JPH0841099A - Monoclonal antibody against chlamydia pneumoniae, its production, its antibody-forming cell and method for detecting chlamydia pneumoniae - Google Patents

Abstract

PURPOSE:To obtain a monoclonal antibody useful for various diagnosing methods for specifically detecting Chlamydia pneumoniae to be specifically reacted with a major outer membrane protein (MOMP) derived from Chlamydia pneumoniae much contained in terms of amount or the MOMP, various reagents, a medicine and a source for antibody gene. CONSTITUTION:This monoclonal antibody has specific reactivity to MOMP. This monoclonal antibody-forming cell is capable of producing the monoclonal antibody. This method for producing the monoclonal antibody comprises culturing the antibody-forming cell. This method for detecting Chlamydia pneumoniae uses the monoclonal antibody.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a monoclonal antibody capable of specifically detecting Chlamydia pneumoniae, which has conventionally been difficult to specifically detect, a method for producing the same, an antibody-producing cell thereof, and a method for detecting Chlamydia pneumoniae.

[0002]

2. Description of the Related Art Four types of Chlamydia are known, namely Chlamydia trachomatis, Chlamydia pneumoniae, Chlamydia sittasi and Chlamydia pecolum. Chlamydia pneumoniae is clinically considered to be the causative agent of pneumonia, and Cho-chou Kuo et al.
First reported in 1986 (J. Clinical Microbiology,
24 (6), 1034-1037 (1986)). Currently, the detection of this microorganism is
Although it is performed by the fluorescent antibody method at the laboratory level, it is not suitable for processing a large amount of specimens. Chlamydia
Lipopolysaccharide (LPS) and protein are known as antigens of Pneumoniae. Lipopolysaccharide has an antigenicity common to the genus Chlamydia. Further, as the protein, those having a molecular weight of 39.5 K daltons, 60 K daltons, 75 K daltons, 98 K daltons, etc. are known. Abbreviated as MOMP).

The production of monoclonal antibodies was carried out in 1975 by Ko
Numerous reports have been made since reported by hler and Milstein. GLXA as an antigen specific to Chlamydia
(a genus-specific glycolipid antigen), lipopolysaccharide,
MOMP is known. Elizabeth S. Stuart et al. (Current Microbiolog
y, 28, 85-90 (1994)), a monoclonal antibody against lipopolysaccharide was added to Harland D. Caldwell AND Penny J. Hitchcock.
(Infection and Immunity, 44 (2), 306-314 (1984)),
Monoclonal antibody against MOMP Ellena M. Pete
rson et al. (Infection and Immunity, 59 (11), 4147-4153
(1991)), Byron E. Batteiger et al. (Infection and Immu
nity, 53 (3), 530-533 (1986)).

Monoclonal antibodies against Chlamydia trachomatis have been reported by Richard S. Stephens et al. (J. I.
After mmunology, 128 (3), 1083-1089 (1982), many reports have been made. H. Puy et al. (Immunology Letters, 23,
217-222 (1989/1990)), a monoclonal antibody against Chlamydia pecolum was reported by Kuroda et al. (Am J Vet Res, 54
(5), 709-712 (1993)). Iijima et al. Have reported various monoclonal antibodies against Chlamydia pneumoniae (J. Clinica).
l Microbiology, 32 (3), 58
3-588 (1994)), and there is no report of a monoclonal antibody that specifically reacts with MOMP. Cho-
Chou Kuo et al. reported a monoclonal antibody (RR402) having reactivity specific to Chlamydia pneumoniae (J. Clinical Microbiology, 24 (6), 1034).
-1037 (1986), Tokuyo Sho 64-500083), the antigen recognized by this monoclonal antibody (RR402) has not been clarified (Y. Iijima, J. Clinical Microbiology, 32 (3),
583-588 (1994)).

[0005]

[Problems to be Solved by the Invention] In order to specifically detect Chlamydia pneumoniae, a large amount is contained,
Furthermore, it is desirable to detect an antigen specific to Chlamydia pneumoniae. M as an antigen that satisfies this condition
OMP is possible. However, it is difficult to produce a monoclonal antibody against MOMP derived from Chlamydia pneumoniae only by using conventional means, and there is no monoclonal antibody that specifically recognizes MOMP derived from Chlamydia pneumoniae to date. Therefore, we earnestly studied to prepare a monoclonal antibody that specifically reacts with Chlamydia pneumoniae-derived MOMP, and completed the present invention.

[0006]

[Means for Solving the Problems] That is, the present invention provides a major outer membrane protein (MOMP) derived from Chlamydia pneumoniae.
A monoclonal antibody having a specific reactivity to
The present invention relates to a method for producing the same, a monoclonal antibody-producing cell capable of producing the antibody, and a method for detecting Chlamydia pneumoniae characterized by using the antibody. The monoclonal antibody-producing cell of the present invention is not particularly limited as long as it can produce a monoclonal antibody having specific reactivity with Chlamydia pneumoniae-derived MOMP, but in general, animal immunity, cell fusion, and fused cells , Which is a fused cell (hybridoma) prepared through the steps of selection of cells, selection of specific antibody-producing cells, cloning, etc., and produces a monoclonal antibody having specific reactivity with Chlamydia pneumoniae-derived outer membrane major protein. It is a thing.

Chlamydia pneumoniae used as an immunizing antigen can use an appropriate strain. For example, YK-41 strain, TWAR strain and the like can be mentioned. Examples of mammals to be immunized include mice, rats, rabbits, sheep, chickens, etc., with mice being particularly preferred. The immunization method can be appropriately performed by combining known appropriate methods. In order to efficiently produce the monoclonal antibody-producing cells of the present invention, for example, first, immunization is induced with a basic body of Chlamydia pneumoniae (hereinafter abbreviated as EB) or its native MOMP, and the final immunization is performed. A method using MOMP modified by treatment with a reducing agent and a surfactant as an antigen is preferable. As a reducing agent,
There is no particular limitation, but 2-mercaptolethanol is preferable. The surfactant is not particularly limited, but SDS
(Sodium dodecylbenzene sulfonate) is preferred. In addition, it is preferable to use an adjuvant during immunization, for example, Freund's complete adjuvant (hereinafter referred to as F
Preferred examples thereof include CA) and Freund's incomplete adjuvant (hereinafter abbreviated as FIA). After sufficient immunization by the above method, spleen cells, lymphocyte B cells and the like are isolated from the immunized mammal as parent strains having antibody-producing ability.

Myeloma cells are generally used as the other parent strain for producing hybridomas.
For example, mouse-derived P3U1, NS-1, 653, S
Rat-derived AG1, such as P2, X63, and MPC-11,
Human-derived SK such as AG2, AG3, RCY3 and 210
O-007 and the like can be used, but those derived from mouse are preferable, and P3U1, 653 and the like are particularly preferable. For cell fusion, a method using polyethylene glycol, a method using Sendai virus, an electrofusion method, or the like can be used, but a method using polyethylene glycol is preferable. Hybridomas can be selected by culturing in a selection medium in which only hybridomas can grow. For example H
Preferred examples include AT (hypoxanthine-aminopterin-thymidine) medium and HAz (hypoxanthine-azaserine) medium.

The selection of the specific antibody-producing cells is carried out by collecting the supernatant of the well in which the growth of the hybridoma is observed in the above culture and examining the presence or absence of antibody production by the immunoblotting method using Chlamydia pneumoniae EB, MOMP and the like. It can be done by What is cloning?
The specific antibody-producing cells are to be a uniform cell population derived from one clone, and examples thereof include a limiting dilution method, a method for picking up a colony on soft agar, a single cell manipulation method, a FACS method, and the like. However, the limiting dilution method is simple and preferable. By the above method,
The monoclonal antibody-producing cells of the present invention can be obtained. Hybridoma CP-6, which is an example of the monoclonal antibody-producing cells of the present invention, was deposited at the Institute of Biotechnology, Institute of Industrial Science and Technology, and deposited on July 20, 1994.
Biology Research Institute No. 14436 (FERM P-1443
6) Accepted as

The monoclonal antibody can be produced by culturing the monoclonal antibody-producing cells obtained by the above method in a suitable medium. Preferred examples of the medium include Dulbecco's modified Eagle medium (MEM medium) supplemented with fetal bovine serum.
Monoclonal antibodies can be collected by collecting culture supernatants, affinity chromatography using protein A column, protein G column, etc., ion exchange chromatography, polyethylene glycol fractionation, ethanol fractionation, ammonium sulfate fractionation, etc. Can be done by. Among these, the affinity chromatography method is preferable, and the method using the protein A column is particularly preferable. It is possible to evaluate whether or not the obtained monoclonal antibody has a specific reactivity with Chlamydia pneumoniae-derived MOMP. For example, Chlamydia pneumoniae EB may be treated with an appropriate treatment solution, electrophoresed, and Western blotting using the same may be used to select one that specifically reacts with the MOMP band having a molecular weight of about 39.5 KDa. it can. It can be judged that the monoclonal antibody thus selected specifically recognizes the primary structure (ie, amino acid sequence) of MOMP derived from Chlamydia pneumoniae as an antigenic determinant.

The thus obtained monoclonal antibody of the present invention comprises chlamydia of other species of the genus Chlamydia.
Those that have substantially no reactivity with the major protein of Trachomatis outer membrane are preferable, and those that have substantially no reactivity with all major proteins of the outer membrane of Chlamydia pneumoniae other than Chlamydia pneumoniae are It is preferable because it is useful for species-specific diagnosis of major proteins. The type of monoclonal antibody of the present invention includes any type (globulin class) currently known. In addition, a partially decomposed product from a monoclonal antibody or one having a partial structure of a monoclonal antibody is also included.

The monoclonal antibody of the present invention can be used in a method for detecting Chlamydia pneumoniae in combination with other monoclonal antibodies or polyclonal antibodies, if necessary. Specifically, it can be used in a detection method utilizing an immunoreaction such as a radioimmunoassay and an enzyme immunoassay. Further, it can be used as a component of a diagnostic agent or a research reagent therefor. Furthermore, it can be used in medicine directly or as a chimeric antibody or a drafted antibody. Also,
The monoclonal antibody of the present invention can also be used in a diagnostic kit in which polymerase chain reaction and the like are combined. The antibody-producing cells of the present invention can also be used as a source of antibody genes. Furthermore, this antibody gene can also be expressed in various cells.

[0013]

【Example】

1. Preparation of Chlamydia pneumoniae EB YK-41 strain was used for the preparation of Chlamydia pneumoniae EB. Infection of human lung-derived cells (abbreviated as HL cells) with the YK-41 strain is performed by the method of Kishimoto et al.
(7), 959-964 (1990)). Infected cells cultured in a 24-well or 6-well dish were detached, the whole medium was collected, and then centrifuged at 6,000 rpm for 30 minutes. The supernatant was removed and the precipitate was collected with a sucrose-phosphate-glutamine solution (7.
5% (w / v) sucrose, 3.8 mM KH ₂ PO ₄ , 7 mM K ₂ HPO ₄ ,
The cells were suspended in 5 mM glutamic acid (pH 7.4), abbreviated as SPG. After crushing the suspension with a homogenizer, at 2,500 rpm
Centrifuged for 10 minutes. The supernatant was removed and the precipitate was suspended in SPG. After repeating this operation 5 times, the supernatant was collected. The recovered solution was mixed with 23% (v / v) urographine (Japan Schering Co.) and 50% (w / v) sucrose at a ratio of 2 volume: 2 volume: 1 volume and centrifuged (8,000 rpm, 1 hour). did. After removing the supernatant, the lower layer (50% (w / v) sucrose layer) is collected and S
Suspend in PG, wash, and centrifuge (10,000 rpm, 30 minutes)
Then, the supernatant was removed by suction, and the precipitate was suspended in SPG.
~ 38% (v / v) Urografin Density Gradient and 1 volume:
The mixture was mixed at a ratio of 4 volumes and centrifuged (8,000 rpm, 1 hour), the intermediate layer was recovered and suspended in SPG. Centrifuge (10,000rp
(m, 30 minutes), the supernatant was removed by suction, and the precipitate was suspended in an appropriate amount of SPG to give an EB solution. Storage was performed at 4 ° C or -70 ° C after dispensing.

2. Preparation of Chlamydia pneumoniae MOMP The EB solution obtained above was separated and purified by disc preparative electrophoresis to prepare MOMP. That is, the obtained EB solution (protein (EB) concentration 727 μg / m
l) 0.5 ml and 0.5 ml double concentration sample buffer (0.31 M Tris-HCl (pH 6.8), 1.6% (w / v) sodium dodecyl sulfate (abbreviated as SDS), 1 mM dithiothreitol, 16% ( v / v) glycerin, 0.005% (w / v) bromophenol blue) were mixed and the protein was solubilized by boiling (100 ° C, 3 minutes), and then the whole amount was disc gel (concentrated gel 4%, separation gel 10%). ) And electrophoresis (apparatus: disk preparative electrophoresis apparatus NA-1800 type, Nippon Eido Co., Ltd., electrophoresis condition 100 V (constant voltage)), and a fixed amount (0.26 ml / fraction) was collected. Perform SDS slab electrophoresis for each fraction (apparatus: electrophoresis apparatus for DNA-PAGE NB-5000 type, Nippon Eido Co., Ltd., electrophoresis condition 100
V (constant voltage)) MOMP with a molecular weight of 39.5K is fraction 3
It was confirmed that the protein was collected in Nos. 3 to 45 (protein concentration 0.21m
g / ml).

3. Production of hybridoma and monoclonal antibody 0.5 ml of physiological saline solution of EB obtained by the above method
Was mixed with 0.6 ml of FCA (Difco), and a water-in-oil type emulsion was prepared using an ultrasonic crusher (Branson) as an immunizing antigen, and BALB / c female mice of 8 weeks old 5
Each mouse was intraperitoneally injected with 200 μl (EB immunization amount of 10 μg / mouse each time). After 6 days and 12 days, immunization was performed using FIA (Difco). Six days later, a partial blood sample was taken and the antibody titer of each serum was determined by Dot Immuno Binding Assay.
Method (hereinafter abbreviated as DIBA method), the antibody with the highest antibody titer was purified by the above method as the final immunization for 3 days from the measurement day.
P immunity amount of 1 μg / mouse) was administered through the tail vein. On the day after the third immunization, the spleen of the mouse was dissected out after cervical dislocation and dispersed as single cells to obtain splenocytes for cell fusion. 10% The partner cells for cell fusion (v / v) were pre-cultured in fetal bovine serum pressurized Dulbecco MEM medium (abbreviated as 10F medium) (Napco manufactured CO ₂ incubator, 37 ° C., 5% carbon dioxide gas, saturated humidity ), And P3U1 mouse myeloma cells were used.

1.1 × 10 ⁸ spleen cells and 2.1 × 10 ⁷ P3U1 mouse myeloma cells were mixed and centrifuged (1,00).
After 10 minutes (0 rpm, 10 minutes), the supernatant was removed by suction, and the cell pellet was loosened by shaking gently to give 0.2 ml of a cell fusion agent (50% (v / v) polyethylene glycol; MW = 4000 Merck).
added. After 1 minute and 45 seconds, add 10 ml of culture solution over 1 minute,
Further, 20 ml of the culture solution was added and suspended, followed by stepwise centrifugation and separation (300 rpm 3 minutes, 500 rpm 3 minutes, 700 rpm 3 minutes). The supernatant was removed by suction, and 10F medium (abbreviated as 10F + Bri medium) containing 5% Blai clone (Dainippon Pharmaceutical Co., Ltd.) was added to the cell pellet to prepare a cell suspension. Dispense the cell suspension into 96-well plastic flat-bottomed multiplates (Corning) at 4 × 10 ⁴ cells / well / 0.1 ml in terms of myeloma cells and incubate at 37 ° C, 5% carbon dioxide gas, and saturated humidity. did. The next day, 10F medium containing HAT (1 × 10 ^-4 M hypoxanthine, 4 × 10 ^-7 M aminopterin, 1.6 × 10 ^-5 M
0.1 ml / well of Dulbecco's MEM medium supplemented with thymidine and 10% (v / v) fetal bovine serum, abbreviated as HAT medium) was added, and half of the culture supernatant was replaced with HAT medium at intervals of 3 or 4 days. HAT selection was carried out for about 2 weeks, and the supernatant of the well in which the hybridoma was observed was collected, and the Dot Immuno Binding Assay method (hereinafter,
The presence or absence of antibody production against EB was screened by the DIBA method).

The DIBA method uses a 96-well U-bottom multi-plate (Corning Co.), and the dot size is set to 0.1 per dot in advance.
3mm square membrane filter (Advantech 47mm diameter filter with filter A0) with 02μg EB as antigen
45b047A) with 10% (v / v) FCS (Fetal Calf Serum) at room temperature for 15 minutes, and after removing the supernatant, add 100 μl of the hybridoma culture supernatant to room temperature for 1 hour, then remove the supernatant and label with peroxidase. 100 μl of a 500-fold diluted solution of rabbit anti-mouse immunoglobulin antibody (Cappel) was added, and they were similarly reacted for 1 hour. Finally, 4-chloro-1naphthol was added as a substrate to develop color. Those in which the color was visually recognized on the membrane filter on which the antigen was immobilized were determined to be positive for antibody production. As a result of this screening, hybridomas in wells positive for antibody production were further cloned by the limiting dilution method. That is, a suspension of the antibody-producing positive hybridoma in a HAT-containing medium was prepared, the cell concentration was measured, and the diluted solution was dispensed and cultured in a new microplate so that the hybridoma was 2 to 100 cells / 100 μl / well. . At this time, the diluted solution contains HT + 5%
Bly clone medium was used.

When hybridoma growth is observed in wells of high dilution, antibody production is measured and observed under a microscope to confirm that it is a monoclone, and the same operation is repeated again to confirm that the antibody with high antibody continuous production ability is high. An EB monoclonal antibody producing clone was obtained. Each of the obtained clones was gradually increased in culture scale and finally cultured in 50 ml of 10F medium, and the properties of the antibody were examined in the culture supernatant. The above-mentioned experimental operation was repeated twice before and after, and finally 10 monoclonal antibody-producing clones shown in Table 1 were obtained. The globulin class of the monoclonal antibody produced by 10 hybridoma clones is ISOTYPE Ab-STAT-I kit (Sang Stat M
edical) (see Table 1). As shown in Table 1, monoclonal antibodies CP-1, CP-2, CP-
4, CP-5, CP-6, CP-7, CP-8 did not show reactivity to Chlamydia trachomatis, but showed reactivity to Chlamydia pneumoniae.

[0019]

[Table 1]

Furthermore, 10 F per hybridoma clone producing each of the monoclonal antibodies shown in Table 1.
Pristane (Wako Pure Chemical Industries) that grows cells cultured in a medium
The monoclonal antibody of the present invention was purified from the ascites obtained 2-3 weeks after intraperitoneal inoculation of BALB / c mice pre-treated with 0.5 ml / mouse by inoculation of 1 × 10 ⁷ cells each. Hybridoma clone No. Ascites of 6 is protein A as follows
Was purified by a method using an immobilized affinity column. That is, 1.5 M glycine +3 equivalent to mouse ascites
After adding MNaCl (pH 8.9) and passing through a 0.45 μm nylon filter (Corning Co.), it was applied to an affinity column of Protein A-Poros (NGK) and washed with 1.5 M glycine + 3 M NaCl (pH 8.9). Then 150mM
Elution was performed with a phosphate-citrate buffer solution (pH 6.0) containing NaCl. The eluate was neutralized with 1 M Tris-HCl buffer (pH 9.0), cooled, dialyzed overnight with PBS, and the absorbance of the recovered solution at 280 nm was measured (U-2000, Hitachi, Ltd.) to give 0.2 μm cellulose. Pass through an acetate filter (Corning) -80 ° C
And used as a stock solution of the purified antibody.

4. Specificity analysis of monoclonal antibody The specificity analysis of the monoclonal antibody was performed by Western blotting. Reactivity with Chlamydia pneumoniae is 1. The EB solution (EB concentration 727 μg / ml) obtained by the method described in 1. was used. Specifically, the EB solution containing 50 μg of EB and a sample treatment solution (0.063M Tris, 5%
(v / v) 2-mercaptoethanol, 10% (v / v) glycerin, 2.3% (w / v) SDS, 0.001% (w / v) BPB (Bromphenol Blu
e) (pH 6.8)) was mixed with 1 volume: 1 volume, boiled for 10 minutes, added with 100 μl, and electrophoresed (electrophoresis device: AE-656).
0 (Ato Co.), gel: Atopagel 1020DL, running buffer: 0.025M Tris, 0.192M glycine, 0.1% (w /
v) SDS (pH 8.3), electrophoresis conditions: 20 mA constant current, 80 minutes electrophoresis). As the transfer device, NA-1510 (Japan Eido Co., Ltd.) was used. Gel after migration in transfer buffer (0.025M Tris, 0.192M glycine, 20% (v / v) methanol (pH 8.3)) pre-soaked in ice, 3 mm thick filter paper (Whatman) and Nitrocellulose membrane (Bio-Rad)
Was immersed in a transfer buffer for 15 minutes. The filter paper, the gel, the nitrocellulose membrane, and the filter paper were placed in this order on the pad, and the pad was set on the scissors device. Place the device on ice,
A constant voltage of 50 V was applied for 2 hours to transfer the protein to the nitrocellulose membrane. Nitrocellulose membrane in PBS
10% after washing with (Nissui Pharmaceutical) (2 minutes, shaking twice)
Blocking with FCS / PBS (30 minutes, shaking at room temperature) was performed. The nitrocellulose membrane was washed with PBS (Nissui Pharmaceutical Co., Ltd.) (2 minutes, 3 times, shaken), and then reacted (1 hour, room temperature, shake) with the culture supernatants of all the hybridomas shown in Table 1. After washing with PBS (2 minutes, 3 times, shaking), it was reacted with a 500-fold diluted peroxidase-labeled rabbit anti-mouse immunoglobulin antibody (Kappel) (1 hour, room temperature, shaking). After washing with PBS (shaking for 2 minutes, 3 times), coloring solution (3 mg / ml 4-chloro-
1-naphthol 1ml, PBS 5ml hydrogen peroxide water 2μ
l) was added and reacted until a protein band appeared, and then the membrane was washed several times with distilled water to stop the reaction. For analysis of reactivity with Chlamydia trachomatis, purified Chlamydia trachomatis EB (L2 strain) was used, and Western blotting was performed under the same conditions as above. As a result, the monoclonal antibodies CP-6 and CP-7 of the present invention did not show reactivity with MOMP of Chlamydia trachomatis, but only with MOMP of Chlamydia pneumoniae.

