry w h il e h o l ding th e Te? o n- co a ted w ire in th e middl e
o f th e p o l yeth yl ene tube. Th is p ermits th e f o rma tio n o f
a circumf erentia l s a l t bridg e dep o s it 3 6 w h ich ef f ec
tiv el y a cts a s th e termina l p a rt o f th e ref erence el ec
tro de, l ies in a p l a ne tra ns v ers e to th e l o ng itudina l a x is
o f th e w ire 2 8 a nd es ta bl is h es el ectrica l co nta ct betw een
th e ref erence a nd s ens ing el ectro des . Th e emp ty a nnu
l a r s p a ce betw een th e Te? o n- co a ted w ire a nd th e p o l y
eth yl ene tube is th en ? l l ed under v a cuum w ith 0. 1 M
p h o s p h a te buf f er, p H =7. 4 co nta ining 9 g / L Na Cl . A
ch l o ridized s il v er w ire 3 8 ( 0. 05 mm o . d. , 5 cm l o ng
p rep a red a s des cribed in Ex a mp l e 1 ) , is intro duced into
th e p o l yeth yl ene tube f ro m. th e rig h t h a nd end th ereo f
a nd th is o p ening is a l s o s ea l ed a s des cribed a bo v e to
p res ent a s ea l ing dep o s it 40. Th e ref erence el ectro de
s h o w s a p o tentia l o f ~ 60: 1 0 mV ( n=6) v s , Ag / Ag Cl
( s a tura ted K CL) a t 3 7 C. Th e enzyme immo bil iza tio n
a nd p o l yureth a ne dep o s itio n s tep s a re th en ca rried o ut
us ing th e p ro cedures des cribed in Ex a mp l e 1 to g iv e th e
inner, neg a tiv el y ch a rg ed membra ne 3 2 0, th e circum
f erentia l indica ting enzyme l a yer 3 3 , a nd o uter p ermea
bl e membra ne 42 il l us tra ted in F I G. 2 . Th e co mp l ete
s ens o r 43 is th en rea dy f o r ca l ibra tio n a nd us e w ith
el ectrica l co nnectio ns a f f o rded by th e a x ia l l y ex tending
ends o f th e w ires 2 8 , 3 8 .
Th e s ens o rs des cribed in th e a bo v e ex a mp l e a re ca l i
bra ted by dip p ing into a th errno s ta ted cel l ( a t 3 7 C. )
co nta ining 1 0 ml o f s tirred 0. 1 M p h o s p h a te buf f ered
s a l ine, p H =7. 4, a nd a p o tentia l o f +600 mV ( f o r h ydro
g en p ero x ide detectio n) is a p p l ied betw een th e w o rk ing
a nd th e ref erence/ indica ting el ectro des . Th e ba ck
g ro und current is a l l o w ed to s ta bil ized f o r 2 0 minutes .
Th e ca l ibra tio n o f th e s ens o r is ca rried o ut by a dding
increa s ing a mo unts o f g l uco s e to th e s tirred buf f er. Th e
current is mea s ured a t th e p l a tea u ( s tea dy s ta te re
s p o ns e) a nd is rel a ted to th e co ncentra tio n o f th e a na
l yte. F o l l o w ing th e ca l ibra tio n p ro cedure, th e s ens o rs
a re s to red in 0. 1 M p h o s p h a te buf f ered s a l ine, p H = 7. 4 a t
ro o m temp era ture.
