JP3772267B2 - Method for analyzing allergen-specific IgE - Google Patents

Description

[0001]
BACKGROUND OF THE INVENTION
The present invention relates to a method for analyzing allergen-specific immunoglobulin E (IgE). The “analysis” in the present specification includes “measurement” for quantitatively or semi-quantitatively determining the amount of allergen-specific IgE to be analyzed, and the presence or absence of allergen-specific IgE to be analyzed. Both “detection” and “determination” are included.
[0002]
[Prior art]
Immunoglobulin E (IgE) is an antibody molecule having a molecular weight of about 190,000 discovered by Ishizaka et al. In 1966 and is known to be deeply involved in type I allergy. As typical diseases of type I allergy, for example, atopic dermatitis, allergic rhinitis, bronchial asthma, or urticaria can be mentioned. In these diseases, serum IgE concentration (total IgE concentration) has a certain medical condition. It is known to reflect.
[0003]
On the other hand, the antibody molecule has a high specific binding property to an antigen as a characteristic of the antibody molecule, and this reactivity is used to identify a disease-causing antigen (allergen). Classical methods include skin tests (eg, scratch test, prick test, intradermal test, or Prausnitz-Kustner test). In this method, an allergen solution is put into a patient's skin through a wound or intradermal injection scratched with a needle, and a causative antigen is estimated based on the degree of biological reaction such as redness or wheal. In addition, a provocation test or the like for observing the degree of a disease state by inhaling or ingesting a component that seems to be an allergen has been conventionally performed. Both of these tests are observing the patient's own biological response, are not only painful for the patient, but are often risky if not performed under good management.
[0004]
In order to avoid such inconvenience, in recent years, a method of measuring allergen-specific IgE amount in a test tube such as CAP-RAST (Pharmacia) or ALASTAT (DPC) using antigen-antibody reaction ( In vitro testing) has been widely performed.
That is, IgE expressed in vivo is reacted with various allergens (for example, allergens such as tick, cedar, dust, or milk), and then the anti-IgE antibody labeled with an enzyme or radioactive substance and the complex And the amount of allergen-specific IgE is measured by detecting a signal derived from the labeling substance.
[0005]
In addition, as a measurement system that performs multi-item allergen search simultaneously, there is a so-called MAST method (MAST). For example, in a commercially available product using the MAST method (for example, MAST26; MAST), a carrier with stretched cellulosic yarn in which 26 different allergens are separately solid-phased is placed in a dedicated reaction container. The corresponding specific IgE in can be measured simultaneously.
[0006]
By the way, typical diseases of type I allergic diseases involving specific IgE include, for example, atopic dermatitis or bronchial asthma. Many of these cases have a childhood age from the first few months of life, and inhalable antigens such as house dust, dust mites, or molds, or dietary antigens such as egg white, milk, or wheat are reported as allergens. Yes. In order to avoid the seriousness and / or lengthening of symptoms in patients of the age group who do not have sufficient ability to transmit subjective symptoms, it is extremely therapeutically important to identify allergens and restrict intake early. Yes, the measurement of allergen-specific IgE concentration in serum by in vitro test is considered to have clinical significance. However, since conventional in vitro tests require a minimum of tens to hundreds of μL of serum, it is difficult to collect such large samples from children (especially infants) and often rely on in vivo tests. The situation was not possible.
[0007]
[Problems to be solved by the invention]
Accordingly, an object of the present invention is to solve the above-mentioned disadvantages of the prior art, and to simultaneously measure the amount of allergen-specific IgE in a small amount of a test sample (for example, serum or dried filter paper blood) for a plurality of allergens. Another object of the present invention is to provide a method for analyzing (particularly measuring) with high sensitivity.
[0008]
[Means for Solving the Problems]
Said subject is according to the invention,
(1) A step of binding different allergens for each color tone to plural kinds of colored beads having a diameter of 2 to 5 mm having different color tones (hereinafter referred to as an allergen binding step);
(2) A step of bringing the allergen-binding colored beads obtained in the step (1) into contact with a test sample that may contain each immunoglobulin E that specifically reacts with each of the allergens (hereinafter referred to as the following) , Referred to as test sample contact step);
(3) a step of isolating the allergen-binding colored beads for each color tone (hereinafter referred to as an isolation step);
(4) A step of bringing each allergen-binding colored bead isolated in the step (3) into contact with an anti-immunoglobulin E antibody labeled with a labeling substance (hereinafter referred to as a labeled secondary antibody contact step). And (5) each signal derived from a labeling substance contained in each immune complex comprising each allergen-binding colored bead, immunoglobulin E specifically reacting with the allergen, and labeled anti-immunoglobulin E antibody; Alternatively, a step of analyzing each signal derived from a labeled substance contained in an unreacted labeled anti-immunoglobulin E antibody not contained in each of the immune complexes (hereinafter referred to as an analysis step).
