U.S. patent application number 10/706208 was filed with the patent office on 2005-02-17 for methods and kits for assays of rapid screening of diabetes.
Invention is credited to Carney, Fiona Patricia, Lane, Jennifer Dawn, Morris, Carol Ann.
Application Number | 20050038329 10/706208 |
Document ID | / |
Family ID | 32326585 |
Filed Date | 2005-02-17 |
United States Patent
Application |
20050038329 |
Kind Code |
A1 |
Morris, Carol Ann ; et
al. |
February 17, 2005 |
Methods and kits for assays of rapid screening of diabetes
Abstract
The invention provides an in vivo screening assay and an in
vitro screening assay for rapid screening of diabetes. A method of
the invention includes determining a first glucose concentration in
an ocular fluid of a patient; administering orally a load of
carbohydrate to the patient; determining a second glucose
concentration in an ocular fluid of the patient at a period of time
of less than 50 minutes after orally administering of the load of
carbohydrate; comparing the second glucose concentration with the
first glucose concentration to determine if the patient is likely
to be a diabetic. The method of the invention is performed by using
a kit of the invention. The kit comprises: (1) a glucose-sensing
ophthalmic device and instructions for using the glucose-sensing
ophthalmic device to screen for diabetes; or (2) two or more
tear-collecting devices, and a testing agent composition which
specifically reacts with glucose to form a detectable signal. The
glucose-sensing ophthalmic device comprises a testing agent
composition which specifically and reversibly interacts with
glucose to form a detectable optical signal which changes in a
concentration-dependent manner.
Inventors: |
Morris, Carol Ann; (Duluth,
GA) ; Carney, Fiona Patricia; (Atlanta, GA) ;
Lane, Jennifer Dawn; (Stone Mountain, GA) |
Correspondence
Address: |
CIBA VISION CORPORATION
PATENT DEPARTMENT
11460 JOHNS CREEK PARKWAY
DULUTH
GA
30097-1556
US
|
Family ID: |
32326585 |
Appl. No.: |
10/706208 |
Filed: |
November 12, 2003 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
60427705 |
Nov 20, 2002 |
|
|
|
Current U.S.
Class: |
600/319 ; 435/14;
977/932 |
Current CPC
Class: |
G01N 2800/042 20130101;
G01N 33/66 20130101; G01N 33/6893 20130101 |
Class at
Publication: |
600/319 ;
435/014 |
International
Class: |
A61B 005/00; C12Q
001/54 |
Claims
What is claimed is:
1. A method for rapidly screening for diabetes, comprising the
steps of: contacting a glucose-sensing ophthalmic device with an
ocular fluid, wherein the glucose-sensing ophthalmic device
comprises a testing agent composition which specifically and
reversibly interacts with glucose to form a detectable signal which
changes in a concentration-dependent manner; determining by means
of the glucose-sensing ophthalmic device a first glucose
concentration in the ocular fluid; administering orally a load of
carbohydrate to the patient; at a period of time of less than 50
minutes after orally administering of the load of carbohydrate,
determining by means of the glucose-sensing ophthalmic device a
second glucose concentration in the ocular fluid; and comparing the
second glucose concentration with the first glucose concentration
to determine if the patient is likely to be a diabetic.
2. A method of claim 1, wherein the second glucose concentration is
determined about 15 minutes after orally administering of the load
of carbohydrate.
3. A method of claim 1, wherein said testing agent composition
comprises a receptor that is capable of reversibly binding glucose
and has a detectable optical signal that changes in a
concentration-dependent manner when the receptor is reversibly
bound to glucose, wherein said detectable optical signal results
from one or more labels associated with the receptor.
4. A method of claim 3, wherein the detectable optical signal
results from a pair of labels associated with the receptor, a first
label and a second label, wherein one of the first and second label
is a fluorescence energy donor and the other is a fluorescence
energy acceptor or a non-fluorescence energy acceptor.
5. A method of claim 1, wherein said testing agent composition
comprises a receptor having a first label associated therewith and
a competitor having a second label associated therewith, wherein
one of the first and second labels is a fluorescent energy donor
and the other one is a fluorescent or non-fluorescent energy
acceptor.
6. A method of claim 1, wherein said load of carbohydrate is at
least 40 grams of carbohydrate.
7. A method for rapidly screening for diabetes, comprising the
steps of: collecting a first tear fluid from a patient using a
first tear-collecting device; assaying a specific amount of the
first tear fluid to determine a first glucose concentration;
administering orally a load of carbohydrate to the patient;
collecting a second tear fluid, at a period of time of less than 50
minutes after orally administering of the load of carbohydrate,
using a second tear-collecting device; assaying a specific amount
of the second tear fluid to determine a second glucose
concentration; and comparing the second glucose concentration with
the first glucose concentration to determine if the patient is
likely to be a diabetic.
8. A method of claim 7, wherein said first and second tear
collecting devices are selected from the group consisting of
capillary tubes, hydrogel strips, and contact lenses.
9. A method of claim 7, wherein at least one of the first and
second tear collecting devices is a strip having a first end and a
second end, wherein said strip is made of a hydrogel material in
substantially dry state and is characterized by having a
substantially uniform swelling along the hydrogel strip from the
first end to the second end when fully wicked by a tear fluid and
by having a correlation between the volume of tear uptake by said
strip and the length of a tear-wicked end portion of said
strip.
10. A method of claim 7, wherein said load of carbohydrate is at
least 40 grams of carbohydrate.
11. A method of claim 7, wherein said second tear fluid is
collected at a period of time of at least 15 minutes after orally
administering of the load of carbohydrate.
12. A kit for rapid screening of diabetes, the kit comprising: a
glucose-sensing ophthalmic device and instructions for using the
glucose-sensing ophthalmic device to screen for diabetes, wherein
the glucose-sensing ophthalmic device comprises a testing agent
composition which specifically and reversibly interacts with
glucose to form a detectable optical signal which changes in a
concentration-dependent manner.
13. A kit of claim 12, wherein said testing agent composition
comprises a receptor that is capable of reversibly binding glucose
and has a detectable optical signal that changes in a
concentration-dependent manner when the receptor is reversibly
bound to glucose, wherein said detectable optical signal results
from one or more labels associated with the receptor.
14. A kit of claim 13, wherein the detectable optical signal
results from a pair of labels associated with the receptor, a first
label and a second label, wherein one of the first and second label
is a fluorescence energy donor and the other is a fluorescence
energy acceptor or a non-fluorescence energy acceptor.
15. A kit of claim 14, wherein said receptor is selected from the
group consisting of GGBP, concanavalin A, inactivated glucose
oxidase, inactivated glucose dehydrogenase, and boronic acid.
16. A kit of claim 14, wherein said fluorescent energy donor is
selected from the group consisting of xanthene-type dyes,
fluorescein-type dyes, rhodamine-type dyes, cyanine-type dyes,
phycobiliproteins.
17. A kit of claim 12, wherein said testing agent composition
comprises a receptor having a first label associated therewith and
a competitor having a second label associated therewith, wherein
one of the first and second labels is a fluorescent energy donor
and the other one is a fluorescent or non-fluorescent energy
acceptor.
18. A kit of claim 12, wherein the ophthalmic device can comprise a
glucose-sensing LbL coating which is not covalently attached to the
core material of the ophthalmic device, wherein the glucose-sensing
LbL coating comprises the testing agent composition.
19. A kit of claim 18, wherein the glucose-sensing LbL coating
comprises one or more layers of a vesicle with a charged surface
and with a receptor or a competitor entrapped therein, wherein the
receptor has a first label associated therewith and the competitor
has a second label associated therewith, wherein one of the first
and second labels is a fluorescent energy donor and the other one
is a fluorescent or non-fluorescent energy acceptor.
20. A kit of claim 18, wherein the glucose-sensing LbL coating
comprises one or more layers of a vesicle with a charged surface
and with a receptor entrapped therein, wherein the receptor is
capable of reversibly binding glucose and has a detectable optical
signal that changes in a concentration-dependent manner when the
receptor is reversibly bound to glucose, wherein said detectable
optical signal results from one or more labels associated with the
receptor.
21. A kit of claim 20, wherein the detectable optical signal
results from a pair of labels associated with the receptor, a first
label and a second label, wherein one of the first and second label
is a fluorescence energy donor and the other is a fluorescence
energy acceptor or a non-fluorescence energy acceptor.
22. A kit for rapid screening of diabetes, the kit comprising: two
or more tear-collecting devices, and a testing agent composition
which specifically reacts with glucose to form a detectable
signal.
23. A kit of claim 22, wherein said two or more tear-collecting
devices are selected from the group consisting of a hydrogel strip,
a capillary tube, and a soft-hydrogel contact lens.
24. A kit of claim 23, wherein said two or more tear-collecting
devices are hydrogel strips, wherein each of said strips has a
first end and an opposite second end, wherein each of said strips
is made of a hydrogel material in substantially dry state and is
characterized by having a substantially uniform swelling along that
hydrogel strip from the first end to the second end when fully
wicked by a tear fluid and by having a correlation between the
volume of tear uptake by that strip and the length of a tear-wicked
end portion of that strip.
25. A kit of claim 24, wherein said hydrogel material is selected
from the group consisting of poly(vinyl alcohol), modified
polyvinylalcohol, poly(hydroxyethyl methacrylate), poly(vinyl
pyrrolidone), poly(vinyl alcohol) with polycarboxylic acids,
polyethylene glycol, polyacrylamide, polymethacrylamide,
silicone-containing hydrogels, polyurethane, polyurea, and mixtures
thereof.
26. A kit of claim 24, wherein said defined correlation between the
volume of tear uptake and the length of the tear-wicked end portion
is a substantially linear relationship.
27. A kit of claim 24, wherein each of said strips has noticeable
marks thereon, wherein each of the marks indicates a volume of the
tear fluid absorbed by the end portion up to that mark of that
strip.
28. A kit of claim 22, wherein said testing agent composition
comprises a receptor that is capable of reversibly binding glucose
and has a detectable optical signal that changes in a
concentration-dependent manner when the receptor is reversibly
bound to glucose, wherein said detectable optical signal results
from one or more labels associated with the receptor.
29. A kit of claim 28, wherein said testing agent composition
comprises a receptor that is capable of reversibly binding glucose
and has a detectable optical signal that changes in a
concentration-dependent manner when the receptor is reversibly
bound to glucose, wherein said detectable optical signal results
from a pair of labels, a first label and a second label, associated
with the receptor, wherein one of the first and second label is a
fluorescence energy donor and the other is a fluorescence energy
acceptor or a non-fluorescence energy acceptor.
30. A kit of claim 22, wherein said testing agent composition
comprises a receptor having a first label associated therewith and
a competitor having a second label associated therewith, wherein
one of the first and second labels is a fluorescent energy donor
and the other one is a fluorescent or non-fluorescent energy
acceptor.
Description
[0001] This application claims under 35 USC .sctn. 119 (e) the
benefit of the filing date of U.S. Provisional Patent Application
Ser. No. 60/427,705 filed Nov. 20, 2002 and all references
incorporated therein.
[0002] The invention is related to methods and kits for rapid
screening of diabetes.
BACKGROUND OF THE INVENTION
[0003] Diabetes is a serious, lifelong disease which can cause
long-term complications that affect almost every part of the body.
This disease often leads to blindness, heart and blood vessel
disease, strokes, kidney failure, amputations, and nerve damage.
Uncontrolled diabetes can complicate pregnancy, and birth defects
are more common in babies born to women with diabetes. Diabetes is
widely recognized as one of the leading causes of death and
disability in the United States.
[0004] It is believed that earlier diagnosis and treatment can
prevent or delay the costly and burdensome complications of
diabetes. Currently, about 5 to 6 million adults in the United
States have diabetes but do not know it. The simpler testing method
of measuring fasting glucose should help identify these people so
they can benefit from treatment sooner. Traditionally, the criteria
for diagnosing diabetes relied heavily on performing an oral
glucose tolerance test (OGTT). In this test, the person must come
in fasting, drink a glucose syrup, and have a blood sample taken 2
hours later. This complicated procedure made detection and
diagnosis of diabetes a difficult and cumbersome process. Recently,
it is recommended that OGTT be eliminated from clinical use and
that fasting plasma glucose is used for detection and diagnosis of
diabetes. A fasting blood glucose of 126 mg/dL or greater is the
value to diagnose diabetes since this value has been found to be
associated with an increased risk of diabetes complications
affecting the eyes, nerves, and kidneys. For such test, a patient
still needs to come in fasting and is forced to draw blood and to
endure discomfort associated with needles to obtain blood samples
for testing fasting blood glucose level. Blood samples are
generally to be sent to a specialized laboratory for testing and it
typically take a couple of days to obtain the testing results. If
the testing results show a fasting blood glucose of 126 mg/dL or
greater, that patient needs to undergo a second test to confirm the
diagnosis. Although the fasting value can be easily obtained during
routine physician visits, in clinics at the place of employment,
and other situations, a fasting test may still be inconvenient,
uncomfortable, and cumbersome. Therefore, there is still a need for
a diabetes-screening method which is fast and can alleviate the
discomfort and inconvenience for patients.
SUMMARY OF THE INVENTION
[0005] One object of the invention is to provide a method for rapid
screening of diabetes.
[0006] Another object of the invention is to provide kits for rapid
screening of diabetes.
[0007] These and other objects of the invention are met by the
various aspects of the invention described herein.
[0008] The invention, in one aspect, provides a method for rapidly
screening for diabetes, the method comprising the steps of:
collecting a first tear fluid from a patient using a first
tear-collecting device; assaying a specific amount of the first
tear fluid to determine a first glucose concentration;
administering orally a load of carbohydrate to the patient;
collecting a second tear fluid, at a period of time of less than 50
minutes after orally administering of the load of carbohydrate,
using a second tear-collecting device; assaying a specific amount
of the second tear fluid to determine a second glucose
concentration; comparing the second glucose concentration with the
first glucose concentration to determine if the patient is likely
to be a diabetic.