A typ ica l res p o ns e curv e to th e g l uco s e a dditio n is
s h o w n in F I G. 3 , f o r a s ens o r ma de in a cco rda nce w ith
F I G. 1 . As il l us tra ted, th e res p o ns e ch a ra cteris tics o f
th e s ens o r o v er th e co ncentra tio n ra ng e o f interes t
( 0- 2 5 mM) a re es s entia l l y l inea r, a nd a re es p ecia l l y s o
1 0
2 5
3 0
3 5
40
45
55
60
65
8
o v er th e ra ng e o f 0 l 5 mM. Th e s ens o r o utp ut is a l s o
es s entia l l y indep endent o f th e s tirring ra te. Th e in v itro
ch a ra cteris tics o f th e s ens o r a re s umma rized in th e f o l
l o w ing Ta bl e. A typ ica l s to ra g e s ta bil ity curv e f o r th e
s ens o r is s h o w n in F I G. 4. D uring th e ? rs t f ew da ys o f
s ens o r p rep a ra tio n, th e p o l yureth a ne membra ne
ch a ng es its p ermea bil ity f o r g l uco s e a s a res ul t o f h ydro
l ytic a nd s w el l ing p ro ces s es , l ea ding to th e increa s ed
p a s s a g e o f g l uco s e a nd a n increa s ed current. Af ter th is
initia l p erio d, h o w ev er, th e s ta bil ity is ex cel l ent.
Th e s ens o rs o f th e inv entio n a re in us e el ectrica l l y
co up l ed w ith s uita bl e s ig na l p ro ces s ing eq uip ment, a nd
imp l a nted into a des ired s ubcuta neo us s ite. Gl uco s e a nd
o x yg en dif f us ing th ro ug h th e o uter s ynth etic p o l ymer
membra ne a re enzyma tica l l y ca ta l yzed by th e G0, . a t
th e indica ting s urf a ce, res ul ting in p ro ductio n o f g l u
co nic a cid a nd h ydro g en p ero x ide. Th e l a tter is mea
s ured a mp ero metrica l l y, w h ich is a mea s urement o f
g l uco s e co ncentra tio n.
TABLE
I n Vitro Ch a ra cteris tics o f F I G. 1 Gl uco s e S ens o r
Pa ra meter Va l ue
Res idua l current ( nA/ mmz) 7 0. 7 i 0. 2
S ens itiv ity ( nA/ mM - mmz) 1 . 2 i 0. 3
Linea r Ra ng e ( up p er l imit) ( mM) 1 5 i 3
Res p o ns e time ( min. ) , T 9 0% 3 . 5 i 1
Res ul ts s h o w n a bo v e a re ex p res s ed a s mea n i S D f o r s ix s ens o rs .
" Res idua l currents a re mea s ured a f ter 1 h o ur o f p o l a riza tio n.
F I GS . 5 a nd 6 il l us tra te a no th er s ens o r 44 in a cco r
da nce w ith th e inv entio n. I n th is ca s e, th e s ens o r bo dy
46 is receiv ed w ith in a s ta inl es s s teel h o l l o w tubul a r
needl e 48 .
Th e s ens o r bo dy 46 incl udes a n innermo s t, Te? o n
co a ted, p l a tinum- iridium w ire 50 ( 9 0% Pt/ 1 0% I r) h a v
ing a to ta l CD . o f a bo ut 0. 2 mm a nd a ca v ity 52 f o rmed
th erein a s des cribed in Ex a mp l e 1 . Th e ca v ity 52 is
a p p ro x ima tel y 1 . 0 mm in l eng th a nd is l o ca ted a bo ut 3 . 0
mm f ro m th e tip o f th e w ire 50. A g l uco s e o x ida s e l a yer
54 is immo bil ized w ith in th e ca v ity 52 , a nd co mp ris es a
cel l ul o s e a ceta te p o l ymer l a yer a tta ch ed to th e s urf a ce
o f th e Pt- I r w ire, w ith g l uco s e o x ida s e cro s s l ink ed
th ro ug h g l uta ra l deh yde o nto th e cel l ul o s e a ceta te. Th is
p ro cedure is in a cco rda nce w ith Ex a mp l e l . B. l . a bo v e.
Th e entirety o f th e indica ting el ectro de is th en co v ered
by a membra ne 56 o f p o l yureth a ne, a g a in us ing th e
meth o d s et f o rth in Ex a mp l e 1 .