It can be solved by a method for analyzing allergen-specific immunoglobulin E, which comprises
The present invention also provides:
(1) A step of binding different kinds of allergens for each color tone to a plurality of types of colored beads having a diameter of 2 to 5 mm having different color tones;
(2) contacting the allergen-binding colored beads obtained in the step (1) with a test sample that may contain each immunoglobulin E that specifically reacts with each of the allergens;
(3) a step of bringing the mixture obtained in the step (2) into contact with the anti-immunoglobulin E antibody labeled with a labeling substance;
(4) a step of isolating the allergen-binding colored beads for each color tone; and (5) each allergen-binding colored bead isolated in the step (4) and immunoglobulin E specifically reacting with the allergen. And a method for analyzing allergen-specific immunoglobulin E, comprising the step of analyzing each signal derived from a labeled substance contained in each immune complex comprising a labeled anti-immunoglobulin E antibody. .
Furthermore, the present invention provides
(1) a plurality of colored beads having a diameter of 2 to 5 mm having different color tones and carrying different allergens on the surface for each color tone;
(2) An immunoglobulin E analysis kit that specifically reacts only with any one of the aforementioned allergens, comprising an anti-immunoglobulin E antibody labeled with a labeling substance.
[0009]
DETAILED DESCRIPTION OF THE INVENTION
Hereinafter, the present invention will be described in detail.
The analysis method of the present invention includes (1) an allergen binding step, (2) a test sample contact step, (3) an isolation step, (4) a labeled secondary antibody contact step, and (5) an analysis step. In the analysis method of the present invention, the steps (1) to (5) can be carried out in this order, or the order of the steps (3) and (4) is reversed, and the allergen binding step and the test sample contact are performed. It can also be carried out in the order of the step, the labeled secondary antibody contact step, the isolation step, and the analysis step.
In the analysis method of the present invention, a plurality of colored beads on which allergens are solid-phased are reacted with a test sample in a single reaction tube, and then divided into individual reaction tanks according to the color of the colored beads. By using a detection system using an anti-human IgE antibody labeled with a labeling substance, the allergen-specific IgE antibody titer can be simultaneously measured. As a result, it is possible to reduce the amount of the test sample, and it is possible to measure serum, blood, filter paper blood, or the like, which has conventionally been difficult to collect a sufficient amount from an infant.
[0010]
Coloring of beads In the analysis method of the present invention, a plurality of kinds of colored beads having different color tones are used. Various colored beads are commercially available, and these commercially available beads can be used in the analysis method of the present invention.
Size of the beads, Ru diameter 2mm~5mm der. If the particle size is smaller than this, the dispensing operation may be complicated. On the other hand, if the particle size is larger than this, a large amount of sample is required, and it is not preferable from the viewpoint of reaction efficiency.
As the material, either an inorganic substance or an organic substance can be used as long as it does not cause a disadvantage in the reaction system, and is not particularly limited, but is preferably a polymer substance such as polystyrene beads or Examples thereof include polypropylene and glass.
[0011]
The colored beads can be prepared by a conventionally known method instead of using a commercially available product. For example, when polystyrene beads [Immunobeads A26 (diameter = 3.2 mm); Immunochemical Co., Ltd.] are used as beads, for example, an appropriate dye [for example, methyl yellow (Tokyo Chemical Industry), Solvent Blue] 35 (Solvent Blue 35; Aldrich Chemical Company), Oil Orange SS (Tokyo Chemical Industry), or Oil Red EGN (Oil Red EGN; Aldrich Chemical Company) etc.] dissolved in a 50% aqueous solution of acetone. It can be prepared by stirring with purified water and then washing with purified water. As the pigment, an oil-soluble dye is easy to use, but a water-soluble dye can also be used.
[0012]
Allergen solid phase to beads The method for binding each allergen to colored beads is not particularly limited, and may be carried out by a conventionally known means such as a so-called physical adsorption or chemical binding method. it can.
For example, in the case of physical adsorption, colored beads are stirred in an aqueous solution obtained by diluting each allergen extract to an appropriate concentration, washed with a buffer solution, and dried to prepare allergen-bound colored beads. can do.
In addition, as a chemical bonding method, for example, biotin is solid-phased on the surface of a bead in advance, and an allergen to which biotin is bonded is bonded through avidin, or an appropriate functional group (for example, carboxyl Examples thereof include a method in which beads having undergone functional group modification such as a group, an amino group, or a sulfhydryl group are bonded to an allergen via a crosslinking agent (for example, carbodiimide or N-hydroxysuccinimide).
[0013]
Allergen The allergen that can be used in the analysis method of the present invention is not particularly limited. For example, house dust, dust mite, food, pollen, fungus, insect, parasite, chemical substance, or Conventionally known allergens such as drugs can be targeted. As each allergen, a commercially available product [for example, available from GREER Co., