JP2001264334A - Reagent for measuring syphilis treponemal antibody and method for producing the same - Google Patents

Abstract

(57) [Summary] [Problem] To provide a highly sensitive, constant quality reagent for measuring treponema pallidum antibodies while generating few false positive or false negative samples. SOLUTION: This is a reagent for measuring a syphilis treponemal antibody using a peptide having a syphilis treponemal antigen activity, wherein at least one peptide among the peptides is:
SEQ ID NOS: 1 to 39 in the sequence listing (for example, SEQ ID NO: 1: Ser
Ala Gly Ala Leu Gln Leu Leu Val Val Gly Cys Gly S
er Ser His His Glu Thr His Tyr Gly Tyr Ala), characterized in that it is a peptide containing at least 5 or more consecutive amino acid sequences in at least one peptide selected from 39 peptides. Syphilis Treponema antibody measuring reagent.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

[0001] The present invention relates to a syphilis treponemal antibody assay reagent using a peptide having syphilis treponemal antigen activity, and a method for producing the same.

[0002]

2. Description of the Related Art In the field of clinical tests, tests for Treponema pallidum antibodies using blood have been conducted, and various measuring methods have been developed and used so far. Representative examples of these measurement methods include a biochemical measurement method using an enzyme reaction, an immunoassay method using an antigen-antibody reaction, and the like.

In recent years, it has been desired to accurately measure Treponema pallidum antibodies in blood, and immunoassays using genetically modified antigens have been increasingly used.

[0004] For example, Japanese Patent Application Laid-Open No. 9-229988 discloses a method for detecting an antibody to Treponema pallidum using a Treponema pallidum antigen prepared by genetic recombination.

[0005] It is generally known that a reagent using a recombinant antigen exhibits the same or better reagent performance than a reagent using a natural antigen. However, even with such a reagent using a recombinant antigen, the problem that false positive or false negative samples are generated in the measurement of T. pallidum antibodies cannot be completely solved.

In addition, since syphilis antigens prepared by genetic recombination are purified from microorganisms such as Escherichia coli, the production process is complicated, the quality tends to vary, and the sensitivity of reagents is insufficient. there were.

[0007]

SUMMARY OF THE INVENTION The present invention has been made to solve the above-mentioned problems, and has as its object to reduce the occurrence of false positive or false negative specimens and to increase the sensitivity,
It is an object of the present invention to provide a reagent for measuring T. pallidum antibodies of constant quality.

[0008]

According to the first aspect of the present invention,
A syphilis treponemal antibody measuring reagent using a peptide having syphilis treponemal antigen activity, wherein at least one of the peptides is selected from 39 peptides represented by SEQ ID NOs: 1 to 39 in the sequence listing. A reagent for measuring an antibody to Treponema pallidum, which is a peptide containing at least 5 consecutive amino acid sequences in at least one kind of the obtained peptides.

[0009] The invention according to claim 2 is a reagent for measuring an antibody to Treponema pallidum using a peptide having an activity of Treponema pallidum, wherein at least one of the peptides has SEQ ID NOS: 11 to 52 in the sequence listing. A syphilis treponemal antibody measuring reagent, characterized in that it is a peptide selected from the 42 peptides shown in (1).

The invention according to claim 3 is the reagent for measuring a syphilis treponemal antibody according to claim 1 or 2, wherein the peptide is supported on an insoluble carrier.

[0011] The invention according to claim 4 is the reagent for measuring a syphilis treponemal antibody according to claim 3, wherein at least a part of the peptide is carried on an insoluble carrier via a carrier protein.

According to a fifth aspect of the present invention, there is provided a method for producing a syphilis treponemal antibody measuring reagent according to the fourth aspect, wherein
Syphilis, characterized in that, after the step of carrying the carrier protein on the insoluble carrier, the step of carrying the peptide on the insoluble carrier in the first step is carried out as the second step. This is a method for producing a treponemal antibody measurement reagent.

Hereinafter, the contents of the present invention will be described in detail.

The selection of the amino acid sequence of the peptide used in the present invention was performed as follows.

Of the syphilis treponemal antigens, 47 kDa
For the antigen, 15 kDa antigen and TmpA antigen, the amino acid sequence of a site (epitope) having a known syphilitic treponemal antigen activity was used. 47k
The amino acid sequence of the epitope of the Da antigen is SEQ ID NO: 1
10 (reported by RE Baughn et al. (Thejo
urnal of Immunology 157 (1996) 720-731)), 15 kD
The amino acid sequence of the epitope of the a antigen has SEQ ID NOS: 11 to 11.
23 (reported by RE Baughn et al. (Infec
tion and Immuity 64 (7) (1996) 2457-2466))
The amino acid sequence of the epitope of the mpA antigen is SEQ ID NO: 3.
0 to 33 (G. An
Report of toni et al. (Journal of Immunological Methods 189
(1996) 137-140)). Among the epitopes of the 47 kDa antigen, the amino acid sequences of sites having particularly high antigen activity are shown in SEQ ID NOs: 40 to 48. Among the epitopes of the TmpA antigen, the amino acid sequences of sites having particularly high antigen activity are shown in SEQ ID NO: 30. It is known to be shown (see the above-mentioned documents).

On the other hand, among the syphilis treponemal antigens, 17
The amino acid sequences of the kDa antigen, TmpB antigen and 4D antigen have already been reported (for the 17 kDa antigen, reports by DR Akins et al. (Infection and Immu
64 (4) (1993) 1202-1210) and the TmpB antigen reported by Hansen, EB et al. (J. Bacteriol 162 (3) (1.
985) 1227-1237), for the 4D antigen, Walfield,
Report by AM et al. (Infection and Immuity 57 (2) (1989)
633-635). ) Has not been reported for the amino acid sequence of the site (epitope) having T. pallidum antigenic activity.

Further, glycerophosphodiester phosphodiesterase derived from Treponema pallidum (Glyce
rophosphodiester phosphodiesterase (GlpQ))
Has a molecular weight of about 38 kDa, which is said to be secreted into the outer membrane of the bacterium by the N-terminal 20 amino acids which are a secretion signal.
The enzyme. Cameron, CE et al.
Has shown to act as a vaccine when immunized to rabbits, concluding that it is a highly reactive immunogen (Treponemal antigen syphilis). The amino acid sequence of Treponema pallidum GlpQ antigen was determined by Stebeck, CE et al. (FEMS Microbio
l. Lett. 154 (2) (1997) 303-310) and Shevchenko, DV
(Infection and Immuity 65 (10) (1997) 4179-4189)
However, the amino acid sequence of a site (epitope) having T. pallidum antigenic activity has not been reported.

For this reason, the present inventors have proposed Hopp, TP, W
oods, KR et al. (Proc. Natle. Acid. Sci. USA 1
78 (6) (1981) 3824-3828), the 17 kDa antigen, T
The amino acid sequences of the epitopes of the mpB antigen 4D antigen and the GlpQ antigen were deduced.

The amino acid sequence of the epitope of the 17 kDa antigen is shown in SEQ ID NOS: 24 to 26. Among them, the amino acid sequence of the site having particularly high antigen activity is estimated to be those shown in SEQ ID NOs: 49 to 52. did. Also, T
The amino acid sequence of the eptB of the mpB antigen is SEQ ID NO: 2.
7 to 29, and among them, the amino acid sequence of the site having particularly high antibody activity was estimated to be that shown in SEQ ID NO: 27. The amino acid sequence of the epitope of the 4D antigen is shown in SEQ ID NOs: 34 to 36, and among them, the amino acid sequence of a site having a high antigen activity was estimated to be shown in SEQ ID NO: 34. Furthermore, Gl
The amino acid sequence of the pQ antigen epitope is SEQ ID NO: 37.
To 39, and among them, the amino acid sequence of the site having particularly high antigen activity was estimated to be that shown in SEQ ID NO: 37.

The present invention is a reagent for measuring an antibody to Treponema pallidum using a peptide having at least a part of the amino acid sequence of the above epitope.

At least one of the peptides used in the present invention includes at least one peptide selected from 39 peptides represented by SEQ ID NOs: 1 to 39 in the sequence listing.
It is necessary that at least 5 or more, more preferably 7 or more consecutive amino acid sequences in a species peptide be included. As such a peptide, for example, those derived from the peptide represented by SEQ ID NO: 1 (24 amino acid residues) include five or more consecutive
In addition to the peptide itself obtained by extracting 24 or less amino acids, a peptide in which an arbitrary peptide is bound to one or both ends of these peptides may be used. Further, the peptide used in the present invention may be a peptide containing at least 5 or more consecutive amino acid sequences extracted from each of two or more peptides selected from the 39 peptides shown in the sequence listing, directly or by another peptide. It may be a peptide bound via a peptide.

At least one of the peptides used in the present invention is represented by SEQ ID NO: 11 to 52 in the sequence listing.
More preferably, it is a peptide selected from the 42 peptides represented by

The method for producing the above peptide is not particularly limited, but generally a peptide obtained by solid phase synthesis can be used.

[0024] In the reagent for measuring an antibody to Treponema pallidum of the present invention, the above-mentioned peptide having Treponema pallidum antigen activity is preferably carried on an insoluble carrier.

Examples of the insoluble carrier include organic polymer powders, microorganisms, blood cells and cell membrane fragments. Further, the insoluble carrier may be a plate or the like used for ELISA or the like.

Examples of the organic polymer powder include natural polymer powders such as insoluble agarose, cellulose, and insoluble dextran; polystyrene, styrene-sulfonic acid (salt) copolymer, styrene-methacrylic acid copolymer, acrylonitrile- Synthetic polymer powders such as a butadiene-styrene copolymer, a vinyl chloride-acrylate copolymer, and a vinyl acetate-acrylate copolymer are exemplified. In addition, the same organic polymer as shown here can be used as a material for a plate or the like used for the above-mentioned ELISA or the like.

Among the above insoluble carriers, it is particularly preferable to use a latex in which a synthetic polymer powder is uniformly suspended. In this case, the particle size of the latex particles is 0.05 to
1.5 μm is preferred, and 0.1 to 0.8 μm is more preferred.

Further, as the insoluble carrier, those in which a sulfonic acid group, a carboxyl group, an amino group, a hydroxyl group or the like is introduced on the surface of the carrier can be used as appropriate.

When the peptide is carried on an insoluble carrier, the peptide may be carried directly by physical adsorption or chemical bonding, or may be carried via a carrier protein as described later.

When the peptide is supported by physical adsorption, the peptide can be supported by, for example, adding a suspension of the insoluble carrier to a solution containing the peptide and stirring the solution.

When a peptide is supported by chemical bonding, an insoluble carrier having a sulfonic acid group, a carboxyl group, an amino group, a hydroxyl group or the like introduced on its surface is used, and an appropriate crosslinking agent is added. It can be carried.

The pH of the solution at the time of carrying out the above supporting step is 3
-10 and the temperature is preferably 2-50 ° C.

In the present invention, it is more preferable that at least a part of the peptide is carried on an insoluble carrier via a carrier protein. In this case, for example, after the peptide is chemically bonded to the carrier protein in advance, the carrier protein can be supported on the insoluble carrier by physical adsorption. Alternatively, a suspension of a carrier protein supported on an insoluble carrier is added to a peptide-containing solution and stirred, whereby the peptide is bound to the carrier protein by physical adsorption. Alternatively, the peptide can be obtained by chemically bonding a peptide to a carrier protein by using an amino group or a carboxyl group on the surface of the carrier protein and adding an appropriate crosslinking agent.

Examples of the carrier protein include serum albumin (human, bovine, horse, rabbit, etc.),
Albumins such as ovalbumin and ovalbumin; biological proteins such as ferritin; hemocyanins such as quinhole limpet hemocyanin (KLH); and enzymes such as galactosidase and alkaline phosphatase. Although not limited, it is preferable to use bovine serum albumin (BSA) in terms of availability. Further, a protein and another substance may be bonded, such as a glycoprotein bonded to a sugar or a lipoprotein bonded to a lipid.

The method for producing the syphilis treponemal antibody measuring reagent of the present invention comprises, as a first step, a step of carrying a carrier protein on an insoluble carrier, followed by a step of:
As a step, the production method of carrying out the step of supporting the peptide on the insoluble carrier in which the first step has been performed is more preferable.

In the first step, the carrier protein may be loaded on the insoluble carrier by either physical adsorption or chemical bonding.

When physical adsorption is used, a suspension containing an insoluble carrier is added to a solution containing a carrier protein, and the solution is supported by stirring.

When a chemical bond is used, an insoluble carrier surface into which a sulfonic acid group, a carboxyl group, an amino group, a hydroxyl group, or the like is introduced is used, and a suitable crosslinking agent is reacted. The carrier protein can be supported on an insoluble carrier. The crosslinking agent is not particularly limited, but N-succiimidyl-4- (N-maleimidomethyl) -1-carboxylate (SMCC) or the like can be used.

When carrying the carrier protein by the physical adsorption or the chemical bonding, the pH of the solution is 3 to 1
0, temperature is preferably 2 to 50 ° C. This is because if the pH is out of this range, problems such as denaturation of the carrier protein occur. When the temperature is lower than 2 ° C., the reaction rate is low, and it is difficult to obtain a reagent having a desired sensitivity. When the temperature is higher than 50 ° C., the carrier protein is denatured. It is.

In the second step, the method of loading the peptide on the insoluble carrier in which the first step is performed may be either physical adsorption or chemical bonding.

When the above-mentioned physical adsorption is used, the suspension containing the carrier protein supported on the insoluble carrier in the first step is added to the solution containing the peptide, and the suspension can be supported by stirring. . When using a chemical bond, a functional group such as an amino group, a carboxyl group, a thiol group, a sulfonic acid group, or a hydroxyl group on the surface of the carrier protein or the insoluble carrier is used, and the above-mentioned crosslinking agent is added. Thereby, the peptide can be supported on the carrier protein.

When the peptide is supported by the above physical adsorption or chemical bonding, the pH of the solution is 3 to 10 and the temperature is
2-50C is preferred.

When the syphilis treponemal antibody measurement reagent of the present invention is used, it may be diluted with an appropriate buffer or the like before use.

The buffer includes, for example, phosphate buffer, Tris buffer, glycine buffer, Good buffer, etc., but is not particularly limited, and may be appropriately selected in consideration of the characteristics of the substrate to be used. It may be used.

[Action] The syphilis treponemal antibody assay reagent of the present invention uses a peptide having the above-mentioned syphilis treponemal antigen activity instead of using the recombinant syphilis treponemal antigen. There is no problem and the measurement sensitivity is high and the production reproducibility is excellent.

[0046]

The present invention will be described in more detail with reference to the following examples, but the present invention is not limited to these examples.

Examples 1 to 7 (1) Preparation of Peptide-Sensitized Latex Solution Bovine serum albumin (hereinafter referred to as BSA) was 1% (w
/ V) containing 100 mM phosphate buffer (pH 7.4)
2 mL of polystyrene latex having an average particle size of 0.4 μm (solid content 10% (w / v), manufactured by Sekisui Chemical Co., Ltd.) 10
Added to 0 μL and stirred at 4 ° C. for 1.5 hours. Next, each of the various peptides shown in Table 1 (each obtained by solid-phase synthesis) was added to the phosphate buffer containing the latex at 2 μg / mL (for example, in the case of Example 1, Peptide solution 4 dissolved at a concentration of 9 peptides shown in Nos. 40 to 48 (total 18 μg / mL)
Add 00 μL and stir for 1 hour. This solution was centrifuged at 18000 rpm at 10 ° C. for 30 minutes. The resulting precipitate contains 100% BSA at 0.25% (w / v).
5 mL of an mM phosphate buffer (pH 7.4) was added, and the latex was suspended to prepare a peptide-sensitized latex solution.

[0048]

[Table 1]

(2) Preparation of Sample Dilution Solution BSA was 0.25% (w / v), pGEMA (glycosylethyl methacrylate homopolymer, average molecular weight 1)
10% containing 140,000 (manufactured by Nippon Seika) 1% (w / v)
Sodium ethylenediaminetetraacetate was dissolved in 0 mM phosphate buffer (pH 7.4) to a concentration of 5 mM to prepare a sample diluent.

(3) Treponema pallidum antibody measuring reagent The reagent for measuring treponema pallidum antibody of the present invention is as described in (1) above.
A two-component reagent comprising the first reagent consisting of the peptide-sensitized latex liquid described in the above section and the second reagent consisting of the sample diluent described in (2) above.

(4) Standard substance Treponema pallidum antibody was used in 0.17.67.475T.
U. Human serum containing (titer unit) was used as a standard treponemal antibody solution.

(5) Measurement of Treponema pallidum antibody titer by an automatic analyzer The following shows a method of measuring an anti-treponema pallidum antibody in a sample using an automatic biochemical analyzer "Hitachi 7050" (manufactured by Hitachi, Ltd.). Measurement mode: Original Abs Parameters: Sample volume 20 μL Peptide-sensitized latex solution (first reagent) 50 μL Sample diluent (second reagent) 350 μL Measurement wavelength: 570 nm Measurement time: Immediately after sample dispensing, sample diluent is added and mixed Thereafter, the peptide-sensitized latex solution is added and mixed. After the addition of the peptide-sensitized latex solution, the amount of change in absorption from 80 seconds to 320 seconds was determined, and this was defined as the reaction amount. The pH of the reaction system was 7.4.

[Embodiments 8 to 14] (1) of Embodiments 1 to 7
The procedure was performed in the same manner as in Examples 1 to 7, except that the section of preparation of the peptide-sensitized latex reagent was as follows. (1) Preparation of peptide-sensitized latex solution 2 mL of 100 mM phosphate buffer (pH 7.4) containing 1% (w / v) of BSA was added to a polystyrene latex having an average particle diameter of 0.4 μm (solid content: 10% (w / v)). / V), manufactured by Sekisui Chemical Co., Ltd.) and stirred at 4 ° C. for 1.5 hours. In addition, SMCC (N-succiimidyl-4-
(N-maleimidomethyl) -1-carboxylate) 8
0.4 mL of a mg / mL dimethylformamide solution was added, and the mixture was stirred at 35 ° C. for 1 hour to bind BSA and SMCC. Next, the above solution was applied to a dialysis membrane (UC-32-10).
0: size 20/32, manufactured by Sanko Junyaku Co., Ltd.), and dialyzed with 3 L of physiological saline (0.9% NaCl) all day and night. Next, 2 μg / mL of each of the peptides shown in Table 1 (each obtained by solid-phase synthesis) in a 100 mM phosphate buffer (pH 7.4) was added to the dialyzed solution (for example, in the case of Example 8). Then, SEQ ID NOs: 40 to 48 in the sequence listing
(9 total peptides, total 18 μg / mL) were added, and the mixture was stirred at 4 ° C. for 1 hour. This solution is heated at 10 ° C. for 30 minutes for 180 minutes.
Centrifuge at 00 rpm. 5 mL of 100 mM phosphate buffer (pH 7.4) containing 0.25% (w / v) of BSA was added to the obtained precipitate, and the latex was suspended to prepare a peptide-sensitized latex solution.

[Comparative Examples 1 to 7] Examples 1 to 7 were repeated except that item (1) in Examples 1 to 7 was performed as follows.
The same was done. (1) Preparation of T. pallidum antigen-sensitized latex solution 15 μl in 2 mL of 100 mM phosphate buffer (pH 7.4)
400 μL of the recombinant T. pallidum antigen solution shown in Table 1 dissolved at a protein concentration of g / mL was added, and this solution was further added to polystyrene latex having an average particle size of 0.4 μm (solid content: 10% (w / v)). , Manufactured by Sekisui Chemical Co., Ltd.) and stirred at 4 ° C. for 1.5 hours. Then 100 mM containing 1% (w / v) BSA
2 mL of a phosphate buffer (pH 7.4) was added, and the mixture was stirred for 1.5 hours. This solution is added at 18000 for 30 minutes at 18000.
Centrifuged at rpm. 0.25 BSA was added to the resulting precipitate.
% (W / v) in 100 mM phosphate buffer (pH
7.4) 5 mL was added, and the latex was suspended to prepare a Treponema pallidum antigen-sensitized latex solution.

[Performance Test 1] The syphilis treponemal antibody measurement reagents of Examples 1 to 14 and Comparative Examples 1 to 7 were prepared 5 times (5 lots) each, and 67T. U. The antibody was measured using the standard T. pallidum antibody solution as a specimen. Coefficient of variation from measured value (CV value: standard deviation ÷ average value x
100) was obtained, and this value is shown in Table 2.

[0056]

[Table 2]

As is clear from Table 2, Examples 1 to 14
In the present invention, compared to Comparative Examples 1 to 7, the variation between lots is small, and the production reproducibility is excellent.

[Performance Test 2] Using the syphilis treponemal antibody measurement reagents of Examples 1 to 14 and Comparative Examples 1 to 7, 50 samples that were previously determined to be negative or positive were measured, and the coincidence rate of the determination was tested. The results are shown in Tables 3 to 9. In addition, 10T. U. The above was judged as positive.

[0059]

[Table 3]

[0060]

[Table 4]

[0061]

[Table 5]

[0062]

[Table 6]

[0063]

[Table 7]

[0064]

[Table 8]

[0065]

[Table 9]

As apparent from Tables 3 to 9, Examples 1 to
In the fourteen inventions, the negative / positive judgment results surely coincided with each other, and no false negative / false positive problem occurred as in Comparative Examples 1 to 7.

[0067]

Since the present invention has the above-mentioned constitution, it has excellent reproducibility in production and has less occurrence of false-negative and false-positive antibodies compared to a measurement reagent using Treponema pallidum antigen by genetic modification. Reagents can be provided.

[0068]

[Sequence List] <110> SEKISUI CHEMICAL CO., LTD. <120> Reagent for measuring syphilis treponemal antibody and its production method <130> 00P02223 <160> 52 <210> 1 <211> 24 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 1 Ser Ala Gly Ala Leu Gln Leu Leu Val Val Gly Cys Gly Ser Ser His 1 5 10 15 His Glu Thr His Tyr Gly Tyr Ala 20 <210> 2 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 2 Ala Gly Glu Leu Gly Gln Ser Arg Asp Val Leu Leu Ala Gly Asn Ala 1 5 10 15 Glu Ala Asp Arg 20 <210> 3 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 3 Arg Lys Lys Trp Glu Tyr Glu Thr Asp Pro Ser Val Thr Lys Met Val 1 5 10 15 Arg Ala Ser Ala 20 <210> 4 <211> 32 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 4 Gly Glu Ile Lys Phe Glu Ala Val Glu Gly Ala Val Ala Leu Ala Asp 1 5 10 15 Arg Ala Ser Ser Phe Met Val Asp Ser Glu Glu Tyr Lys Ile Thr Asn 20 25 30 <210> 5 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence : Peptide <400> 5 Glu Asp Ser Arg Val Thr Glu Asn Thr Asn Gly Leu Lys Thr Met Leu 1 5 10 15 Thr Glu Asp Ser 20 <210> 6 <211> 24 <212> PRT <213> Artificial Sequence <220 > <223> Description of Artificial Sequence: Peptide <400> 6 Met Glu Ser Pro His Asp Leu Val Val Asp Thr Val Gly Thr Val Tyr 1 5 10 15 His Ser Arg Phe Gly Ser Asp Ala 20 <210> 7 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 7 Asn Phe Asn Thr Val Arg Tyr Asp Tyr Tyr Gly Asp Asp Ala Ser Tyr 1 5 10 15 Thr Asn Leu Met 20 <210> 8 <211> 40 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 8 Phe Asp Arg Phe Lys Gly Ser Gly Pro Gly Tyr Tyr Arg Leu Thr Leu 1 5 10 15 Ile Ala Asn Gly Tyr Arg Asp Val Val Ala Asp Val Arg Phe Leu Pro 20 25 30 Lys Tyr Glu Gly Asn Ile Asp Ile 35 40 <210> 9 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 9 Ile Gly Gly Ala Asp Ala Glu Thr Leu Met Asp Ala Ala Val Asp Val 1 5 10 15 Phe Ala Asp Gly 20 <210> 10 <211> 40 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 10 Val Ser Asp Gln Ala Val Ser Leu Gly Gln Asn Val Leu Ser Ala Asp 1 5 10 15 Phe Thr Pro Gly Thr Glu Tyr Thr Val Glu Val Arg Phe Lys Glu Phe 20 25 30 Gly Ser Val Arg Ala Lys Val Val 35 40 <210> 11 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 11 Val Met Tyr Ala Ser Ser Gly 1 5 <210> 12 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 12 Asp Tyr His Arg Val Met Tyr 1 5 <210> 13 <211> 8 <212> PRT <213> Artificial Sequence <220 > <223> Description of Artificial Sequence: Peptide <400> 13 Lys Met Val Gln Val Val Tyr Asp 1 5 <210> 14 < 211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 14 Met Val Gln Val Val Tyr Asp Tyr 1 5 <210> 15 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 15 Val Gln Val Val Tyr Asp Tyr Gln 1 5 <210> 16 <211> 8 <212> PRT <213> Artificial Sequence <220 > <223> Description of Artificial Sequence: Peptide <400> 16 Asp Tyr His Arg Val Met Tyr Ala 1 5 <210> 17 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 17 Met Tyr Ala Ser Ser Gly Ile Gly 1 5 <210> 18 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 18 Pro Glu Lys Ala Phe Arg Glu Leu 1 5 <210> 19 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 19 Glu Lys Ala Phe Arg Glu Leu Ala 1 5 <210> 20 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial S equence: Peptide <400> 20 Lys Ala Phe Arg Glu Leu Ala Asp 1 5 <210> 21 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 21 Phe Arg Glu Leu Ala Asp Ala Leu 1 5 <210> 22 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 22 Arg Glu Leu Ala Asp Ala Leu Leu 1 5 <210> 23 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 23 Val Thr Gly Ala Thr Val Ser Ser 1 5 <210> 24 <211> 25 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 24 Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu Lys Val Glu 1 5 10 15 Cys Ala Leu Lys Gly Gly Ile Phe Arg 20 25 <210> 25 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 25 Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro Ser Pro Leu Thr Tyr 1 5 10 15 Arg Gly Thr Trp 20 <210> 26 <211> 16 <212> PRT <213> A rtificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 26 Arg Tyr Met Gly Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met Ala 1 5 10 15 <210> 27 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 27 Gln Glu Glu Arg Lys Arg Ala Glu Glu Glu 1 5 10 <210> 28 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 28 Leu Val Ile Pro Ser Leu Lys Gly Glu Glu Arg Glu Gly Leu Tyr 1 5 10 15 <210> 29 <211> 15 <212> PRT <213 > Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 29 Lys Lys Leu Tyr Glu Ala Asn Lys Asp Lys Ile Pro Gln Ser Lys 1 5 10 15 <210> 30 <211> 19 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 30 Ala Ser Gly Ala Lys Glu Glu Ala Glu Lys Lys Ala Ala Glu Gln Arg 1 5 10 15 Ala Leu Leu <210> 31 <211 > 35 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequen ce: Peptide <400> 31 Glu Ala Glu Lys Ala Leu Gln Ser Ala Lys Thr Lys Gln Lys Ala Ser 1 5 10 15 Ser Asp Leu Ala Arg Ser Ala Asp Lys Ser Ala Pro Leu Pro Glu Asn 20 25 30 Ala Gln Gly 35 <210> 32 <211> 37 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 32 Pro Leu Pro Glu Asn Ala Gln Gly Phe Ser Lys Glu Pro Ile Glu Val 1 5 10 15 Glu Pro Leu Pro Asn Asp Arg Leu Asn Thr Thr Gln Ala Asp Glu Ser 20 25 30 Ala Pro Ile Pro Ile 35 <210> 33 <211> 40 <212> PRT <213> Artificial Sequence <220> <223 > Description of Artificial Sequence: Peptide <400> 33 Asp Glu Ser Ala Pro Ile Pro Ile Ser Asp Thr Ser Ser Pro Ser Arg 1 5 10 15 Val Gln Ser Arg Gly Val Glu Asp Gly Gly Arg Ser Pro Lys Ser Ser 20 25 30 Met Asn Glu Glu Gly Ala Ser Arg 35 40 <210> 34 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 34 Ala Glu Glu Thr Glu Lys Glu Ile Thr Ile 1 5 10 <210> 35 <211> 15 <212> PRT <213> Artificial Sequence <220> <2 23> Description of Artificial Sequence: Peptide <400> 35 Val Ser Ala Leu Ala Arg Val Lys Arg Asp Phe Glu Tyr Leu Ser 1 5 10 15 <210> 36 <211> 20 <212> PRT <213> Artificial Sequence <220 > <223> Description of Artificial Sequence: Peptide <400> 36 Glu Ala Phe Asp Thr Ile Ala Glu Arg Leu Leu Gln Leu Gly Ala Gln 1 5 10 15 Ala Pro Ala Ser 20 <210> 37 <211> 30 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 37 Val Gln Thr Tyr Asp Phe Asn Glu Leu Lys Arg Ile Lys Arg Glu Leu 1 5 10 15 Leu Pro Lys Tyr Glu Met Asn Val Lys Leu Ile Gln Arg Val 20 25 30 <210> 38 <211> 30 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 38 Ala Tyr Thr Asp Gln Arg Glu Thr Gln Glu Lys Asp Ser Arg Gly Lys 1 5 10 15 Trp Ile Asn Tyr Asn Tyr Asn Trp Met Phe Glu Pro Gly Gly 20 25 30 <210> 39 <211> 30 <212> PRT <213> Artificial Sequence <220> < 223> Description of Artificial Sequence: Peptide <400> 39 Asp Val Ala Glu Lys Phe Pro Arg Arg Gln Arg Ala Asp Gly His Phe 1 5 10 15 Tyr Val Ile Asp Phe Thr Val Glu Glu Leu Ser Leu Leu Arg 20 25 30 <210> 40 <211> 12 <212> PRT <213> Artificial Sequence <220> < 223> Description of Artificial Sequence: Peptide <400> 40 Gly Gln Ser Arg Asp Val Leu Leu Ala Gly Asn Ala 1 5 10 <210> 41 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 41 Glu Tyr Glu Thr Asp Pro Ser Val Thr Lys Met Val 1 5 10 <210> 42 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 42 Gly Glu Ile Lys Phe Glu Ala Val Glu Gly Ala Val 1 5 10 <210> 43 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence : Peptide <400> 43 Glu Asp Ser Arg Val Thr Glu Asn Thr Asn Gly Leu 1 5 10 <210> 44 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 44 Met Glu Ser Pro His Asp Leu Val Val Asp Thr Val 1 5 10 <210> 45 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 45 Asn Phe Asn Thr Val Arg Tyr Asp Tyr Tyr Gly Asp 1 5 10 <210> 46 <211> 12 <212> PRT <213 > Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 46 Lys Gly Ser Gly Pro Gly Tyr Tyr Arg Leu Thr Leu 1 5 10 <210> 47 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 47 Ile Gly Gly Ala Asp Ala Glu Thr Leu Met Asp Ala 1 5 10 <210> 48 <211> 12 <212> PRT <213> Artificial Sequence < 220> <223> Description of Artificial Sequence: Peptide <400> 48 Phe Thr Pro Gly Thr Glu Tyr Thr Val Glu Val Arg 1 5 10 <210> 49 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 49 Lys Ala Lys Ala Glu Lys 1 5 <210> 50 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 50 Lys Ala Glu Lys Val Glu 1 5 <210> 51 <211> 6 <212> PRT <213> Artificial S equence <220> <223> Description of Artificial Sequence: Peptide <400> 51 Glu Lys Lys Ser Ala Pro 1 5 <210> 52 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Peptide <400> 52 Gly Lys Pro Ser Lys Glu 1 5