Th e s ens o r bo dy 46 is th ereup o n ins erted into a 2 5
g a ug e dis p o s a bl e s ta inl es s s teel h yp o dermic needl e, th e
l a tter h a v ing a n a p erture 58 a dj a cent th e f o rw a rd,
s h a rp ened ins ertio n end 60 th ereo f . Th e s ens o r bo dy 46
is ins ta l l ed in s uch ma nner th a t th e g l uco s e o x ida s e l a yer
54 co mes into reg is try w ith th e s idew a l l o p ening 58 ,
th ereby ex p o s ing th e l a yer 54 to th e bio l o g ica l env iro n
ment. A s il ico ne rubber p l ug 62 is ins ta l l ed in th e f o r
w a rd end o f th e needl e 48 a s s h o w n.
As il l us tra ted in F I G. 5, th e w ire 50 ex tends rea r
w a rdl y o ut o f th e end o f needl e 48 , a nd is a da p ted to be
co nnected w ith a p p ro p ria te ins trumenta tio n f o r mea
s uring g l uco s e co ncentra tio ns . I n o rder to s ea l th e rea r
w a rd end o f th e s ens o r 44, a bea d 64 o f ep o x y is a p p l ied
a ro und th e w ire 50 a nd th e butt end o f th e needl e 48 a nd
s ens o r bo dy 46.
Th e o v era l l s ens o r 44 is co mp l eted by p ro v is io n o f a
h o l der 66 ex tending tra ns v ers el y o f th e needl e 48 . Th e
h o l der 66 is p ref era bl y in th e f o rm o f a p l a s tic s h eet
w ra p p ed a ro und th e rea rw a rd end o f th e needl e 48 a s
s h o w n, a nd s ecured by mea ns o f ep o x y ' o r p o l ycya no a
5, 1 65, 407
9
cryl a te g l ue. Th e h o l der 66 p ermits rea dy ma nip ul a tio n
a nd ins ertio n o f th e s ens o r 44 ev en by th e p a tient.
I n th e us e o f s ens o r 44, th e ref erence el ectro de ma y
be eith er ex terna l l y a p p l ied o r imp l a nted. As a n ex terna l
el ectro de, us e ma y be ma de o f a co mmercia l el ectro ca r
dio g ra m s k in el ectro de des cribed p rev io us l y ma y be
us ed. An ex terna l ref erence el ectro de s h o ul d be a p p l ied
in cl o s e p ro x imity to th e imp l a nted s ens o r f o r th e bes t
mea s urement res ul ts . Th e h o l der 66 ma y a l s o be us ed to
s up p o rt a n ex terna l el ectro de o f th e typ e des cribed
p rev io us l y. I na s much a s th e h o l der l ies cl o s el y a dj a cent
th e s k in up o n imp l a nta tio n, th e h o l der ma y s erv e a s a n
idea l p l a tf o rm f o r th e ex terna l el ectro de.
F I G. 7 il l us tra tes a n embo diment w h erein us e is ma de
o f a n imp l a nta bl e ref erence el ectro de. I n th is ca s e, th e
needl e 48 h a s a n el ectro dep o s ited l a yer 68 o f s il v er o n
th e ex terna l s urf a ce th ereo f , w ith th is l a yer being a no d
ized in th e p res ence o f ch l o ride io n to crea te a Ag
/ Ag Cl ref erence el ectro de. A s il v er l ea d w ire 70 is
co nductiv el y a f ? x ed to th e rea rw a rd end o f needl e 48
by mea ns o f s il v er ep o x y o r s imil a r ex p edient, a nd th e
h o l der 66 is w ra p p ed a bo ut th is co nnectio n a s s h o w n.
Al terna tel y, th e inner w a l l o f th e s ta inl es s s teel needl e
48 ma y be p ro v ided w ith a n el ectro dep o s ited, a no dized
s il v er l a yer, w ith co nducting g el betw een th is l a yer a nd
th e s ens o r bo dy 46. A s il v er l ea d w ire w o ul d th en be
co nductiv el y s ecured to th e inner needl e s urf a ce. I n th is
embo diment, el ectrica l current f l o w s th ro ug h th e g el
betw een th e indica ting el ectro de a nd th e ref erence el ec
tro de. .
S ens o rs co ns tructed in a cco rda nce w ith F I GS . 5- 7,
a nd us ing eith er ex terna l o r imp l a nted ref erence el ec
tro des , g iv e es s entia l l y th e s a me l inea r res p o ns e a s th o s e
co ns tructed in a cco rda nce w ith F I GS . 1 - 2 .
Actua l ex p erience w ith s ens o rs in a cco rda nce w ith
th e inv entio n h a s demo ns tra ted th a t, up o n imp l a nta tio n,
th e cel l s a nd ca p il l a ries o f p ro x ima l tis s ue a re s l ig h tl y
da ma g ed. Af ter f o ur o r ? v e da ys , h o w ev er, s uch tis s ues
reg enera te a ro und th e s ens o r, f o rming a co l l a g en l a yer.
Neo v a s cul a riza tio n h a s a l s o been o bs erv ed in th e co l l a
g en l a yer, a nd th is p h eno meno n ma y p a rtia l l y a cco unt
f o r th e s ens itiv ity o f th e s ens o r. Th is is indica tiv e o f
o p era tio n o f th e p a tient s immune s ys tem. I n a ny ev ent,
th e p res ence o f a neo v a s cul a rized co l l a g en l a yer a dj a
cent th e imp l a nted s ens o r p ermits p a s s a g e o f o x yg en
a nd g l uco s e. I n a dditio n, it h a s been f o und th a t in th e
? rs t h o urs a f ter imp l a nta tio n, th e s ens o r res p o ns e is
s o mew h a t v a ria bl e. Ov er time, h o w ev er, th is v a ria bil ity
is decrea s ed a nd th e p erf o rma nce o f th e imp l a nted s en
s o r increa s es . Th is is bel iev ed to be due to th e s ta bil iza
tio n o f th e tis s ue a ro und th e imp l a nted s ens o r. Th e end
res ul t is th a t' th e s ens o rs o f th e p res ent inv entio n ma y be
s ucces s f ul l y imp l a nted a nd l ef t in p l a ce f o r p erio ds o f
time h ereto f o re th o ug h t imp ra ctica l , e. g . , p erio ds o f
f ro m s ev en da ys to th ree w eek s a re f ea s ibl e.
Th o s e s k il l ed in th e a rt w il l unders ta nd th a t th e s en
s o rs o f th e inv entio n ma y req uire in v iv o ca l ibra tio n.
Th is w o ul d typ ica l l y be do ne by mea s uring tw o bl o o d
2 0
45
55
60
65
1 0
g l uco s e l ev el s by co nv entio na l mea ns , a nd co rrel a ting
th es e k no w n v a l ues w ith th e o utp ut o f th e s ens o r.
I t w il l th us be s een th a t th e enzyma tic s ens o rs in
a cco rda nce w ith th e inv entio n ex h ibit p ro p erties h ere
to f o re dif ? cul t to a ch iev e, incl uding s ma l l , f ul l y im
p l a nta bl e s ize; l inea rity in res p o ns e o v er th e co ncentra
tio n ra ng es o f interes t; s to ra g e s ta bil ity; a nd th e a bil ity
to be co ns is tentl y ma nuf a ctured w ith o ut undue rej ec
tio n ra tes . .
