U.S. patent application number 10/553660 was filed with the patent office on 2006-09-21 for test specimen and production thereof.
This patent application is currently assigned to Canon Kabushiki Kaisha. Invention is credited to Hiroyuki Hashimoto, Manabu Komatsu.
Application Number | 20060211106 10/553660 |
Document ID | / |
Family ID | 34712971 |
Filed Date | 2006-09-21 |
United States Patent
Application |
20060211106 |
Kind Code |
A1 |
Komatsu; Manabu ; et
al. |
September 21, 2006 |
Test specimen and production thereof
Abstract
A test specimen is provided which has one or more chemical
substances fixed to prescribed plural independent positions on a
substrate, and the quantities of the chemical substances fixed at
the respective prescribed positions are the total of integer
multiples of existence quantity units defined for the respective
chemical substances in the range from 1 amol to 1 nmol (excluding
the case in which the total quantity is zero).
Inventors: |
Komatsu; Manabu; (Tokyo,
JP) ; Hashimoto; Hiroyuki; (Tokyo, JP) |
Correspondence
Address: |
FITZPATRICK CELLA HARPER & SCINTO
30 ROCKEFELLER PLAZA
NEW YORK
NY
10112
US
|
Assignee: |
Canon Kabushiki Kaisha
3-30-2, Shimomaruko,
Tokyo
JP
|
Family ID: |
34712971 |
Appl. No.: |
10/553660 |
Filed: |
December 22, 2004 |
PCT Filed: |
December 22, 2004 |
PCT NO: |
PCT/JP04/19716 |
371 Date: |
October 17, 2005 |
Current U.S.
Class: |
435/287.1 ;
436/86 |
Current CPC
Class: |
Y10T 436/11 20150115;
H01J 49/40 20130101; H01J 49/10 20130101; Y10T 436/2575 20150115;
Y10T 436/112499 20150115; Y10T 436/25 20150115; B01L 3/5085
20130101 |
Class at
Publication: |
435/287.1 ;
436/086 |
International
Class: |
C12M 1/34 20060101
C12M001/34; G01N 33/00 20060101 G01N033/00 |
Foreign Application Data
Date |
Code |
Application Number |
Dec 22, 2003 |
JP |
2003-424994 |
Sep 24, 2004 |
JP |
2004-277678 |
Claims
1. A test specimen having one or more chemical substances fixed to
prescribed plural independent positions on a substrate, wherein the
quantities of the chemical substances existing in the respective
prescribed positions are totals of integer multiples of existence
quantity units defined for the respective chemical substances.
2. The test specimen according to claim 1, wherein at least one
kind of the fixed chemical substances is applied onto the substrate
by an inkjet system.
3. The test specimen according to claim 2, wherein all kinds of the
fixed chemical substances are applied onto the substrate by an
inkjet system.
4. The test specimen according to claim 2, wherein the quantity of
the chemical substance applied onto the prescribed positions by the
inkjet system is controlled by a number of liquid droplets which
contain the chemical substance and ejected by the inkjet
system.
5. The test specimen according to claim 4, wherein one liquid
droplet has a volume of not more than 50 pL.
6. The test specimen according to claim 1, wherein the prescribed
positions are arranged in a matrix, and are different in the
existing ratios of the chemical substance.
7. The test specimen according to claim 1, wherein the chemical
substance is selected from the group consisting of metals, metal
compounds, semiconductor materials, organic compounds of a
number-average molecular weight of not more than 10,000, biological
substances, metal ions, metal complexes, halogen ions, and
substances having solubility of 1 ppb or more in water or an
organic solvent at an ordinary temperature and pressure.
8. The test specimen according to claim 7, wherein the metal, the
metal compound, or the semiconductor material is applied in a state
of a fine particle of a diameter of not larger than 1 .mu.m.
9. The test specimen according to claim 1, wherein the test
specimen is used as a standard sample for quantitative
analysis.
10. The test specimen according to claim 9, wherein the
quantitative analysis is conducted by time-of-flight secondary ion
mass spectrometry (TOF-SIMS).
11. A screening method, wherein a test object is applied by inkjet
system onto the chemical substance fixed to the prescribed
positions on the test specimen set forth in claim 1, and a reaction
is detected.
12. The screening method according to claim 11, wherein the test
object contains a biological substance or a medical substance.
13. The screening method according to claim 11, wherein the
reaction is detected by time-of-flight secondary ion mass
spectrometry (TOF-SIMS).
Description
TECHNICAL FIELD
[0001] The present invention relates to a test specimen having a
chemical substance fixed at plural positions on a substrate, and to
a screening method employing the test specimen.