[0009] The invention, in another aspect, provides a method for
rapidly screening diabetes, the method comprising the steps of:
contacting a glucose-sensing ophthalmic device with an ocular
fluid, wherein the glucose-sensing ophthalmic device comprises a
testing agent composition which specifically and reversibly
interacts with glucose to form a detectable signal which changes in
a concentration-dependent manner; determining by means of the
glucose-sensing ophthalmic device a first glucose concentration in
the ocular fluid; administering orally a load of carbohydrate to
the patient; at a period of time of less than 50 minutes after
orally administering of the load of carbohydrate, determining by
means of the glucose-sensing ophthalmic device a second glucose
concentration in the ocular fluid; and comparing the second glucose
concentration with the first glucose concentration to determine if
the patient is likely to be a diabetic.
[0010] The invention, in a still further aspect, provides a kit for
screening for diabetes, the kit comprising: (1) a glucose-sensing
ophthalmic device, wherein the glucose-sensing ophthalmic device
comprises a testing agent composition which specifically and
reversibly interacts with glucose to form a detectable signal which
changes in a concentration-dependent manner; or (2) two or more
tear-collecting devices selected from the group consisting of a
strip, a capillary tube, and a soft-hydrogel contact lens, and a
testing agent composition which specifically reacts or interacts
with glucose to form a detectable signal which changes in a
concentration-dependent manner, wherein said strip has a first end
and an opposite second end and preferably has substantially uniform
cross-sections from the first end to the second end, wherein said
strip is made of a hydrogel material in a substantially dry state
and is characterized by having a substantially uniform swelling
along the hydrogel strip from the first end to the second end when
fully wicked by a tear fluid and by having a correlation between
the volume of tear uptake by said strip and the length of a
tear-wicked end portion of said strip.
BRIEF DESCRIPTION OF THE DRAWINGS
[0011] FIG. 1 shows glucose concentrations in a tear fluid
collected at every 15 minutes after oral administration of a
carbohydrate load to a subject.
DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS
[0012] Reference now will be made in detail to the embodiments of
the invention, one or more examples of which are set forth below.
Each example is provided by way of explanation of the invention,
and is not a limitation of the invention. In fact, it will be
apparent to those skilled in the art that various modifications and
variations can be made in the present invention without departing
from the scope or spirit of the invention. For instance, features
illustrated or described as part of one embodiment, can be used on
another embodiment to yield a still further embodiment. Thus, it is
intended that the present invention covers such modifications and
variations as come within the scope of the appended claims and
their equivalents. Other objects, features and aspects of the
present invention are disclosed in or are obvious from the
following detailed description. It is to be understood by one of
ordinary skill in the art that the present discussion is a
description of exemplary embodiments only, and is not intended as
limiting the broader aspects of the present invention.
[0013] Unless defined otherwise, all technical and scientific terms
used herein have the same meaning as commonly understood by one of
ordinary skill in the art to which this invention belongs.
Generally, the nomenclature used herein and the laboratory
procedures are well known and commonly employed in the art.
Conventional methods are used for these procedures, such as those
provided in the art and various general references. Where a term is
provided in the singular, the inventors also contemplate the plural
of that term. As employed throughout the disclosure, the following
terms, unless otherwise indicated, shall be understood to have the
following meanings.
[0014] The invention, in one aspect, provides a method for rapidly
screening for diabetes, the method comprising the steps of:
collecting a first tear fluid from a patient using a first
tear-collecting device; assaying a specific amount of the first
tear fluid to determine a first glucose concentration;
administering orally a load of carbohydrate to the patient;
collecting a second tear fluid, at a period of time of less than 50
minutes after orally administering of the load of carbohydrate,
using a second tear-collecting device; assaying a specific amount
of the second tear fluid to determine a second glucose
concentration; comparing the second glucose concentration with the
first glucose concentration to determine if the patient is likely
to be a diabetic.
[0015] Any tear-collecting device known to a person skilled in the
art can be used. Examples of tear-collecting devices are glass
capillary tubes, hydrogel strips, and contact lenses.
[0016] Preferably, a tear-collecting device is a hydrogel strip,
which is disclosed in a copending U.S. patent application Ser. No.,
entitled "Methods and Kits For Assays Of Analytes Of Interest In
Tears", filed on Oct. 1, 2002, herein incorporated by reference in
its entirety. The hydrogel strip is made of a hydrogel material in
substantially dry state and has a uniform cross-section, wherein
said strip is characterized by having a substantially uniform
swelling along the hydrogel strip when fully wicked by a tear fluid
and characterized by having a defined correlation between the
volume of tear uptake by said strip and the length of the
tear-wicked end portion of said strip. A hydrogel strip as a
tear-collecting device can offer some advantages over a glass
capillary tube, including, for example, easy of handling, safety,
and low irritation. Furthermore, assays for glucose in an ocular
fluid can be carried out directly on and in one or more divided
pieces of the tear-wicked portion of a hydrogel strip. Or, a tear
fluid absorbed by a hydrogel strip can be substantially recovered
by a method known to a person skilled in the art.
[0017] A "hydrogel material" refers to a polymeric material which
can absorb at least 10 percent by weight of water when it is fully
hydrated. Generally, a hydrogel material is obtained by
polymerization or copolymerization of at least one hydrophilic
monomer in the presence of or in the absence of additional monomers
and/or macromers.
[0018] A "monomer" means a low molecular weight compound that can
be polymerized. Low molecular weight typically means average
molecular weights less than 700 Daltons.
[0019] A "macromer" refers to a medium and high molecular weight
compound or polymer that contains functional groups capable of
further polymerization. Medium and high molecular weight typically
means average molecular weights greater than 700 Daltons.
[0020] A "hydrophilic vinylic monomer" refers to a monomer which as
a homopolymer typically yields a polymer that is water-soluble or
can absorb at least 10 percent by weight water. Suitable
hydrophilic vinylic comonomers include, without limitation,
hydroxy-substituted lower alkylacrylates and -methacrylates,
acrylamide, methacrylamide, lower alkyl-acrylamides and
-methacrylamides, ethoxylated acrylates and methacrylates,
hydroxy-substituted lower alkyl-acrylamides and -methacrylamides,
hydroxy-substituted lower alkylvinyl-ethers, sodium ethylene
sulphonate, sodium styrene sulphonate, 2-acrylamido-2-methyl-pro-
pane-sulphonic acid, N-vinyl pyrrole, N-vinyl succinimide, N-vinyl
pyrrolidone, 2- or 4-vinyl pyridine, acrylic acid, methacrylic
acid, amino- (whereby the term "amino" also includes quaternary
ammonium), mono-lower-alkylamino- or
di-lower-alkylamino-lower-alkyl-acrylates and -methacrylates, allyl
alcohol and the like. Preference is given e.g. to
hydroxy-substituted C.sub.2-C.sub.4-alkyl(meth)acrylates, five- to
seven-membered N-vinyl-lactams,
N,N-di-C.sub.1-C.sub.4-alkyl-methacrylami- des and vinylically
unsaturated carboxylic acids with a total of 3 to 5 carbon atoms.