W e cl a im: I
1 . A s ens o r a da p ted f o r p o s itio ning in a n env iro nment
ch a ra cterized by th e p res ence o f bio l o g ica l mo l ecul es
w h ich a re s ubs tra tes f o r o r p ro ducts p ro duced by en
zymes in o rder to determine th e p res ence o f s a id mo l e
cul es , s a id s ens o r co mp ris ing :
a n el o ng a ted ? ex ibl e bo dy co mp ris ing a l eng th o f
el ectrica l l y co nductiv e indica ting el ectro de w ire
co v ered w ith a n el ectrica l l y ins ul a tiv e ma teria l ,
th ere being a p o rtio n o f s a id ma teria l remo v ed
f ro m s a id el ectro de w ire to de? ne a n enzyme
receiv ing zo ne;
a n enzyme o p era bl y immo bil ized o n s a id zo ne to
p res ent a n enzyma tic indica ting s urf a ce;
a ref erence el ectro de s up p o rted o n s a id bo dy a nd
p res enting a s ectio n a da p ted f o r ex p o s ure to s a id
env iro nment,
s a id indica ting s urf a ce a nd s a id ref erence el ectro de
s ectio n being l a tera l l y s p a ced a p a rt a l o ng th e
l eng th o f s a id bo dy a nd ea ch s ubs ta ntia l l y circum
s cribing th e bo dy, _
s ubs ta ntia l l y th e entireties o f s a id circums cribing
indica ting s urf a ce a nd s a id circums cribing ref er
ence el ectro de s ectio n being ex p o s ed f o r rea ctio n
w ith s a id env iro nment w h en th e s ens o r is p l a ced
th erein,
s a id indica ting el ectro de w ire s erv ing to s up p o rt s a id
s ens o r w ith in s a id env iro nment w ith o ut th e need
f o r a ca rrier.
2 . Th e s ens o r o f cl a im 1 , s a id indica ting s urf a ce being
l o ca ted intermedia te th e ends o f s a id l eng th o f el ectro de
w ire w ith res p ectiv e s eg ments o f s a id ins ul a ting ma te
ria l being ' o n o p p o s ite s ides o f a nd de? ning s a id enzyme
receiv ing zo ne.
3 . Th e s ens o r o f cl a im 1 , s a id ref erence el ectro de
co mp ris ing a co il dis p o s ed a bo ut s a id bo dy.
4. Th e s ens o r o f cl a im 1 , ' incl uding a n o uter s ynth etic
p o l ymer membra ne dis p o s ed o v er s a id indica ting s ur
f a ce a nd ref erence el ectro de s ectio n, s a id membra ne
being p ermea bl e to s a id bio l o g ica l mo l ecul es .
5. Th e s ens o r o f cl a im 4, s a id membra ne being f o rmed
o f p o l yureth a ne.
6. Th e s ens o r o f cl a im 4, s a id membra ne h a v ing a
th ick nes s o f f ro m a bo ut 5 to 1 0 micro ns .
7. Th e s ens o r o f cl a im 1 , incl uding a n inner mem
bra ne a p p l ied to s a id el ectro de w ire a l o ng th e l eng th o f
s a id enzyme- receiv ing zo ne.
8 . Th e s ens o r o f cl a im 7, s a id membra ne being neg a
tiv el y ch a rg ed.
9 . Th e s ens o r o f cl a im 1 , s a id s ens o r being a g l uco s e
s ens o r, s a id enzyme being g l uco s e o x ida s e.
# i i #
UNI TED S TATES PATENT AND TRAD EMARK OF F I CE
CERTI F I CATE OF CORRECTI ON
PATENTNO. 2 5, 1 65, 407
D ATED : No v ember 2 4, 1 9 9 2
' NVENTOR( S ) I GEORGE 3 . W I LS ON, D I LBI R s . BI ND RA, BRI AN s . H I LL, D ANI EL R.
TH EVENOT, ROBERT S TERNB ERG , GERARD REACH & YANAN Z H ANG
I t is certif ied th a t erro r a p p ea rs in th e a bo v e- indentif ied p a tent a nd th a t s a id Letters Pa tent is h ereby
co rrected a s s h o w n bel o w :
Co l umn 1 , l ine 6 bef o re th e " Ba ck ro und f '
h H -
f o l l o w ing p a ra g rg p h : g o t e I nv entio n , ins ert th e
- Th is inv entio n w a s ma de w ith Go v ernment s up p o rt under Gra nt
No . ROl D K 3 071 8 a w a rded by th e Na tio na l I ns titutes o f H ea l th .
Th e Go v ernment h a s certa in rig h ts in th e inv entio n.
S ig ned a nd S ea l ed th is
F o urth D a y o f J a nua ry, 1 9 9 4
Ang s t" 6M W
BRUCE LEH MAN
Arres ting O? icer Co mmis s io ner o f Pa rents a nd Tra dema rk s