BACKGROUND ART
[0002] With development of film formation technique in recent
years, various materials and devices are coming to be constructed
mainly of a thin film of 1 .mu.m or thinner. Lately, high-speed
film-formation techniques have been developed, which enables
formation of a thin film structure by holding plural functional
films on a substrate as a fine part of high functionality such as
electronic devices and bio-chips. Further, thin film structures
having a sensor function have become important which detect a micro
quantity of a chemical reaction product by utilizing plural
components in the thin film.
[0003] With such improvement of the function of thin film parts,
methods are being developed for more precise and finer analysis and
evaluation of the thin films. The examples of the methods are:
(1) Direct measurement of electroconductivity, hardness, optical
properties, X-ray reactivity, and ionic reactivity for measurement
of functions of a thin film;
(2) Indirect analysis of the components of a thin film by
fractionation of a thin film component by gas chromatography,
high-speed liquid chromatography, ICP-MS analysis, or a like
method;
(3) Marker insertion to an objective substance in the thin film,
such as addition of a fluorescent functional substance, or
substitution by an isotopic element;
[0004] and combinations thereof. In particular, precise and
accurate analysis of the film components is indispensable since the
functions of the thin film are affected delicately by the component
ratios. The extreme small thickness of the thin film tends to cause
a problem of dependence of the functions of the thin film on the
state of the substrate for the thin film, and a problem of adverse
effect of a contamination with a foreign matter or change of the
quality or quantity of the thin film by pretreatment. The
dependence of the function of the thin film on the film component
ratio is investigated frequently by formation of thin film samples
constituted of various component concentration ratios, direct
measurement of the function as mentioned in the above method (1),
and preparation of a calibration curve regarding the dependency of
the obtained signal intensity on the component concentration ratio.
For preparation of more accurate calibration curve, the components
should be uniformly distributed in the thin film sample as well as
precise control of the component concentration ratio in the
samples.
[0005] P. Lazzeri et al. (Surface and Interface Analysis, Vol. 29,
798 (2000)) describes formation of a thin film by spin coating and
analysis thereof with a time-of-flight secondary ion mass
spectrometer (hereinafter referred to as a "TOF-SIMS"). In this
method, the size of one thin film is several millimeters square.
This size is about ten thousand times the size of thin films used
currently in devices in which the size of the one thin film is
being decreased to tens of micrometers square. Such a large
difference in the size causes difference between the practical thin
films and the thin films for calibration samples owing to local
flocculation and mixing state of the respective components, and
other conditions. Therefore, ideally, the one entire thin film is
to be measured and analyzed in one time. However, one measurement
region of the TOF-SIMS is as small as several hundreds of
micrometers square, which cannot cover the entire thin film at one
time. Therefore, the measurement sectional regions are introduced
successively into a measurement chamber for the measurement. In
such a measurement process, during the waiting time for the
measurement, the component ratio is liable to vary by adhesion of
moisture or an impurity from the environmental atmosphere to the
measurement regions or evaporation of the sample component from the
measurement regions.
[0006] (1) Direct measurement of electrical conductivity, hardness,
optical properties, X-ray reactivity, and ion reactivity as the
thin film functionality.
[0007] Energy dispersive fluorescent X-ray analysis is capable of
simultaneous measurement of Na and heavier elements by use of a
fluorescent X-ray. The fluorescent X-ray intensities are
proportional in first approximation to the concentrations of the
respective elements, but are affected greatly by ratio of
coexisting component elements by absorption and secondary
excitation effect thereof. Therefore, in the fluorescent X-ray
analysis also, the standard specimens for the calibration should be
prepared by strict control of the component mixing ratio for the
quantitative determination of the film components and evaluation of
the functionality.
[0008] U.S. Pat. No. 5,365,563 evaluates influence of component
mixing ratio in the thin film by calculation means in quantitative
determination by fluorescent X-ray measurement. In this method,
however, precise calculation is difficult since the fluorescent
X-ray intensity is not necessarily in a linear relation to the
component ratio. Further in this method, the samples should be
prepared in a number corresponding to the number of the film
components, which requires finally test specimens of high accuracy
for the quantitative determination.
[0009] By the above reasons, precise quantitative determination is
not practicable by any of the conventional methods. Therefore, in
many analysis methods including ionic analysis and fluorescent
X-ray analysis, standard specimens should be prepared with accurate
and precise control of the component concentration ratio.
DISCLOSURE OF THE INVENTION
[0010] According to an aspect of the present invention, there is
provided a test specimen having one or more chemical substances
fixed to prescribed plural independent positions on a substrate,
wherein the quantities of the chemical substances existing in the
respective prescribed positions are totals of integer multiples of
existence quantity units defined for the respective chemical
substances.
[0011] At least one kind of the fixed chemical substances is
preferably applied onto the substrate by an inkjet system. All
kinds of the fixed chemical substances are more preferably applied
onto the substrate by an inkjet system.
[0012] The quantity of the chemical substance applied onto the
prescribed positions by the inkjet system is preferably controlled
by a number of liquid droplets which contain the chemical substance
and ejected by the inkjet system. One liquid droplet has preferably
a volume of not more than 50 pL.
[0013] The prescribed positions are preferably arranged in a
matrix, and are different in the existing ratios of the chemical
substance.
[0014] The chemical substance is preferably selected from the group
consisting of metals, metal compounds, semiconductor materials,
organic compounds of a number-average molecular weight of not more
than 10,000, biological substances, metal ions, metal complexes,
halogen ions, and substances having solubility of 1 ppb or more in
water or an organic solvent at an ordinary temperature and
pressure. The metal, the metal compound, or the semiconductor
material is preferably applied in a state of a fine particle of a
diameter of not larger than 1 .mu.m.
[0015] The test specimen is preferably used as a standard sample
for quantitative analysis. The quantitative analysis is more
preferably conducted by time-of-flight secondary ion mass
spectrometry (TOF-SIMS).
[0016] According to another aspect of the present invention, there
is provided a screening method, wherein a test object is applied by
inkjet system onto the chemical substance fixed to the prescribed
positions on the above test specimen, and a reaction is
detected.
[0017] The test object preferably contains a biological substance
or a medical substance.
[0018] The reaction is preferably detected by time-of-flight
secondary ion mass spectrometry (TOF-SIMS).
EFFECTS OF THE INVENTION
[0019] In the present invention, a test specimen is prepared
quickly which contains components in varying ratio in a thin film
in fine regions placed on a substrate for observing influences of
slight change of constituting component ratio and contamination of
impurity. With this test specimen, precise quantitative
determination can be conducted. As shown later in Example 1, an
effective calibration curve can be formed for the measurement in
which a slight change of component like impurity contamination may
cause change in the signal intensity. Further as shown later in
Example 2, an effective calibration curve can be formed for the
quantity of a chemical reaction product of plural mixture
components.
BRIEF DESCRIPTION OF THE DRAWINGS
[0020] FIG. 1 illustrates schematically a planar arrangement of
constitution elements in a quantitative determination specimen of
the present invention.
[0021] FIGS. 2A and 2B are graphs showing relations of a measured
secondary ionic strength to a quantity of a substance in a dot in a
quantitative determination specimen.
[0022] FIG. 3 illustrates schematically a planar arrangement of
constitution elements in a quantitative determination specimen of
the present invention.
[0023] FIGS. 4A and 4B are graphs showing relations of measured
secondary ionic strengths of a chemical reaction product and an
unreacted substance to a quantity of a substance in a dot in a
quantitative determination specimen.
[0024] FIGS. 5A, 5B, 5C, 5D, 5E and 5F show measured ion images of
secondary ions of specimens on which peptides have been deposited
in superposition.
BEST MODE FOR CARRYING OUT THE INVENTION
[0025] In the test specimen of the present invention, one or more
chemical substances are immobilized or fixed on each of prescribed
positions on a substrate. The quantity of the chemical substance is
separated by a quantity unit. The quantity unit by which represents
the quantity of the chemical substance existing in the respective
position is called an "existence quantity unit." Thereby, the
quantity of each of the chemical substances at a prescribed
position is indicated by an integer multiple of the existence
quantity unit. For example, in the case where the existence
quantity units of five chemical substances .alpha., .beta.,
.gamma., .delta., and .epsilon. are denoted respectively by a, b,
c, d, and e, and four chemical substances .alpha., .beta., .gamma.,
.delta., and .epsilon. exist at a certain prescribed position, the
sum of the quantities of the existing chemical substances is
c.sub.1a+c.sub.2b+c.sub.3c+c.sub.4d+c.sub.5e=c.sub.1a+c.sub.2b+c.sub.3c+0-
.times.d+c.sub.5e, where c.sub.1, C.sub.2, C.sub.3, C.sub.4, and
c.sub.5 are respectively an integer, and the coefficient C.sub.4
for d is zero because of the absence of the substance .delta..
[0026] When the chemical substance is fixed without volatilization
or diffusion at the prescribed positions, the existence quantity
unit is occasionally called a "fixation quantity unit."
[0027] The application and fixation of the chemical substance to a
substrate is conducted suitably by an inkjet system typified by a
bubble jet system. A region containing at least one kind of
chemical substance can be formed by applying and fixing the
chemical substance onto a substrate by an inkjet system
(hereinafter the region is occasionally called a "fixation
region"). Incidentally, a solution of chemical substance .gamma.,
for instance, is referred to a ".gamma. solution."
[0028] For precise control of the fixation quantity of chemical
substances, the respective chemical substances are preferably
applied separately and independently onto a substrate by an inkjet
system. The inkjet system enables application of a necessary number
of fine liquid droplets containing the chemical substance onto an
intended spot. The application of plural liquid droplets of a
chemical substance solution to mix the plural liquid droplets is
called "superposed dotting" or "dotting in superposition" in this
Specification. The volume of one liquid droplet applied by the
inkjet system is preferably not more than 50 pL. Here the existence
quantity unit is preferably defined by the quantity of the chemical
substance contained in the one liquid droplet of the inkjet or an
integer multiple thereof. Otherwise, the existence quantity unit
may be defined to bring the integer multiple of the existence
quantity unit within a suitable range in the calibration.
[0029] The test specimen of the present invention is constituted by
controlled application of the chemical substance by an inkjet
system. Therefore a precise calibration curve for quantitative
analysis can be obtained by use of this test specimen.
[0030] For quantitative analysis by use of the test specimen of the
present invention as a standard sample, the plural positions of the
chemical substance fixation are preferably arranged in a matrix.
The matrix arrangement is suitable for changing the fixation
quantity of the chemical substance for the quantitative analysis.
The fixation positions of the chemical substance are arranged in a
matrix of X lines and Y columns. The matrix may be divided into
submatrix elements. One submatrix is called a "block." The
prescribed position constituting the matrix of the present
invention is called an "element position." Of course, one block may
consists of one element position. The composition having been
applied to the element position and containing the chemical
substance is called a "spot." The operation of registering the
ejection head to the element position and applying the liquid
droplets onto the element position by ejection is called
"spotting." The spot formed by one liquid droplet is called a
"dot." One liquid droplet traveling in the air for formation of a
spot is also called a "dot" occasionally. The dot of a .gamma.
solution is called a ".gamma. dot." Two or more dots may be put in
superposition to form one spot. The operation of ejection or
non-ejection of a droplet of a solution from an ejection head onto
an element position is called an "application operation." The
application operation includes stop of the head at an element
position not to conduct spotting to the element position on the
basis of the determination of applying no dot to the element
position. One scanning cycle is completed by application operation
of all the solution ejection heads respectively on every element
position of the XY matrix.
[0031] The chemical substance in the present invention includes
metals, metal compounds, semiconductor materials, organic compounds
of a number-average molecular weight of not more than 10,000,
biological substances, metal ions, metal complexes, halogen ions,
and substances having solubility of 1 ppb or more in water or an
organic solvent at an ordinary temperature and pressure. A metal,
metal compound, or semiconductor material, on application onto a
substrate, is preferably in state of a fine particles of not larger
than 1 .mu.m in diameter on the substrate.
[0032] The test specimen of the present invention is useful
suitably as a standard specimen in quantitative analysis by
time-of-flight secondary ion mass spectrometry (TOF-SIMS). A
primary use of the test specimen is a standard specimen for
quantitative analysis by time-of-flight secondary ion mass
spectrometry (TOF-SIMS) and like analysis methods.
[0033] The analysis methods by use of the test specimen of the
present invention as a standard specimen include fluorescent X-ray
analysis, optical response analysis, and electrical conductivity
analysis in addition to the TOF-SIMS.
[0034] In quantitative analysis by TOF-SIMS, the dose quantity of
primary ions is kept constant at a level of not higher than
1.times.10.sup.13/cm.sup.2, and the integrated intensity (count
number) of specified secondary ions emitted from a prescribed area
is measured.
[0035] A secondary use of the test specimen of the present
invention is use for various screening. In the screening, a third
chemical substance is applied by an inkjet system onto the chemical
substance fixed on plural points on the test specimen and the
resulting chemical reaction is used for the screening. The third
chemical substance is preferably a biological substance or a
medical substance. The test is preferably conducted by
time-of-flight secondary ion mass spectrometry (TOF-SIMS).
[0036] When the objective chemical substance is a water-soluble
metal complex, a material disclosed in Japanese Patent Application
Laid-Open No. 2000-251665, for instance, can be used as it is. Such
a material is preferably applied by a bubble jet system. The test
specimen of the present invention prepared by applying a chemical
substance solution onto a substrate by an inkjet system may be
heat-treated, if necessary, after the application. The dotting in
superposition by the inkjet system may be conducted with a driving
system described in Japanese Patent Application Laid-Open No.
H04-361055.
[0037] Before application of the test chemical substance onto the
substrate, the substrate surface may be treated for fixation of the
chemical substance. This treatment may be conducted, for instance,
by the method described in Japanese Patent Application Laid-Open
No. H11-187900. This method is preferably employed when the
chemical substance to be applied is an organic compound having an
SH group.
[0038] According to the present invention, a test specimen can be
prepared for precise evaluation of the dependency of the
performance of a thin film on a slight difference in the components
ratio in the film, the film thickness, and the kind of the
substrate. This is one of the features of the present
invention.
[0039] For instance, the intensity of signals according to
ionization is affected greatly by the state of a specimen.
Therefore, dependence of a noticed function on the state of the
specimen can be evaluated by using signals obtained from the
ionization. For this evaluation, the test specimen of the present
invention and preparation method thereof will be effective.
[0040] A generalized embodiment of the present invention is
explained below. For simplicity of explanation, two kinds of
chemical substances, .alpha. and .beta., are used to prepare a test
specimen.
[0041] Solutions of the respective chemical substances are
prepared, and are stored respectively in printer head tanks.
[0042] In this embodiment, the element positions are arranged in a
matrix having X lines and Y columns. This XY matrix is divided into
submatrixes having respectively m lines and n columns, where
m<n. One submatrix is called a "block." The block on the i-th
line on the j-th column of the XY matrix is represented by
B.sub.ij. For simplicity of explanation, all the blocks are assumed
to be constituted of elements in x lines and y columns. Therefore,
x.m=X and y.n=Y.
[0043] All the spots fixed at element positions in one block are
made to have the same composition within that block.
[0044] One droplet of an a solution ejected from a printer head is
assumed to contain a chemical substance .alpha. in an amount of
"a," and one droplet of a .beta. solution ejected from a printer
head is assumed to contain a chemical substance .beta. in an amount
of "b."
[0045] In one scanning cycle of the spotting operation, the spots
may be formed on all the element positions in one block, and
thereafter sequentially in the next blocks (this process being
called a "sequence process." Otherwise, the spots may be formed on
a specific element position in all of the blocks and then this
application operation is repeated by changing the specific
positions at all of the positions on all of the blocks (this
process being called a "correspondence process").
[0046] Naturally, the heads may be provided in a number
corresponding to the number of the lines or columns of the blocks
to conduct the spotting operation on the respective lines or
columns simultaneously.
[0047] In the first scanning cycle, .alpha. dots are put at all the
element positions in all of the blocks except the blocks on the
first line, and .beta. dots are put at all the element positions in
all of the blocks except the blocks on the first column.
[0048] In the second scanning cycle, .alpha. dots are put in
superposition at all the element positions in all of the blocks
except the blocks on the first and second lines, and .beta. dots
are put in superposition at all the element positions in all of the
blocks except the blocks of the first and second columns.
[0049] Similarly, in the i-th scanning cycle, .alpha. dots are put
in superposition at all the element positions in all of the blocks
except the blocks of the first to i-th lines, and .beta. dots are
put in superposition at all the element positions in all of the
blocks except the blocks of the first to i-th columns.
[0050] In the m-th scanning cycle, since there is no (m+1)th line,
the dotting of the a solution is not conducted, and .beta. dots are
put in superposition at all the element positions in all of the
blocks except the blocks of the first to m-th columns. Thus, in the
m-th scanning cycle and later scanning cycles, the .alpha. solution
is not applied.
[0051] In the (n-1)th scanning cycle, the a solution is not put,
and .beta. dots are put in superposition at all the element
positions in the blocks except the 1 to (n-1)th columns, namely in
the n-th column.
[0052] In the n-th scanning cycle, since there is no columns except
the 1 to n columns, the .beta. solution is not applied, and the
scanning is completed.
[0053] After the above scanning cycles, no a dot is formed in the
blocks on the first line, and no .alpha. dot is formed in the
blocks on the first column: there is no spot in the block
B.sub.11.
[0054] In the block B.sub.ij, the spot at one element position is
formed by (i-1) .alpha. dots, and (j-1) .beta. dots. Therefore, the
quantity of the chemical substance a in that spot is a(i-1), and
the quantity of the chemical substance .beta. is b(j-1). Since each
of the blocks has element positions in x lines and y columns, the
block B.sub.ij contains totally the chemical substance .alpha. in a
quantity of a(i-1)xy and the chemical substance .beta. in a
quantity of b(j-1)xy. For instance, in the block B.sub.45, the
number of the a dots is 4-1=3, and the number of the .beta. dots is
5-1=4 for formation of the one spot on the respective element
positions: the spot contains the chemical substance .alpha. in a
quantity of 3a and the chemical substance .beta. in a quantity of
4b. In the entire block B.sub.45, the chemical substance .alpha. is
contained in a quantity of 3axy, and the chemical substance .beta.
is contained in a quantity of 4bxy. In the above method, the
concentration ratios are changed only by an integer ratio between
the blocks. Dots having any existence ratio can be formed by
providing plural aqueous solutions having different concentrations
for standard specimen preparation.
[0055] A specimen having continuous condition change in the entire
XY matrix can be prepared by varying the element positional
conditions and subdividing the blocks, and conducting the spot
formation by the aforementioned correspondence method to give
conditional gradient in the one block.
EXAMPLES
[0056] The present invention is described below in more detail by
reference to Examples. The examples below show best modes for
carrying out the invention, but do not limit the invention
thereto.
Example 1
[0057] In analysis of components of a biological material by SIMS
or fluorescent X-ray analysis, a trace amount of sodium (Na) or
potassium (K) as a contaminant may affect the intensity of the
signals. In this Example, a calibration curve was obtained for
quantitative determination of phosphorus (P) by TOF-SIMS by using a
standard sample of ammonium phosphate (NH.sub.4H.sub.2PO.sub.4,
ammonium dihydrogenphosphate) containing Na and K as an example of
quantitative determination of a biological material by using the
method of the present invention.
(1) Substrate Cleaning
[0058] A silicon substrate (high-resistance p-type, commercial
product) having a size of 10 mm.times.12 mm.times.1 mm was
subjected to supersonic cleaning in high-purity acetone, ethanol,
and ultrapure water respectively for 10 minutes.
(2) Preparation of Aqueous Solutions of Component Mixtures
[0059] Aqueous standard solutions for IPC-MS (SPEX Co.) containing
respectively P (10.1%), Na (10.1%), or K (5.0%) were diluted
respectively with pure water to 100 .mu.M to prepare aqueous
solutions for standard sample preparation (hereinafter referred to
as a "P solution," an "Na solution, and a "K solution"
respectively). Two standard specimens were prepared: a matrix
having elements composed of mixtures of the P solution and the Na
solution as the elements, and a matrix having elements composed of
mixtures of the P solution and the K solution (hereinafter the
former is referred to as an "Na matrix," and the latter is referred
to as a "K matrix"). The spot formation procedure in this Example
is shown below specifically for formation of the Na matrix from the
P dots and the Na dots. For formation of the K matrix, the Na
solution is replaced by the K solution in the Na matrix formation
procedure.
(3) Application of Solutions for Preparation of Quantitative
Determination Specimen by Inkjet System
[0060] The printer employed was a bubble jet printer (BJF-950:
Canon K.K.) of a bubble jet type, a kind of a thermal jet type. The
above-prepared aqueous standard solutions were placed respectively
in a several hundred microliter portion in the three tanks of the
printer head of the printer. The volume of the one liquid droplet
of the respective solutions ejected from the printer heads was 4
pL/droplet. The dot formed by one liquid droplet on the element
position had a diameter of about 50 .mu.m. The spots were formed by
dotting in superposition. The content of P in one ejected droplet
is 2.4.times.10.sup.8 atoms and the content of Na therein is
2.4.times.10.sup.8 atoms. The content of K in one ejected droplet
regarding K matrix is alos 2.4.times.10.sup.8 atoms. The Na matrix
and the K matrix were formed respectively of 157 lines and 236
columns at a density of 200 dpi, namely 127 .mu.m pitch, in a range
of 20 mm.times.30 mm on the surface of the silicon wafer having
been cleaned in Step (1) above. The two matrixes were placed side
by side as shown in FIG. 1. The Na matrix was divided into blocks
in 10 lines and 10 columns. The reminders of the divisions were
ignored.
[0061] In the first scanning cycle, P dots were applied on all
lines except the first line, and Na dots were applied on all
columns except the first column.
[0062] In the second scanning cycle, P dots were applied on all
lines except the first and second lines in superposition on the
spots having formed in the first scanning cycle, and Na dots were
applied on all columns except the first and second columns in
superposition on the having formed in the first scanning cycle.
[0063] In the later scanning cycles, the P dots and the Na dots
were put in superposition by decreasing one line and one column of
the application in each scanning cycle. That is, in the i-th
scanning cycle, P dots were put in superposition on all the spots
in all of the lines except the first to i-th lines, and Na dots
were put in superposition on all the spots in all of the columns
except the first to i-th columns.
[0064] In the ninth scanning cycle, the P dots were put only on the
tenth lines, and the Na dots were put only on the tenth
columns.
[0065] The pattern was completed by the above nine scanning cycles.
In the block B.sub.ij on the i-th line on the j-th column, the spot
at one element position is formed from (i-1) dots of P, and (j-1)
dots of Na. Therefore, in that spot, the quantity of P is a(i-1),
and the quantity of Na is b(j-1). When the number of the element
position in the block B.sub.ij is n.sub.ij, the total quantity of P
in the block B.sub.ij is a(i-1)n.sub.ij, and the total quantity of
Na in this block is b(j-1)n.sub.ij.
[0066] No dot was put on the first line on the first column in the
block B.sub.11 (at the upper left corner) of the Na matrix shown in
FIG. 1, so that no spot was formed there.
[0067] Although a bubble jet printer was employed in this Example,
the same result will be obtained by use of a piezo type printer or
a like printer.
(4) TOF-SIMS Measurement
[0068] The concentration standard specimen shown in FIG. 1 was
subjected to analysis by means of a time-of-flight secondary ion
mass spectrometer (TOF-SIMSIV: ION-TOF Co.). The irradiation was
conducted to a primary ion injection dose of 1.times.10.sup.12
atoms/cm.sup.2 under the conditions shown in Table 1, the intensity
of P.sup.- of the secondary ions detected during the irradiation
was integrated, and the cumulative intensity was derived for the
mixing ratios of Na or K. TABLE-US-00001 TABLE 1 TOF-SIMS
Measurement Conditions Primary ions Secondary ions Ion species
Ga.sup.+ Ion species C.sup.- Acceleration 25 kV Measurement 300
.times. 300 .mu.m.sup.2 voltage region Pulse 10 kHz Integration 32
times times
[0069] FIGS. 2A and 2B show relations between the intensity of
phosphorus secondary ion and the quantity of existing phosphorus
for each of the spots on the third line (j=4), the fourth line
(j=5), and the fifth line (j=6) for each of the Na and K,
respectively. The phosphorus secondary ion intensity increased
slightly with increase of the mixed Na and K. Thus, the matrix
effects of the impurities or the like can be quantitatively
understood, which enables precise quantitative analysis by
TOF-SIMS.
Example 2
[0070] By the technique of the present invention, an effective
calibration curve can be obtained for the quantity of a product of
a chemical reaction of mixture components. In this Example, a test
specimen for a chemical reaction product was prepared by dropping
an aqueous solution of sodium carbonate (Na.sub.2CO.sub.3) onto a
film of a peptide (Morphiceptin: mass number 521 amu) formed on a
substrate, and was evaluated.
[0071] An aqueous solution of Morphiceptin, a well-known
intracerebral neurotransmitter, and an aqueous solution of sodium
carbonate as a weak acid salt were prepared. Spots were formed by
changing the quantity of the respective components by dotting in
superposition by an inkjet system. The quantity of the chemical
reaction product of the two components in the spot was evaluated by
secondary ion intensity by TOF-SIMS.
(1) Preparation of Specimen
[0072] Morphiceptin and powdery sodium carbonate were dissolved
respectively in water to prepare an aqueous Morphiceptin solution
(1.9.times.10.sup.-4 mol/L) and an aqueous sodium carbonate
solution (2.4.times.10.sup.-4 mol/L). Spots were formed on a
silicon wafer surface as shown in FIG. 3 by a bubble jet printer in
the same manner as in Example 1. In the block B.sub.ij on the i-th
line on the j-th column, the spot at one element position is formed
by (i-1) dots of Morphiceptin, and (j-1) dots of sodium carbonate.
Therefore, in that spot, the quantity of Morphiceptin is a(i-1) and
the quantity of sodium carbonate is b(j-1), where "a" and "b" are
respectively a quantity of Morphiceptin or sodium carbonate
contained in one ejected liquid droplet. When the number of the
element position in the block B.sub.ij is n.sub.ij, the total
quantity of Morphiceptin in the block B.sub.ij is a(i-1)n.sub.ij,
and the total quantity of Na is b(j-1)n.sub.ij in this block.
(2) TOF-SIMS Measurement
[0073] The concentration standard specimen shown in FIG. 3 was
subjected to TOF-SIMS measurement. The intensity of ions formed
from the molecular Morphiceptin (hydrogen atom addition, mass
number 522 amu), and the intensity of ions (mass number: 544 amu)
of the chemical reaction product (Morphiceptin molecule+sodium)
ions were integrated, and the cumulative intensities for the
chemical reaction product was derived for the existence ratios of
the substances.
[0074] Incidentally, the chemical reaction of the Morphiceptin
molecule and sodium carbonate is substitution of the hydrogen atom
of the terminal carboxyl group (COOH) of the Morphiceptin molecule
by sodium.
[0075] FIGS. 4A and 4B show the dependency of the average of the
secondary ionic intensities of the reaction product (Morphiceptin
molecule+sodium) and the unreacted reactant (Morphiceptin
molecule+hydrogen) on the existence quantity of sodium carbonate in
each of the spots at the positions of i=6 (fixed) and j=1, 2, 3, .
. . , 10. As shown in FIGS. 4A and 4B, before the chemical
equilibrium, the secondary ion intensity of the chemical reaction
product in the presence of molecular sodium carbonate increases in
linear proportion, and the secondary ion intensity of the unreacted
reactant decreases in linear proportion.
Example 3
[0076] This Example shows detection of plural parent peptide
molecular moieties by TOF-SIMS.
(1) A silicon wafer substrate was prepared in the similar manner as
in Example 1.
[0077] (2) A first synthetic peptide SEQ ID NO: 1 (GGGGCGGGGG)
(hereinafter referred to as a "peptide G," mass number: 634 amu), a
second synthetic peptide SEQ ID NO: 2 (YYYYCYYYYY) (hereinafter
referred to as a "peptide Y," mass number: 1588 amu), and a powdery
insulin (mass number: 5807 amu) material were dissolved
respectively in 2 mL of water containing a small amount of a
surfactant (0.1 wt %), the solutions having respectively a
concentration of 7.9.times.10.sup.-5 mol/L, 1.1.times.10.sup.-5
mol/L, and 8.2.times.10.sup.-6 mol/L. The solutions are referred to
respectively as a "G solution," a "Y solution," and an "insulin
solution."
[0078] (3) Ten matrixes (having 10 lines and 10 columns), M.sub.1
to M.sub.10 were prepared by use of the G solution and the Y
solution in place of the P solution and the Na solution in Example
1 in the same manner for Na matrix formation as in Example 1. The
existence quantity units of the G solution, the Y solution, and the
insulin solution in one dot are represented respectively by a, b,
and c. In every matrix, in the block B.sub.ij on the i-th line on
the j-th column, the spot at one element position is formed by
(i-1) dots of the peptide G (hereinafter G dots), and (j-1) dots of
the peptide Y (hereinafter Y dots). Therefore, in that spot, the
quantity of the peptide G is a(i-1), and the quantity of the
peptide Y is b(j-1). When the number of the element position in the
block B.sub.ij is n.sub.ij, the total quantity of the peptide G in
the block B.sub.ij is a(i-1)n.sub.ij, and the total quantity of the
peptide Y therein is b(j-1)n.sub.ij. No dot was put on the block
B.sub.11 on the first line on the first column (at the upper left
end) in the respective blocks, so that no spot was formed
there.
(4) Onto the spots in the ten matrixes M.sub.1, . . . , M.sub.k, .
. . , M.sub.10, including the element positions in the respective
blocks B.sub.11, insulin was dotted in superposition.
[0079] On each of the spots in matrix M.sub.k, k-1 dots of insulin
were put in superposition.
[0080] By TOF-SIMS measurement, as shown in FIGS. 5A to 5F, ion
images were obtained as secondary ions: ion images of the G peptide
parent ions, the Y peptide parent ions and Na atom adducts thereof
(FIGS. 5A and 5B), ion images of the Y peptide parent ions, and Na
atom adducts thereof (FIGS. 5C and 5D), and ion images of insulin
fragments ions (mass number: 804 amu, 1198 amu) (FIGS. 5E and 5F).
The quantities of the peptides in the one spot in the ion images
were calculated to be 2 pg of the G peptide, 0.7 pg of the Y
peptide, and 19 pg of the insulin fragments. In FIGS. 5A to 5F,
"mc" and "tc" are abbreviations of "maximum counts" and "total
counts," respectively. By combination of the above result with the
procedure in Example 2 for "quantitative detection of a chemical
reaction product by dropwise addition of aqueous sodium carbonate
onto a peptide film substrate," a test of reactivity of several
tens of picograms of a peptide with a medical substance (screening)
is practicable in principle.
Sequence Listing Free Text
<210> 1
<223> sample for detection by TOF-SIMS
<210> 2
<223> sample for detection by TOF-SIMS
[0081] This application claims priorities from Japanese Patent
Applications No. 2003-424994 filed on Dec. 22, 2003 and No.
2004-277678 filed on Sep. 24, 2004, which are hereby incorporated
by reference herein.
Sequence CWU 1
1
2 1 10 PRT Artificial SAMPLE FOR DETECTION BY TOF-SIMS 1 Gly Gly
Gly Gly Cys Gly Gly Gly Gly Gly 1 5 10 2 10 PRT Artificial SAMPLE
FOR DETECTION BY TOF-SIMS 2 Tyr Tyr Tyr Tyr Cys Tyr Tyr Tyr Tyr Tyr
1 5 10
* * * * *