Examples of suitable hydrophilic vinylic comonomers include
hydroxyethyl methacrylate, hydroxyethyl acrylate, acrylamide,
methacrylamide, dimethylacrylamide, allyl alcohol, vinyl pyridine,
vinyl pyrrolidone, glycerol methacrylate,
N-(1,1-dimethyl-3-oxobutyl)acrylamide- , and the like.
[0021] Any known, suitable hydrogels can be used in the invention.
Exemplary hydrogels include, but are not limited to, poly(vinyl
alcohol) (PVA), modified polyvinylalcohol (e.g., as nelfilcon A),
poly(hydroxyethyl methacrylate), poly(vinyl pyrrolidone), PVAs with
polycarboxylic acids (e.g., carbopol), polyethylene glycol,
polyacrylamide, polymethacrylamide, silicone-containing hydrogels,
polyurethane, polyurea, and the like. A hydrogel can be prepared
according to any methods known to a person skilled in the art.
[0022] Preferably, a hydrogel strip is placed at a location near
the lateral canthus of an eye to collect tear fluids. "Lateral
canthus" refers to one of the two canthuses of an eye, which is
located away from the nose.
[0023] A hydrogel strip can have any dimension suitable for
collecting tear fluids. A hydrogel strip of the invention has a
length sufficient long to absorb a minimum volume of tear (e.g., at
least about 1 .mu.l). A hydrogel strip is preferably at least 15 mm
in length, more preferably at least 30 mm in length.
[0024] Preferably, the dimension of the cross-section (e.g,
diameter, width, height, etc.) of a hydrogel strip is neither too
small nor too large. Where the dimension of the cross-section of a
hydrogel strip is too small, the hydrogel strip becomes not
structurally steady and/or can become sharp so that it can
potentially cause damages to eye tissues. Where the dimension of
the cross-section of a hydrogel strip is too large, the hydrogel
strip can not access the lateral canthus.
[0025] A hydrogel strip preferably has a uniform cross-section
along the strip. The cross-section of a hydrogel strip of the
invention can have any geometric shape, for example, such as
rectangular, square, circular, triangular, annular ring, or the
like. Preferably, the cross-section of a hydrogel strip has a
rectangular shape. The rectangular cross-section has a width of
from about 1 mm to about 3 mm, preferably from 1.5 mm to 2 mm, and
a height of from 0.5 mm to 1.5 mm, preferably from 0.8 mm to 1.2
mm. Where the cross-section of a hydrogel strip of the invention is
circular, the diameter of the circular cross-section is preferably
from 1 mm to 3 mm, more preferably from 1.5 mm to 2.2 mm.
[0026] A "substantially uniform swelling along the hydrogel strip
when fully wicked by a tear fluid" means that when a hydrogel strip
is fully wicked by a fluid (e.g., a tear), it has a substantially
uniform increase in volume along the length of the hydrogel strip
and no significant change in the geometric shape of the strip can
be observed.
[0027] Correlation between the volume of fluid (e.g., tear) uptake
by said strip and the length of the fluid-wicked end portion of
said strip preferably is a substantially linear relationship. With
a substantially linear correlation, the volume of tear uptake by a
hydrogel strip can be easily quantified. In a preferred embodiment,
the volume of tear uptake is noticeably marked on a hydrogel
strip.
[0028] For example, a hydrogel strip is prepared from poly(vinyl
alcohol) (PVA) and has a dimension of 1.5 mm in width, 1.0 mm in
height, and 30 mm in length.
[0029] Glucose can be assayed directly on a fraction or all of the
tear-wicked portion of the strip or by first recovering the tear
sample from the wicked portion of the strip and then assaying
glucose in the recovered tear sample.
[0030] It is well known to a skilled artisan that the assay of
glucose can be carried out with the help of a testing agent
composition which specifically reacts or interacts with glucose,
leading to formation of a detectable signal. A detectable signal,
for example, can be electrical signals (electrochemical assays), or
optical signals (enzyme assays, binding assays or competitive
binding assays). Exemplary electrical signals are electrical
potentials and currents. "Optical signals" refer to changes in the
optical properties, including, but not limited to, a color
formation, a change in color, fluorescence, luminescence,
chemiluminescence, changes in fluorescence or luminescence
intensity, changes in fluorescence or luminescence lifetimes,
fluorescent anisotropy or polarization, a spectral shift of the
emission spectrum, time-resolved anisotropy decay, and the
like.
[0031] Electrochemical assay of glucose is largely carried out by
using an enzymatic electrode (or biosensor) which consists of a
thin layer of enzymes adsorbed to the active surface of a
transducer. Along with a suitable reference electrode and a
circuit, a biosensor allows to measure either the potential
difference generated between the two electrodes (for potentiometric
measurements) or the current that flows between the two electrodes
(for amperometric measurements). For, example, a glucose biosensor
consists of a carbon electrode with a conductive coating containing
a mixture of glucose oxidase and mediator (e.g., ferrocene or
derivatives thereof) (see, for example, Cass et al.,
"Ferrocene-mediated Enzyme Electrode for Amperometric Determination
of Glucose", Anal. Chem. 56: 667-671 (1984), herein incorporated ny
reference in its entirety). At the working electrode surface
glucose is oxidized by the glucose oxidase enzyme. This reaction
causes the mediator to be reduced. At the fixed potential, applied
between the two electrodes the mediators is oxidized, generating a
signal response which correlates with the glucose concentration in
a sample.
[0032] The hydrogel strip can be served as a medium for performing
an electrochemical assay. For example, the electrochemical assay of
glucose in a tear fluid can be carried out by first collecting an
amount of the tear fluid using a hydrogel strip, then by placing
the whole or fractional tear-wicked portion of the hydrogel strip
in direct contact with an enzyme electrode and a reference
electrode, and finally by applying a fixed potential between the
two electrodes to obtain an amperometric signal (current) that
correlates with the concentration of the analyte of interest.
[0033] Glucose can be assayed based on the Trinder reaction.
Typically in the Trinder reaction, glucose oxidase, in the presence
of oxygen, oxidizes glucose to form gluconic acid and hydrogen
peroxide which in turn reacts with a chromogenic
oxidation/reduction indicator (e.g., phenol,
3-hydroxy-2,4,6-triiodobenzoic acid, 3-hydroxy-2,4,6-tribromobenz-
oic acid, etc.) in the presence of peroxidase to form a color
different from its original color or to generate a
chemiluminescence.
[0034] Binding assays and competitive binding assays have been
widely used in the determination of an analyte of interest in a
sample. Typically, a binding assay (without use of any competitor)
is generally carried out by using a protein or fragment thereof or
a chemical compound (as a receptor) that is capable of binding said
analyte (ligand) in said sample and has a detectable optical signal
(or other detectable signal) that changes in a
concentration-dependent manner when the receptor is bound to said
analyte. A competitive binding assay is based on the competition
between a labeled ligand (analyte) or ligand analogue
(analyte-analogue) and an unlabeled ligand (analyte) in the
reaction with a receptor (e.g., antibody, receptor, transport
protein, chemical compound). The labeled ligand (analyte) or ligand
analogue (analyte-analogue) also is called as a competitor.
[0035] The detectable optical signal results from one or more
labels associated with a receptor and/or a competitor. A label may
be covalently or non-covalently bound to a receptor or a
competitor. A "receptor" refers to a protein or fragment thereof or
a chemical compound that is capable of binding reversibly glucose
in a sample. A "competitor" refers to a molecule or moiety that
competes with glucose for binding to a receptor.
[0036] A wide range of suitable labels are known. For example, the
label may be a fluorescent label. "A fluorescent label" refers to a
moiety that comprises at least one fluorophore and that, when
attached to a molecule, renders such molecules detectable using
fluorescent detection means. Exemplary fluorophores include
xanthene-type dyes, fluorescein-type dyes, rhodamine-type dyes,
cyanine-type dyes, and the like. A fluorophore can also be a
fluorescent protein such as phycobiliproteins.
[0037] The detectable optical signal can be derived from a pair of
fluorophores, a first fluorophore and a second fluorophore. One of
the two fluorophores can be an energy donor, for example the first
fluorophore, which absorbs energy upon excitation at an excitation
wavelength within its absorption spectrum and emits energy at a
wavelength within its emission spectrum, and the other fluorophore
can be an energy acceptor, for example the second fluorophore,
which accepts the energy emitted by the donor at a wavelength
within the absorption spectrum of the acceptor and emits energy at
a wavelength within the emission spectrum of the acceptor. The
wavelength of the absorption maximum of the donor fluorophore is
shorter than the wavelength of the absorption maximum of the
acceptor fluorophore; and the wavelength of the emission maximum of
the donor fluorophore is shorter than the wavelength of the
emission maximum of the acceptor fluorophore. It is known that the
energy transfer efficiency depends on the several factors such as
spectral overlap between the emission spectrum of the donor and the
absorption spectrum of the acceptor, spatial distance between donor
and acceptor fluorophores, relative orientation of donor and
acceptor fluorophore, quantum yield of the donor and excited state
lifetime of the donor. It is well known to a person skilled in the
art how to select a donor fluorophore and a acceptor fluorophore.
In a binding assay system, the energy donor fluorophore and the
energy acceptor fluorophore each can be bound to a receptor and
spaced such that there is a detectable optical signal when the
receptor is bound to the analyte. In a competitive binding assay
system, one of the energy donor fluorophore and the energy acceptor
fluorophore can be bound to the receptor and the other can be bound
to the competitor.
[0038] It is understood that the above energy acceptor fluorophore
can be replaced by a non-fluorescent energy transfer acceptor, for
example, such as a dye which accepts the energy emitted by the
donor fluorophore at a wavelength within the absorption spectrum of
the acceptor but does not emits energy in the form of fluorescence
or luminescence.
[0039] A fluorescent label can intrinsically be part of the
receptor. For example, a receptor can be a fusion protein
comprising at least the fluorescent part of a fluorescent protein
and at least the binding part of a receptor protein. Alternatively,
the fluorescent label can be a fluorescent label which is not
naturally associated with the receptor moiety but which is attached
by means of a chemical linkage, such as a covalent bond.
[0040] A fluorescent label can intrinsically be part of the
competitor. Alternatively, the fluorescent label can be a
fluorescent label which is not naturally associated with the
competitor moiety but which is attached by means of a chemical
linkage, such as a covalent bond.
[0041] One example of binding assay is an assay for glucose
disclosed in U.S. Pat. No. 6,197,534, using an E. coli
glucose/galactose binding protein ("GGBP") as previously described
(Scholle, et al., Mol. Gen. Genet 208:247-253 (1987)), or
functionally equivalent fragments thereof. As a sensor for glucose
monitoring, GGBP has several favorable features including a single
glucose binding site and high affinity for glucose; GGBP binds
glucose with a dissociation constant near 0.8 .mu.M. Like similar
transport proteins from other bacteria, GGBP is highly specific for
binding glucose and/or galactose. The apparent binding affinity of
GGBP for sugars other than glucose or galactose is typically
100-1000 fold weaker [Boos, et al., J. Biol. Chem. 247(3):917-924
(1972); Boos, W., J. Biol. Chem. 247(17):5414-5424 (1972); Strange
and Koshland, Proc. Nat'l Acad. Sci. USA 73(3):762-766 (1976);
Zukin, et al., Biochemistry 16(3):381-386 (1977)). The high
affinity for glucose also will allow to measure .mu.M glucose
concentrations in a tear fluid. GGBP can be labeled with one
fluorescence energy donor moiety and one fluorescence energy
acceptor at two specific position on GGBP in a manner so that there
is a detectable spectral change (e.g., change in fluorescence
intensity or lifetime) when GGBP is bound to glucose.
[0042] One example of a competitive binding assay is a glucose
assay disclosed in U.S. patent application Ser. No. 09/784,471
(herein incorporated by reference), using a glucose-sensing system
which comprises tetramethylrhodamine isothiocyanate concanavalin A
(TRITC-ConA) as a receptor, fluorescein isothiocyanate dextran
(FITC-dextran) as a competitor. While the FITC-dextran is bound to
the TRITC-ConA, the FITC fluorescence is quenched by TRITC via a
fluorescence resonance energy transfer. Increased glucose
concentration frees the FITC-dextran and results in fluorescence
which is proportional to glucose concentration.
[0043] The hydrogel strip can be served as a medium for performing
a binding assay or a competitive binding assay using a testing
agent composition which specifically reacts or interacts with
glucose to form a detectable signal that changes in a
concentration-dependent manner.
[0044] Where glucose in a tear fluid is assayed based on a binding
assay, the testing agent composition preferably comprises a
receptor that is capable of binding glucose and has a detectable
optical signal that changes in a concentration-dependent manner
when the receptor is bound reversibly to glucose, wherein said
detectable optical signal results from one or more labels
associated with the receptor. More preferably, the testing agent
composition comprises: (1) a fluorescence energy donor and a
fluorescence energy acceptor; or (2) a fluorescence energy donor
and a non-fluorescence energy acceptor.
[0045] Where glucose in a tear fluid is assayed based on a
competitive binding assay, the testing agent composition preferably
comprises a receptor having a first label associated therewith, a
competitor having a second label associated therewith, wherein one
of the first and second labels is a fluorescent energy donor and
the other one is a fluorescent or non-fluorescent energy acceptor.
Binding of both the competitor and glucose to the receptor is
reversible.
[0046] A testing agent composition can be a solution or can be
incorporated partially or fully in a hydrogel strip. For example,
the receptor can be covalently bound to the strip material. The
receptor can be covalently linked to the strip material according
to any known, suitable methods.