 ──────────────────────────────────────────────────続 き Continuation of the front page (31) Priority claim number Japanese Patent Application No. 11-299790 (32) Priority date October 21, 1999 (Oct. 21, 1999) (33) Priority claim country Japan (JP) (31) Priority claim number Japanese Patent Application No. 11-290969 (32) Priority date October 13, 1999 (October 13, 1999) (33) Priority claim country Japan (JP) (31) Priority claim number Japanese Patent Application No. 11-287233 (32) Priority date October 7, 1999 (Oct. 7, 1999) (33) Priority claiming country Japan (JP) F-term (reference) 4H045 AA30 BA14 BA15 BA16 BA17 BA18 BA19 BA60 CA11 DA86 EA52

Claims (5)

[Claims] 1. A reagent for measuring a syphilitic treponemal antibody using a peptide having a syphilitic treponemal antigen activity, wherein at least one of the peptides is represented by SEQ ID NOS: 1 to 39 in the sequence listing. A syphilis treponemal antibody measurement reagent, which is a peptide containing at least five consecutive amino acid sequences in at least one kind of peptide selected from species of peptides. 2. A reagent for measuring a syphilitic treponemal antibody using a peptide having a syphilitic treponemal antigen activity, wherein at least one of the peptides is represented by SEQ ID NOS: 11 to 52 in the sequence listing. A reagent for measuring an antibody to Treponema pallidum, which is a peptide selected from species of peptides. 3. The reagent for measuring an antibody to Treponema pallidum according to claim 1, wherein the peptide is carried on an insoluble carrier. 4. The reagent for measuring a syphilis treponemal antibody according to claim 3, wherein at least a part of the peptide is supported on an insoluble carrier via a carrier protein. 5. The method for producing a syphilis treponemal antibody measurement reagent according to claim 4, wherein the first step comprises a step of supporting a carrier protein on an insoluble carrier, and the second step comprises: And carrying out the step of allowing the insoluble carrier subjected to the first step to carry out the method.