U.S. patent application number 13/985731 was filed with the patent office on 2014-03-27 for near-infrared silver sulfide quantum dot, preparation method therefor and biological application thereof.
This patent application is currently assigned to Suzhou Institute of Nano-Bionics, Shinese Academy of Sciences. The applicant listed for this patent is Qiangbin Wang, Yejun Zhang. Invention is credited to Qiangbin Wang, Yejun Zhang.
Application Number | 20140087409 13/985731 |
Document ID | / |
Family ID | 45102899 |
Filed Date | 2014-03-27 |
United States Patent
Application |
20140087409 |
Kind Code |
A1 |
Wang; Qiangbin ; et
al. |
March 27, 2014 |
NEAR-INFRARED SILVER SULFIDE QUANTUM DOT, PREPARATION METHOD
THEREFOR AND BIOLOGICAL APPLICATION THEREOF
Abstract
Provided are near-infrared silver sulfide quantum dots, a
preparation method thereof and a biological application thereof.
The silver sulfide quantum dots have hydrophilic groups derived
from a mercapto-containing hydrophilic reagent attached on the
surface thereof, and the hydrophilic reagent is any one of
mercaptoacetic acid, mercaptopropionic acid, cysteine, cysteamine,
thioctic acid and ammonium mercaptoacetate or any combination
thereof. The silver sulfide quantum dots have high fluorescence
yield, good fluorescence stability, good biocompatibility and
uniform sizes. The preparation method has moderate reaction
conditions, simple operation, short production cycle, good
reproducibility and is easy to control. The silver sulfide quantum
dots can be used in the application of cellular imaging and
biological tissue imaging.
Inventors: |
Wang; Qiangbin; (Suzhou,
CN) ; Zhang; Yejun; (Suzhou, CN) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Wang; Qiangbin
Zhang; Yejun |
Suzhou
Suzhou |
|
CN
CN |
|
|
Assignee: |
Suzhou Institute of Nano-Bionics,
Shinese Academy of Sciences
Suzhou
CN
|
Family ID: |
45102899 |
Appl. No.: |
13/985731 |
Filed: |
February 10, 2012 |
PCT Filed: |
February 10, 2012 |
PCT NO: |
PCT/CN2012/000167 |
371 Date: |
August 15, 2013 |
Current U.S.
Class: |
435/29 ; 549/39;
556/117 |
Current CPC
Class: |
C07F 1/005 20130101;
A61K 49/0067 20130101; C01G 5/00 20130101; A61K 49/0013 20130101;
C09K 11/025 20130101; C09K 11/582 20130101; G01N 33/588
20130101 |
Class at
Publication: |
435/29 ; 556/117;
549/39 |
International
Class: |
G01N 33/58 20060101
G01N033/58 |
Foreign Application Data
Date |
Code |
Application Number |
May 30, 2011 |
CN |
201110142093.8 |
Claims
1. Near-infrared silver sulfide quantum dots, characterized in that
the silver sulfide quantum dots have hydrophilic groups attached to
the surface thereof, the hydrophilic groups are derived from
mercapto-containing hydrophilic reagent, and the hydrophilic
reagent is any one of mercaptoacetic acid, mercaptopropionic acid,
cysteine, cysteamine, thioctic acid and ammonium mercaptoacetate or
any combination thereof.
2. A method for preparation of near-infrared silver sulfide quantum
dots, characterized in that the method comprises the following
steps: 1) preparing hydrophobic silver sulfide quantum dots; and 2)
reacting the hydrophobic silver sulfide quantum dots in step 1)
with stoichiometric or excessive amount of mercapto-containing
hydrophilic reagent in polar organic solvent to allow the surface
thereof to be attached with hydrophilic groups, so as to obtain the
hydrophilic near-infrared silver sulfide quantum dots; the
hydrophilic reagent is any one of mercaptoacetic acid,
mercaptopropionic acid, cysteine, cysteamine, thioctic acid and
ammonium mercaptoacetate or any combination thereof.