5. Analysis of antibodies by fluorescent antibody method (reactivity with Trachomatis, Sittasi, Pecolum) Chlamydia trachomatis, Sittasi, and Pecolum isolates were placed on a slide glass on which 1 μl of each hybridoma shown in Table 1 was cultured. 8 μl of the supernatant was placed, placed in a wet box, reacted at 37 ° C. for 3 hours, washed with PBS and distilled water, and air-dried. After air-drying, place 8 μl of 10-fold diluted FITC-labeled anti-mouse immunoglobulin / goat serum (KPL) so that all the spotted sites are covered, put in a wet box, react at 37 ° C for 30 minutes, wash, air-dry and mount liquid. (50% glycerin-PBS) 10
The cells were sealed with 0 μl and observed with a fluorescence microscope. As a result, it was found that the monoclonal antibodies CP-6 and CP-7 of the present invention brilliantly appeared only with Chlamydia pneumoniae EB, and the antibody reacted only with Chlamydia pneumoniae EB. The hybridoma producing this monoclonal antibody CP-6 was deposited as CP-6 at the Institute of Biotechnology, Institute of Biotechnology (Deposit No.
36 (FERM P-14436)).

[0023]

INDUSTRIAL APPLICABILITY The monoclonal antibody against Chlamydia pneumoniae of the present invention contains a large amount of its M.
Since it specifically reacts with OMP, it is useful as various diagnostic methods and various reagents for specifically detecting Chlamydia pneumoniae or MOMP thereof, and also as a drug. The antibody-producing cells of the present invention are also useful as a source of antibody genes in addition to producing the above-mentioned antibodies.

─────────────────────────────────────────────────── ───

[Procedure amendment]

[Document name] Consignment number change notification

[Submission date] July 20, 1995

[Former name of depositary institution] Institute of Biotechnology, Institute of Biotechnology, AIST

[Old consignment number] FERM P-14436

[Name of new depositary institution] Institute of Biotechnology, Industrial Technology Institute

[New contract number] FERM BP-5155

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁶ Identification code Internal reference number FI Technical display location G01N 33/569 F 33/577 B // C12N 15/02 (C12P 21/08 C12R 1:91) (72) Inventor Kiyotaka Kawagoe 4-13-1, Higashimachi, Hitachi, Ibaraki Hitachi Chemical Co., Ltd. Pharmaceutical Research Laboratories (72) Inventor Akifumi Iguchi 4-13-1, Higashimachi, Hitachi, Ibaraki Hitachi Chemical Co., Ltd. Company Pharmaceutical Research Institute (72) Inventor Toshihide Morikawa 4-13-1, Higashimachi, Hitachi City, Ibaraki Hitachi Chemical Co., Ltd. Pharmaceutical Research Institute (72) Inventor Yoshiki Nakao 4-13-1, Higashimachi, Ibaraki Prefecture Yamazaki Within Industry Co., Ltd.

Claims (7)

[Claims] 1. A monoclonal antibody having specific reactivity with a major protein of the outer membrane derived from Chlamydia pneumoniae. 2. The monoclonal antibody according to claim 1, which has substantially no reactivity with a major outer membrane protein derived from Chlamydia trachomatis. 3. A monoclonal antibody-producing cell having the ability to produce the monoclonal antibody according to claim 1 or 2. 4. The monoclonal antibody-producing cell according to claim 3, which is a hybridoma of a mouse-derived antibody-producing cell and a mouse myeloma cell. 5. The monoclonal antibody-producing cell according to claim 4, which is a hybridoma CP-6 (Life Science Research Institute No. 14436). 6. A method for producing a monoclonal antibody, which comprises culturing the monoclonal antibody-producing cell according to claim 3, 4, or 5. 7. A method for detecting Chlamydia pneumoniae, which comprises using the monoclonal antibody according to claim 1 or 2.