[0047] Similarly, a competitor can be tethered, preferably via a
flexible linker, to the strip material according to any known,
suitable methods. Introduction of flexible linkers into a polymer
or a competitor or receptor is known to a person skilled in the
art.
[0048] Any known suitable competitor can be used in the competitive
binding assays of glucose. For example, a glucose competitor can be
a dextran (which competes with glucose for binding to Concanavalin
A). Another exemplary glucose competitors are 2-deoxy-D-glucose,
D-mannose and D-galactose (which competes with glucose for binding
to inactivated glucose oxidase). Further exemplary glucose
competitors are glucose-protein conjugates (such as a conjugate of
glucose and albumin, obtained by covalently attaching glucose onto
the surface of albumin).
[0049] Exemplary receptors for glucose include, but are not limited
to Concanavalin A (Mansouri & Schultz, Bio/Tech 2, 385, 1984),
GGBP, inactivated glucose oxidase (e.g., apo-glucose oxidase or the
like), inactivated glucose dehydrogenase (e.g., apo-glucose
dehydrogenase or the like), boronic acid, or a genetically
engineered glucose binding protein or fragments thereof.
[0050] Preferably, at least one component or all components of a
testing agent composition can be impregnated in a hydrogel strip
for rapidly screening for diabetes.
[0051] Detectable signals can be detected by any method known to a
person skilled in the art. For example, if the label is a
luminescent label, the detector may include a luminometer; if the
label is a calorimetric label, the detector may include a
colorimeter; if the label is a fluorescent label, the detector may
include a fluorophotometer. Construction of such devices is well
known in the art. Light with wavelengths which will excite the
fluorescent label can be provided, for example, by a laser or a
light source, such as a light-emitting diode.
[0052] The invention, in another aspect, provides a method for
rapidly screening of diabetes, the method comprising the steps of:
contacting a glucose-sensing ophthalmic device with an ocular
fluid, wherein the glucose-sensing ophthalmic device comprises a
testing agent composition which specifically and reversibly
interacts with glucose to form a detectable signal which changes in
a concentration-dependent manner; determining by means of the
glucose-sensing ophthalmic device a first glucose concentration in
the ocular fluid; administering orally a load of carbohydrate to
the patient; at a specific period of time (less than 50 minutes)
after orally administering the load of carbohydrate, determining by
means of the glucose-sensing ophthalmic device a second glucose
concentration in the ocular fluid; and comparing the second glucose
concentration with the first glucose concentration to determine if
the patient is likely to be a diabetic.
[0053] In a preferred embodiment, the glucose-sensing ophthalmic
device comprises a receptor (e.g., a protein or fragment thereof or
a chemical compound) that is capable of binding glucose and has a
detectable optical signal that changes in a concentration-dependent
manner when the receptor is bound to glucose, wherein said
detectable optical signal results from one or more labels
associated with the receptor. More preferably, the detectable
optical signal results from: (1) a fluorescence energy donor and a
fluorescence energy acceptor; or (2) a fluorescence energy donor
and a non-fluorescence energy acceptor, wherein the energy donor
and acceptor are associated with the receptor. As described above,
a fluorescent energy donor can be a fluorescent label; a
fluorescent energy acceptor can be a fluorescent label; and a
non-fluorescent energy donor can be a dye moiety.
[0054] An "ophthalmic device", as used herein, refers to a contact
lens (hard or soft), an intraocular lens, a corneal onlay, other
ophthalmic devices (e.g., stents, implants, or the like) used on or
about the eye or ocular vicinity, and cases or containers for
storing ophthalmic devices or ophthalmic solutions.
[0055] A fluorescent energy acceptor and/or acceptor can
intrinsically be part of the receptor. For example, a receptor can
be a fusion protein comprising at least the fluorescent part of a
fluorescent protein and at least the binding part of a receptor
protein. Alternatively, the fluorescent label can be a fluorescent
label which is not naturally associated with the receptor moiety
but which is attached by means of a chemical linkage, such as a
covalent bond.
[0056] In another preferred embodiment, the glucose-sensing
ophthalmic device comprises a receptor having a first label
associated therewith, a competitor having a second label associated
therewith, wherein one of the first and second labels is a
fluorescent energy donor and the other one is a fluorescent or
non-fluorescent energy acceptor. Binding of both the competitor and
glucose to the receptor is reversible. Exemplary glucose-sensing
ophthalmic devices are those disclosed in copending U.S. patent
application Ser. No. 09/784,471 (herein incorporated by
reference).
[0057] A fluorescent energy donor can intrinsically be part of the
receptor. For example, a receptor can be a fusion protein
comprising at least the fluorescent part of a fluorescent protein
and at least the binding part of a receptor protein. Alternatively,
the fluorescent energy donor can be a fluorescent label which is
not naturally associated with the receptor moiety but which is
attached by means of a chemical linkage, such as a covalent
bond.
[0058] A fluorescent or non-fluorescent energy acceptor can
intrinsically be part of the competitor. Alternatively, the
fluorescent or non-fluorescent energy acceptor can be a fluorescent
label or dye which is not naturally associated with the competitor
moiety but which is attached by means of a chemical linkage, such
as a covalent bond.
[0059] A variety of options are available for providing the
receptor and competitor moieties in an ophthalmic lens.
Construction of various types of ophthalmic devices is well known
in the art. Construction of contact lenses is taught, for example,
in U.S. Pat. Nos. 5,965,631, 5,894,002, 5,849,811, 5,807,944,
5,776,381, 5,426,158, 4,099,859, 4,229,273, 4,168,112, 4,217,038,
4,409,258, 4,388,164, 4,332,922, 4,143,949, 4,311,573, 4,589,964,
and 3,925,178.
[0060] Construction of intraocular lens implants is taught, inter
alia, in U.S. Pat. Nos. 6,051,025, 5,868,697, 5,762,836, 5,609,640,
5,071,432, 5,041,133, and 5,007,928. Subconjunctival lenses are
taught, for example, in U.S. Pat. Nos. 5,476,511, 5,400,114, and
5,127,901. Intracorneal lenses are taught, inter alia, in U.S. Pat.
Nos. 6,090,141, 5,984,961, 5,123,921, and 4,799,931.
[0061] The receptor and/or receptor can be covalently bound or
tethered to the ophthalmic device material which comprises a
polymer meshwork containing pores. The pores are of a size which
permit glucose to be bound reversibly to the receptor.
[0062] The receptor can be covalently linked to a polymer meshwork
according to any known, suitable methods. A polymer meshwork can
comprise or be modified to comprise reactive moieties such as
groups containing amine, hydroxy, isothiocyanate, isocyanate,
monochlorotriazine, dichlorotriazine, mono- or di-halogen
substituted pyridine, mono- or di-halogen substituted diazine,
phosphoramidite, maleimide, aziridine, sulfonyl halide, acid
halide, hydroxysuccinimide ester, hydroxysulfosuccinimide ester,
imido ester, hydrazine, axidonitrophenyl, azide, 3-(2-pyridyl
dithio)proprionamide, glyoxal and aldehyde. For example, a cyclic
acetate can be introduced into a PVA meshwork via an aldehyde group
containing a function end-group such as an amine. The amino
end-group can be modified into an isocyanate or isothiocyanate. A
reactive moieties of a polymer meshwork can be reacted with a
function group on the receptor to form a covalent bond. Exemplary
functional groups include but are not limited to amine, hydroxy and
sulfhydryl.