3. The method for preparation of near-infrared silver sulfide
quantum dots according to claim 2, characterized in that in step
2), the hydrophobic silver sulfide quantum dots are reacted with
the mercapto-containing hydrophilic reagent in the polar organic
solvent at 2-80.degree. C. for 3 or more hours.
4. The method for preparation of near-infrared silver sulfide
quantum dots according to claim 2, characterized in that in step
2), the pH value of the reaction system is adjusted to 7-14.
5. The method for preparation near-infrared silver sulfide quantum
dots according to claim 2, characterized in that in step 2), the
polar organic solvent comprises any one or more of ethanol,
methanol, acetone and 1-methyl-2-pyrrolidone.
6. The method for preparation of near-infrared silver sulfide
quantum dots according to claim 2, characterized in that the step
1) comprises the following steps: 1-1) heating a mixed reaction
system containing a silver source and a long chain thiol to
80-350.degree. C. in a closed environment, to react sufficiently;
and 1-2) naturally cooling the mixed reaction system to room
temperature and then adding a polar solvent, centrifuging and
washing to obtain the hydrophobic near-infrared silver sulfide
quantum dots; wherein the silver source comprises one or more of
silver nitrate, silver diethyldithiocarbamate, silver
dihydrocarbyldithiophosphate, dioctyl silver sulfosuccinate, silver
thiobenzoate, silver acetate, silver dodecanoate, silver
tetradecanoate and silver octadecanoate; and the long chain thiol
comprises one or more of octanethiol, undecanethiol, dodecanethiol,
tridecanethiol, tetradecanethiol, pentadecanethiol,
hexadecanethiol, octadecanethiol, eicosanethiol, hexanethiol,
1,6-hexanedithiol, and 1,8-octanedithiol.
7. The method for preparation of near-infrared silver sulfide
quantum dots according to claim 6, characterized in that in step
1-2), the mixed reaction system further comprises a surfactant
having coordination property, the surfactant is any one of a long
chain alkyl acid, alkylamine, a long chain alcohol, a long chain
thiol and ether or any combination thereof; and the mixed reaction
system is placed in a closed environment to react.
8. The method for preparation of the near-infrared silver sulfide
quantum dots according to claim 2, characterized in that in step
2), the hydrophobic silver sulfide quantum dots are reacted with
the mercapto-containing hydrophilic reagent under the condition of
continuous stirring and/or vibrating and/or sonicating in the polar
organic solvent at 2-80.degree. C. for 3 or more hours.
9. Use of the near-infrared silver sulfide quantum dots according
to claim 1 in cellular imaging and biological tissue imaging.
Description
TECHNICAL FIELD
[0001] The present invention relates to the technical field of
material chemistry and biology. In particular, the present
invention relates to near-infrared silver sulfide quantum dots, a
preparation method thereof and a biological application
thereof.
BACKGROUND
[0002] As a fundamental method of biomedical research, the
fluorescence labelling and detecting technologies play an important
role in the studies at subcellular level, cellular level and in
vivo level. In in vivo imaging, the fluorescence imaging technology
with near-infrared quantum dots has many unique advantages. For
example, it has deeper depth of tissue penetration, and can
overcome the defect that the deep tissue imaging with visible-light
quantum dots is susceptible to the background fluorescence. Thus it
draws broad attention in medical diagnostics, molecular biology,
cellular biology and the like. Currently, all the common
near-infrared quantum dots contain toxic elements such as Cd, Hg,
Pb and the like. Silver sulfide (Ag.sub.2S) quantum dots with low
toxicity or even without toxicity exhibiting near-infrared
fluorescence have been reported (Near-infrared photoluminescent
Ag.sub.2S quantum dots from a single source precursor. J. Am. Chem.
Soc., 2010, 132, 1470), but the particles are relatively large and
the near-infrared fluorescence intensity is not strong enough.
Other literatures regarding Ag.sub.2S have not given the report
regarding fluorescence. Moreover, the Ag.sub.2S reported in those
literatures has poor homogeneity and dispersity and is made by
complex preparation methods. Furthermore, the surface
functionalization of quantum dots, i.e. transformation from
hydrophobic form into hydrophilic form, to make the quantum dots to
be used for biomedical research, has been reported in many
literatures. However, the surface functionalization processes
reported is substantially not suitable for Ag.sub.2S quantum dots,
because all the Ag.sub.2S quantum dots have superlattice structure
and it is difficult to modify the superlattice with conventional
processes. Furthermore, reagents with strong oxidability are not
suitable for the transformation of Ag.sub.2S to its hydrophilic
form. Therefore, it is of great significance to develop a method
for preparation and surface functionalization of Ag.sub.2S quantum
dots, which is simple process and can produce high quality of
Ag.sub.2S quantum dots with uniform particle sizes, good particle
dispersity, high fluorescence intensity and good reproducibility,
so that the Ag.sub.2S quantum dots can be used in the biological
field.
SUMMARY
[0003] To overcome the above problems, an object of the invention
is to provide near-infrared silver sulfide quantum dots. The
near-infrared silver sulfide quantum dots have the advantages such
as high fluorescence yield, fluorescence stability, uniform sizes,
easy preparation process and the like, and may further have good
biocompatibility after surface functionalization, which makes them
useful in biological imaging.
[0004] The near-infrared silver sulfide quantum dots according to
the invention have hydrophilic groups attached to the surface
thereof, which are derived from a mercapto-containing hydrophilic
reagent. The hydrophilic reagent is any one of mercaptoacetic acid,
mercaptopropionic acid, cysteine, cysteamine, thioctic acid and
ammonium mercaptoacetate or any combination thereof.
[0005] In order to overcome the above problems, another object of
the invention is to provide a method for preparation of
near-infrared silver sulfide quantum dots, wherein the method
comprises the following steps:
[0006] 1) preparing hydrophobic silver sulfide quantum dots;
and
[0007] 2) reacting the hydrophobic silver sulfide quantum dots
obtained in step 1) with equivalent or excessive amount of
mercapto-containing hydrophilic reagent in polar organic solvent,
so that the surface of the silver sulfide quantum dots is attached
with hydrophilic groups, to obtain the near-infrared silver sulfide
quantum dots. In the invention, the hydrophilic silver sulfide
quantum dots as prepared have good performance, provided that the
mole number of the mercapto-containing hydrophilic reagent is more
than or equal to that of the hydrophobic silver sulfide quantum
dots. The ratio of the mole number of the mercapto-containing
hydrophilic reagent to that the hydrophobic silver sulfide quantum
dots can be adjusted depending on the actual requirement during the
preparation process, so that the objects of the invention can be
achieved.
[0008] The hydrophilic reagent is any one of mercaptoacetic acid,
mercaptopropionic acid, cysteine, cysteamine, thioctic acid and
ammonium mercaptoacetate or any combination thereof.
[0009] In the method for preparation of near-infrared silver
sulfide quantum dots according to the invention, the polar organic
solvent in step 2) comprises, but not limited to, any one of
ethanol, methanol, acetone and 1-methyl-2-pyrrolidone or any
combination thereof. The pH value of the mixed system of the
hydrophobic silver sulfide quantum dots and the mercapto-containing
hydrophilic reagent in step 2) is adjusted to 7-14, and the mixed
system is reacted in the polar organic solvent at 2-80.degree. C.
for 3 or more hours. In the present invention, the hydrophilic
silver sulfide quantum dots as prepared have good performances,
provided that the reaction time is more than or equal to 3 hours.
The reaction time can be adjusted depending on the actual
requirement during the preparation process, so that the objects of
the invention can be achieved.
[0010] Preferably, the method for preparation of the hydrophobic
silver sulfide quantum dots in step 1) comprises the following
steps:
[0011] 1-1) heating a mixed reaction system containing a silver
source and a long chain thiol to 80-350.degree. C. in a closed
environment to react sufficiently; and
[0012] 1-2) naturally cooling the mixed reaction system to room
temperature, then adding a polar solvent, centrifuging and washing
to obtain the hydrophobic near-infrared silver sulfide quantum
dots;
[0013] wherein the silver source comprises one or more of silver
nitrate, silver diethyldithiocarbamate, silver
dihydrocarbyldithiophosphate, dioctyl silver sulfosuccinate, silver
thiobenzoate, silver acetate, silver dodecanoate, silver
tetradecanoate and silver octadecanoate; and
[0014] the long chain thiol comprises one or more of octanethiol,
undecanethiol, dodecanethiol, tridecanethiol, tetradecanethiol,
pentadecanethiol, hexadecanethiol, octadecanethiol, eicosanethiol,
hexanethiol, 1,6-hexanedithiol, and 1,8-octanedithiol.
[0015] In the method for preparation of near-infrared silver
sulfide quantum dots according to the invention, it is preferred
that the mixed reaction system in step 1-2) further comprises a
surfactant with coordination property, which is any one of a long
chain alkyl acid, alkylamine, a long chain alcohol, and a long
chain thiol and ether or any combination thereof; and the mixture
reaction system is placed in a closed environment to react. More
preferably, in step 2), the hydrophobic silver sulfide quantum dots
were reacted with the mercapto-containing hydrophilic reagent under
the condition of continuous stirring and/or vibrating and/or
sonicating in the polar organic solvent at 2-80.degree. C. for 3 or
more hours.
[0016] In the method for preparation of near-infrared silver
sulfide quantum dots according to the invention, the near-infrared
silver sulfide quantum dots prepared by the method described in the
invention have monoclinic structure and the particle sizes thereof
are below 8 nm.
[0017] The use of the near-infrared silver sulfide quantum dots
according to the invention in the imaging of biological tissues is
provided.
[0018] In the invention, the silver source and the long chain thiol
are used as reactants, and the hydrophobic silver sulfide quantum
dots are nucleated and grown in reaction systems in the presence of
the surfactant with different coordination properties, to obtain
the hydrophobic silver sulfide quantum dots, wherein the long chain
thiol provides the sulfur source and can be used as solvent and
surfactant. Then the surface functionalization of the hydrophobic
silver sulfide quantum dots as prepared is conducted with the
mercapto-containing hydrophilic reagent. Since the mercapto groups
have an excellent binding ability with silver, they can replace
other groups on the surface of the silver sulfide quantum dots,
resulting in the near-infrared silver sulfide quantum dots with low
toxicity, good biocompatibility and high fluorescence yield. The
difference from the modification of the hydrophobic material to the
hydrophilic material in the prior art is that the Ag.sub.2S quantum
dots firstly prepared according to the invention, which have
superlattice structure, can not be modified to hydrophilic
Ag.sub.2S quantum dots by the experimental conditions for
modification in the prior art due to this special structure.
Through numerous experiments, summaries in combination with the
experiences of the inventor, it has been found that the
modification time, which has a significant impact on the
modification effect, is a key experimental condition when the
Ag.sub.2S quantum dots with the special structure are modified.
Moreover, it has been found that better modification effect can be
achieved when the modification time is equal to or more than 3
hours. The longer the time is, the better the modification effect
is. Therefore, the time may be adjusted depending on the actual
requirement during the preparation process. However, the objects of
the invention can be achieved, provided that the time is 3 or more
hours. In addition, the hydrophilic Ag.sub.2S quantum dots after
modification are monodispersed, do not aggregate, have good
hydrophilicity and stability, and can be used for cellular imaging,
and in particular, for the in vivo imaging.
[0019] Specifically, the process of the invention comprises the
following steps: mixing a silver source, a long chain thiol and a
suitable surfactant; placing the mixture into a closed device and
heating to an appropriate temperature for a certain time to conduct
nucleation and growth; then cooling naturally and adding excessive
amount of ethanol; centrifuging and washing to obtain the
hydrophobic silver sulfide quantum dots; then mixing the prepared
hydrophobic silver sulfide quantum dots, a certain amount of
mercapto-containing hydrophilic reagent and ethanol; and stirring,
vibrating or sonicating the mixture to react completely;
centrifuging and washing with water to obtain a low toxic
near-infrared silver sulfide quantum dots with a good
biocompatibility and high fluorescence yield which can be used for
cellular imaging.
[0020] In addition, the above technical solution may further
comprise the following embodiments:
[0021] 1. Different silver sources, different long chain thiols,
different surfactants, different reaction temperatures and
different reaction time can be used in the reaction to adjust the
sizes of the silver sulfide nano particles. For example, it is
possible to enlarge particle sizes by raising the temperature or
extending the reaction time.
[0022] 2. The dispersity of the functionalized Ag.sub.2S quantum
dots in aqueous solution can be changed by adjusting the pH value
(a better dispersity is obtained at pH 7-14). And the emission peak
can be adjusted depending on the different modification by
different mercapto-containing hydrophilic reagent in the
reaction.
[0023] Compared with the prior art, the advantages of the technical
solution of the invention are that the process of the invention has
moderate reaction conditions, simple operation, short production
cycle, and good reproducibility, and is easy to be controlled. The
Ag.sub.2S quantum dots prepared have high fluorescence yield, good
fluorescence stability, excellent biocompatibility and homogeneous
sizes, and can be used for in vitro cellular imaging and in vivo
imaging.
DESCRIPTION OF DRAWINGS
[0024] FIG. 1 is the TEM image of the hydrophobic Ag.sub.2S quantum
dots in Example 1;
[0025] FIG. 2 is the near-infrared fluorescence spectrum of the
hydrophobic Ag.sub.2S quantum dots in Example 1;
[0026] FIG. 3 is the near-infrared fluorescence spectrum of the
hydrophilic Ag.sub.2S quantum dots in Example 1;
[0027] FIG. 4 is the fluorescence photograph of cells specifically
labelled with the near-infrared quantum dots of silver sulfide in
Example 1; and
[0028] FIG. 5 is the fluorescence photograph of the tumor in a
living mouse specifically labelled with the near-infrared quantum
dots of silver sulfide.
DESCRIPTION OF EMBODIMENTS
[0029] The preparation process of the invention is explained in
detail by the specific examples below.
Example 1
[0030] 0.1 mmol of silver diethyldithiocarbamate and 10 g of
dodecanethiol were mixed in a flask, and heated to 200.degree. C.
under a N.sub.2 atmosphere for 1 h. 50 mL of anhydrous ethanol was
added to the solution after the solution was cooled naturally to
room temperature, and then the resultant mixture was centrifuged,
washed and dispersed in cyclohexane. The sample obtained was
identified to be monoclinic Ag.sub.2S quantum dots by X ray
diffraction and transmission electron microscopy (the particle size
thereof is about 5 nm, as shown in FIG. 1), which has a good
near-infrared fluorescence emission spectrum, as shown in FIG. 2.
0.15 g of thioctic acid was added to the above cyclohexane
dispersion, and equal volume of anhydrous ethanol was added, then
the resultant mixture was sonicated in an ultrasonic cleaner for 4
h, centrifuged and washed with deionized water to obtain
water-soluble Ag.sub.2S quantum dots with particle sizes of about 5
nm which still have very strong fluorescence emission, as shown in
FIG. 3. 0.25 mg of the above Ag.sub.2S quantum dots were dispersed
in 100 .mu.L of dimethyl sulfoxide (DMSO), and 50 .mu.L of DMSO
solution containing 0.01 mmol of NHS was mixed with the above
solution. Then 50 .mu.L of DMSO solution containing 0.01 mmol of
EDC was added to the above mixed solution, and the resultant
mixture was packed with aluminum foil, stirred for 1 h, centrifuged
and further dispersed in 100 .mu.L of DMSO. The mixed solution of
15 .mu.L of 2 mg/mL Erbitux and 185 .mu.L of 1.times.PBS was added
to 100 .mu.L of Ag.sub.2S/DMSO mixed solution, and the resultant
mixture was reacted in darkness at 4.degree. C. for 12 h, then
centrifuged at 400 g for 4 min, and then the supernatant was taken.
MDA-MB-468 cells were added to the mixed solution of 100 .mu.L of
the above supernatant and 100 .mu.L of 1.times.PBS, coloured at
4.degree. C. for 2 h, and then washed 3 times with 1.times.PBS
solution. It can clearly be seen that the luminescence was given by
Ag.sub.2S quantum dots in cells by exciting with 658 nm laser,
using 1100 nm filter, and photographing with a 2D InGaAs camera
(see FIG. 4).
Example 2
[0031] 0.1 mmol of silver nitrate, 8 g of dodecanethiol and 5.4 g
of oleylamine were mixed in a three-necked flask, and heated to
180.degree. C. under air for 1 h. After the solution was cooled
naturally to room temperature, 50 mL of anhydrous ethanol was
added. The resultant mixture was centrifuged, washed and dispersed
in cyclohexane. The sample obtained was identified to be monoclinic
Ag.sub.2S quantum dots by X ray diffraction and transmission
electron microscopy, with the particle size below 8 nm, which has a
good near-infrared fluorescence emission spectrum. 0.2 g of
L-cysteine was added to the above cyclohexane dispersion, then
equal volume of anhydrous ethanol was added. The resultant mixture
was stirred for 24 h, then centrifuged and washed with deionized
water to obtain water-soluble Ag.sub.2S quantum dots with particle
sizes of about 8 nm, which still have very strong fluorescence
emission. 0.25 mg of the above Ag.sub.2S quantum dots was dispersed
in 100 .mu.L of dimethyl sulfoxide (DMSO), and 50 .mu.L of DMSO
solution containing 0.01 mmol of NHS was mixed with the above
solution. Then 50 .mu.L of DMSO solution containing 0.01 mmol of
EDC was added to the above mixed solution. The resultant mixture
was packed with aluminum foil, stirred for 1 h, centrifuged and
further dispersed in 100 .mu.L of DMSO. The mixed solution of 15
.mu.L of 2 mg/mL Erbitux and 185 .mu.L of 1.times.PBS was added to
100 .mu.L of Ag.sub.2S/DMSO mixed solution. The resultant mixture
was reacted in darkness at 4.degree. C. for 12 h, then centrifuged
at 400 g for 4 min, and the supernatant was taken. MDA-MB-468 cells
were added to the mixed solution of 100 .mu.L of the above
supernatant and 100 .mu.L of 1.times.PBS, coloured at 4.degree. C.
for 2 h, and then washed 3 times with 1.times.PBS solution. It can
clearly be seen that the luminescence was given by Ag.sub.2S
quantum dots in cells by exciting with 658 nm laser, using 1100 nm
filter, and photographing with a 2D InGaAs camera.
Example 3
[0032] 0.1 mmol of silver thiobenzoate, 8 g of hexadecanethiol and
2 g of trioctylphosphine oxide were mixed in a three-necked flask,
heated to 160.degree. C. under air for 4 h. After the solution was
cooled naturally to room temperature, 50 mL of anhydrous ethanol
was added. The resultant mixture was centrifuged, washed and
dispersed in cyclohexane. 0.1 g of mercaptopropionic acid was added
to the above cyclohexane dispersion, then equal volume of anhydrous
ethanol was added. The resultant mixture was vibrated in a vibrator
for 8 h, centrifuged and washed with deionized water to obtain
water-soluble Ag.sub.2S quantum dots with a particle size of about
6 nm, which still have very strong fluorescence emission. 0.25 mg
of the above Ag.sub.2S quantum dots was dispersed in 100 .mu.L of
dimethyl sulfoxide (DMSO), and 50 .mu.L of DMSO solution containing
0.01 mmol of NHS was mixed with the above solution. Then 50 .mu.L
of DMSO solution containing 0.01 mmol of EDC was added to the above
mixed solution. The resultant mixture was packed with aluminum
foil, stirred for 1 h, centrifuged and further dispersed in 100
.mu.L of DMSO. The mixed solution of 15 .mu.L of 2 mg/mL Erbitux
and 185 .mu.L of 1.times.PBS was added to 100 .mu.L of
Ag.sub.2S/DMSO mixed solution. The resultant mixture was reacted in
darkness at 4.degree. C. for 12 h, then centrifuged at 400 g for 4
min, and then the supernatant was taken. MDA-MB-468 cells were
added to the mixed solution of 100 .mu.L of the above supernatant
and 100 .mu.L of 1.times.PBS, coloured at 4.degree. C. for 2 h, and
then washed 3 times with 1.times.PBS solution. It can clearly be
seen that the luminescence was given by Ag.sub.2S quantum dots in
cells by exciting with 658 nm laser, using 1100 nm filter, and
photographing with a 2D InGaAs camera.
Example 4
[0033] 0.1 mmol of silver hexadecanoate, 5 g of hexadecanethiol and
4 g of octadecylamine were mixed in a three-necked flask and heated
to 200.degree. C. under an Ar atmosphere for 1 h. After the
solution was cooled naturally to room temperature, 50 mL of
anhydrous ethanol was added. The resultant mixture was centrifuged,
washed and dispersed in cyclohexane. 0.12 g of mercaptoacetic acid
was added to the above cyclohexane dispersion, then equal volume of
anhydrous ethanol was added. The resultant mixture was stirred for
24 h, then centrifuged and washed with deionized water to obtain
water-soluble Ag.sub.2S quantum dots with a particle size of about
6 nm, which still have very strong fluorescence emission. 0.25 mg
of the above Ag.sub.2S quantum dots was dispersed in 100 .mu.L of
dimethyl sulfoxide (DMSO), and 50 .mu.L of DMSO solution containing
0.01 mmol of NHS was mixed with the above solution. Then 50 .mu.L
of DMSO solution containing 0.01 mmol of EDC was added to the above
mixed solution. The resultant mixture was packed with aluminum
foil, stirred for 1 h, centrifuged and further dispersed in 100
.mu.L of DMSO. The mixed solution of 15 .mu.L of 2 mg/mL Erbitux
and 185 .mu.L of 1.times.PBS was added to 100 .mu.L of
Ag.sub.2S/DMSO mixed solution and the resultant mixture was reacted
in darkness at 4.degree. C. for 12 h, then centrifuged at 400 g for
4 min, and then the supernatant was taken. MDA-MB-468 cells were
added to the mixed solution of 100 .mu.L of the above supernatant
and 100 .mu.L of 1.times.PBS, coloured at 4.degree. C. for 2 h, and
then washed 3 times with 1.times.PBS solution. It can clearly be
seen that the luminescence was given by Ag.sub.2S quantum dots in
cells by exciting with 658 nm laser, using 1100 nm filter, and
photographing with a 2D InGaAs camera.
Example 5
[0034] 0.1 mmol of silver dihydrocarbyldithiophosphate, 10 g
eicosanethiol and 4 g of hexadecylamine were mixed in a
three-necked flask and heated to 230.degree. C. under an Ar
atmosphere for 0.5 h. After the solution was cooled naturally to
room temperature, 50 mL of anhydrous ethanol was added. The
resultant mixture was centrifuged, washed and dispersed in
cyclohexane. 0.1 g of cysteamine was added to the above cyclohexane
dispersion, then equal volume of anhydrous ethanol was added. The
resultant mixture was stirred for 24 h, then centrifuged and washed
with deionized water to obtain water-soluble Ag.sub.2S quantum dots
with a particle size of about 5 nm, which still have very strong
fluorescence emission. 0.25 mg of the above Ag.sub.2S quantum dots
was dispersed in 100 .mu.L of dimethyl sulfoxide (DMSO), and 50
.mu.L of DMSO solution containing 0.01 mmol of NHS was mixed with
the above solution. Then 50 .mu.L of DMSO solution containing 0.01
mmol of EDC was added to the above mixed solution. The resultant
mixture was packed with aluminum foil, stirred for 1 h, centrifuged
and further dispersed in 100 .mu.L of DMSO. The mixed solution of
15 .mu.L of 2 mg/mL Erbitux and 185 .mu.L of 1.times.PBS was added
to 100 .mu.L of Ag.sub.2S/DMSO mixed solution, and the resultant
mixture was reacted in darkness at 4.degree. C. for 12 h, then
centrifuged at 400 g for 4 min, and the supernatant was taken.
MDA-MB-468 cells were added to the mixed solution of 100 .mu.L of
the above supernatant and 100 .mu.L of 1.times.PBS, coloured at
4.degree. C. for 2 h, and then washed 3 times with 1.times.PBS
solution. It can clearly be seen that the luminescence was given by
Ag.sub.2S quantum dots in cells by exciting with 658 nm laser,
using 1100 nm filter, and photographing with a 2D InGaAs
camera.
Example 6
[0035] 0.1 mmol of silver dodecanoate, 8 g of octanethiol and 4 g
of dodecylamine were mixed in a three-necked flask and heated to
200.degree. C. under an Ar atmosphere for 0.5 h. After the solution
was cooled naturally to room temperature, 50 mL of anhydrous
ethanol was added. The resultant mixture was centrifuged, washed
and dispersed in cyclohexane. 0.12 g of mercaptoacetic acid was
added to the above cyclohexane, then equal volume of anhydrous
ethanol was added, stirred for 24 h, then the resultant mixture was
centrifuged and washed with deionized water to obtain water-soluble
Ag.sub.2S quantum dots with a particle size of about 5 nm, which
still have very strong fluorescence emission. 0.25 mg of the above
Ag.sub.2S quantum dots was dispersed in 100 .mu.L of dimethyl
sulfoxide (DMSO), and 50 .mu.L of DMSO solution containing 0.01
mmol of NHS was mixed with the above solution. Then 50 .mu.L of
DMSO solution containing 0.01 mmol of EDC was added to the above
mixed solution, and the resultant mixture was packed with aluminum
foil, stirred for 1 h, centrifuged and further dispersed in 100
.mu.L of DMSO. The mixed solution of 15 .mu.L of 2 mg/mL Erbitux
and 185 .mu.L of 1.times.PBS was added to 100 .mu.L of
Ag.sub.2S/DMSO mixed solution, and the resultant mixture was
reacted in darkness at 4.degree. C. for 12 h, and then centrifuged
at 400 g for 4 min, and the supernatant was taken. MDA-MB-468 cells
were added to the mixed solution of 100 .mu.L of the above
supernatant and 100 .mu.L of 1.times.PBS, coloured at 4.degree. C.
for 2 h, and then washed 3 times with 1.times.PBS solution. It can
clearly be seen that the luminescence was given by Ag.sub.2S
quantum dots in cells by exciting with 658 nm laser, using 1100 nm
filter, and photographing with a 2D InGaAs camera.
[0036] In conclusion, the method of the invention has moderate
reaction conditions, simple operation, short production cycle, good
reproducibility, and is easy to control. The as-prepared Ag.sub.2S
quantum dots have high fluorescence yield, good fluorescence
stability, excellent biocompatibility and homogeneous sizes, and
can well be used for cellular imaging and in vivo animal tissue
imaging. Furthermore, the method of the present invention is easy
to be implemented in large scale, thus is applicable for the
industrial production.
[0037] The above examples are only the representative ones of
numerous examples of the invention, and do not limit the protection
scope of the invention at all. All the technical solutions having
the equivalent variations or equivalent substitutions fall within
the protection scope of the invention.
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