[0063] Similarly, a competitor can be tethered, preferably via a
flexible linker, to a polymer meshwork according to any known,
suitable methods. Introduction of flexible linkers into a polymer
meshwork or a competitor is known to a person skilled in the art. A
flexible linker may not have significantly adverse effects on the
binding affinity of the competitor to the receptor while
eliminating the out-diffusion of the competitor, especially small
competitor molecule.
[0064] Where the receptor and/or competitor are not covalently
bound to the ophthalmic device material which comprises a polymer
meshwork containing pores. The pores are of a size which permit the
receptor to bind reversibly glucose and/or the receptor, but which
prevent the receptor and the competitor from diffusing out of the
ophthalmic device. Suitable polymers for this purpose are known in
the art and include hydrogels, such as stable polymers of
polyethylene glycol hydrogel (PEGH), polyvinylalcohols (PVA),
modified polyvinylalcohol (e.g., as nelfilcon A), PVAs with
polycarboxylic acids (e.g., carbopol) which may contain
crosslinkable functional groups, water-soluble macromer or
polymers, starpolymers, dendrimers, and other biopolymers
[0065] In another embodiment, the ophthalmic device can comprise a
glucose-sensing LbL coating which is not covalently attached to the
core material of the ophthalmic device, wherein the glucose-sensing
LbL coating comprises a testing agent composition which
specifically and reversibly interacts with glucose to form a
detectable signal which changes in a concentration-dependent
manner.
[0066] In a preferred embodiment, the glucose-sensing LbL coating
comprises at least one layer of a receptor (e.g., a protein or
fragment thereof or a chemical compound) that is capable of binding
glucose and has a detectable optical signal that changes in a
concentration-dependent manner when the receptor is bound to
glucose, wherein said detectable optical signal results from one or
more labels associated with the receptor. More preferably, the
detectable optical signal results from: (1) a fluorescence energy
donor and a fluorescence energy acceptor; or (2) a fluorescence
energy donor and a non-fluorescence energy acceptor, wherein the
energy donor and acceptor are associated with the receptor. As
described above, a fluorescent energy donor can be a fluorescent
label; a fluorescent energy acceptor can be a fluorescent label;
and a non-fluorescent energy donor can be a dye moiety.
[0067] In another preferred embodiment, the glucose-sensing LbL
coating comprises one or more layers of a receptor having a first
label associated therewith and one or more layers of a competitor
having a second label associated therewith, wherein one of the
first and second labels is a fluorescent energy donor and the other
one is a fluorescent or non-fluorescent energy acceptor, wherein
each layer of the receptor is separated from each layer of the
competitor by one or more spacing polyelectrolyte layers. Each
polyelectrolyte layer includes one or more polyelectrolytes, which
are generally high molecular weight polymers with multiple ionic or
ionizable functional groups.
[0068] "LbL coating", as used herein, refers to a coating that is
not covalently attached to the surface of an article and is
obtained by layer-by-layer ("LbL") deposition of polyelectrolytes
on the article. An LbL coating can be a single layer or a bilayer
or multiple bilayers.
[0069] The term "bilayer" is employed herein in a broad sense and
is intended to encompass, a coating structure formed by
non-covalently applying first one layer of a first coating material
and then one layer of a second coating material having charges
opposite the charges of the first coating material. It should be
understood that the layers of the first and second coating
materials may be intertwined with each other in the bilayer.
[0070] As used herein, a "polyionic material" refers to a polymeric
material that has a plurality of charged groups, such as
polyelectrolytes, p- and n-type doped conducting polymers.
Polyionic materials include both polycationic (having positive
charges) and polyanionic (having negative charges) materials.
[0071] A polycationic material used in the present invention can
generally include any material known in the art to have a plurality
of positively charged groups along a polymer chain. For instance,
suitable examples of such polycationic materials can include, but
are not limited to, poly(allylamine hydrochloride) (PAH),
poly(ethyleneimine) (PEI), poly(vinylbenzyltriamethylamine) (PVBT),
polyaniline (PAN or PANI) (p-type doped) [or sulphonated
polyaniline], polypyrrole (PPY) (p-typed doped), and
poly(pyridinium acetylene).
[0072] A polyanionic material used in the present invention can
generally include any material known in the art to have a plurality
of negatively charged groups along a polymer chain. For example,
suitable polyanionic materials can include, but are not limited to,
polymethacrylic acid (PMA), polyacrylic acid (PAA),
poly(thiophene-3-acetic acid) (PTAA), poly(4-styrenesulfonic acid)
(PSS), sodium poly(styrene sulfonate) (SPS) and poly(sodium styrene
sulfonate) (PSSS).
[0073] The foregoing lists are intended to be exemplary, but
clearly are not exhaustive. A person skilled in the art, given the
disclosure and teaching herein, would be able to select a number of
other useful polyionic materials.
[0074] In order to alter various characteristics of the coating,
such as thickness, the molecular weight of the polyionic materials
can be varied. In particular, as the molecular weight is increased,
the coating thickness generally increases. However, if the increase
in molecular weight increase is too substantial, the difficulty in
handling may also increase. As such, polyionic materials used in a
process of the present invention will typically have a molecular
weight M.sub.n of about 2,000 to about 150,000. In some
embodiments, the molecular weight is about 5,000 to about 100,000,
and in other embodiments, from about 75,000 to about 100,000.
[0075] Any suitable LbL polyelectrolyte deposition techniques can
be used in the LbL coating. It has been discovered and disclosed in
U.S. application Ser. No. 09/005,317 that complex and
time-consuming pretreatment of a core material (medical device) is
not required prior to non-covalently binding of a polyionic
material to the core material. By simply contacting a core material
of a medical device, for example, a contact lens, with one or more
solutions each containing one or more polyionic materials, an LbL
coating can be formed on a medical device to modify the surface
properties of the core material of the medical device.
[0076] Application of an LbL coating may be accomplished in a
number of ways as described in pending U.S. patent applications
(application Ser. Nos. 09/005,317,09/774,942, 09/775,104), herein
incorporated by reference in their entireties. One coating process
embodiment involves solely dip-coating and dip-rinsing steps.
Another coating process embodiment involves solely spray-coating
and spray-rinsing steps. However, a number of alternatives involve
various combinations of spray- and dip-coating and rinsing steps
may be designed by a person having ordinary skill in the art.
[0077] One dip-coating alternative involves the steps of applying a
coating of a first polyionic material to a core material of a
medical device by immersing said medical device in a first solution
of a first polyionic material; rinsing the medical device by
immersing the medical device in a rinsing solution; and,
optionally, drying the medical device. This procedure can be
repeated using a second polyionic material, with the second
polyionic material having charges opposite of the charges of the
first polyionic material, in order to form a polyionic bilayer.
This bilayer formation process may be repeated a plurality of times
in order to produce a thicker LbL coating. A preferred number of
bilayers is about 5 to about 20 bilayers. While more than 20
bilayers are possible, it has been found that delamination may
occur in some LbL coatings having an excessive number of
bilayers.
[0078] The immersion time for each of the coating and rinsing steps
may vary depending on a number of factors. Preferably, immersion of
the core material into the polyionic solution occurs over a period
of about 1 to 30 minutes, more preferably about 2 to 20 minutes,
and most preferably about 1 to 5 minutes. Rinsing may be
accomplished in one step, but a plurality of rinsing steps can be
quite efficient.
[0079] Another embodiment of the coating process is a single
dip-coating process as described in U.S. application Ser. No.
09/775,104, herein incorporated by reference in its entirety. Such
single dip-coating process involves dipping a core material of a
medical device in a solution containing a negatively charged
polyionic material and a positively charged polyionic material in
an amount such that the molar charge ratio of said solution is from
about 3:1 to about 100:1. Multiple bilayers can be formed on a
medical device by using this single dip-coating process.
[0080] A sensing layer (receptor or competitor layer) can be
prepared by adding a receptor or a competitor into, a coating
solution for forming part of a bilayer. When receptor or
competitor, which is added into a coating solution, preferably, has
a charge. By having a positive or negative charge, the receptor or
competitor can be substituted for the charged polymeric material in
solution at the same molar ratio. It should be understood, however,
that non-charged receptor or competitor can also be applied to the
core material of an article by entrapment.
[0081] Alternatively, a sensing layer (receptor or competitor
layer) can be prepared by first entrapping within a vesicle with a
charged surface and then non-covalently applying a layer of the
vesicle with a receptor and/or receptor entrapped therein.
[0082] In accordance with the present invention, vesicles include
liposomes, polymerized micelles, and nanocapsules and microcapsules
each having a multilayered shell of polyelectrolytes. The
entrapment of a receptor and/or competitor in a vesicle can be
carried out according to any known suitable method. Then, an LbL
coating can be formed on an article by any suitable known
layer-by-layer deposition technique to contain at least one bilayer
of a vesicle with a charged surface and with a receptor and a
competitor entrapped therein and a polyionic material having
charges opposite of the charges of the vesicle.
[0083] In another preferred embodiment, the glucose-sensing LbL
coating comprises one or more layers of a vesicle with a charged
surface and with a receptor and a competitor entrapped therein,
wherein the receptor has a first label associated therewith and the
competitor has a second label associated therewith, wherein one of
the first and second labels is a fluorescent energy donor and the
other one is a fluorescent or non-fluorescent energy acceptor. Each
polyelectrolyte layer includes one or more polyelectrolytes, which
are generally high molecular weight polymers with multiple ionic or
ionizable functional groups.
[0084] The sensing layers (receptor layers and/or competitor
layers) and spacing polyelectrolyte layers are deposited as uniform
thin films (1-10 nm) in 10-15 deposition cycles onto the core
material of an ophthalmic device, resulting in only a 100-500 nm
thick coating for the sensing film, which is highly biocompatible.
A typical sequence for construction of an ophthalmic lens suitable
for glucose detection involves a deposition cycle of ultrathin
(1-10 nm) films of PAA, PAH, PM, concanavalin A, PM, PAH, PM,
fluorescein dextran, PM, PAH, PM, concanavalin A, PM, fluorescein
dextran, PM, etc.
[0085] The invention, in a still further aspect, provides a kit for
screening for diabetes, the kit comprising: (1) a glucose-sensing
ophthalmic device, wherein the glucose-sensing ophthalmic device
comprises a testing agent composition which specifically and
reversibly interacts with glucose to form a detectable signal which
changes in a concentration-dependent manner; or (2) two or more
tear-collecting devices selected from the group consisting of a
strip, a capillary tube, and a soft-hydrogel contact lens, and a
testing agent composition which specifically reacts or interacts
with glucose to form a detectable signal which changes in a
concentration-dependent manner, wherein said strip has a first end
and an opposite second end and preferably has substantially uniform
cross-sections from the first end to the second end, wherein said
strip is made of a hydrogel material in a substantially dry state
and is characterized by having a substantially uniform swelling
along the hydrogel strip from the first end to the second end when
fully wicked by a tear fluid and by having a correlation between
the volume of tear uptake by said strip and the length of a
tear-wicked end portion of said strip.
[0086] Methods of kits of the invention according to embodiments of
the invention are useful for rapid screening for diabetes. For
example, screenings for diabetes can be carried as follows.
[0087] In an in vitro screening assay, subjects could come into an
eye practitioner with or without fasting and have a tear sample
taken using a first strip of the invention. The subject would then
be given an oral carbohydrate load (e.g. 75 g of glucose) and a
subsequent sample taken with a second strip of the invention after
a defined time period (e.g. from 15 minutes to 30 minutes). The
wicked portion of each of the first and second strips would be
assayed for glucose, and, if there was a substantial rise in the
tear glucose value during that time interval (e.g. 1.5 fold), then
the person would be referred for follow-up and diagnosis to a
general physician. No follow-up would be required if the person did
not get this rise in tear glucose.
[0088] In an in vivo screening assay, subjects could come into an
eye practitioner with or without fasting and have a tear sample
taken using a first strip of the invention. The subject would then
be given an oral carbohydrate load (e.g. 75 g of glucose) and a
subsequent sample taken with a second strip of the invention after
a defined time period (e.g. from 15 minutes to 30 minutes). The
wicked portion of each of the first and second strips would be
assayed for glucose, and, if there was a substantial rise in the
tear glucose value during that time interval (e.g. 1.5 fold), then
the person would be referred for follow-up and diagnosis to a
general physician. No follow-up would be required if the person did
not get this rise in tear glucose.
[0089] The previous disclosure will enable one having ordinary
skill in the art to practice the invention. In order to better
enable the reader to understand specific embodiments and the
advantages thereof, reference to the following examples is
suggested.
EXAMPLE 1
[0090] A tear sample is collected using a microcapillary tube from
a fasting subject before being given orally a 54 g carbohydrate
load. Another tear sample is collected from the same subject at
every 15 minutes after oral administration of the carbohydrate
load. The last tear sample is collected at 180 minutes after oral
administration of the carbohydrate load. 12 subjects (3 normals, 9
diabetics) have been screened. Glucose concentration, in these tear
samples, is measured using a modified Amplex Red Assay (glucose
oxidase and peroxidase, with a fluorescent substrate). FIG. 1 shows
the average tear glucose values (+Standard Deviations) between
normals and diabetics every 15 minutes. At 15 minutes, there is
noticeable difference between normals and diabetics. At 30, 45 and
60 minutes there is definitively a statistically significant
difference between normals and diabetics.
* * * * *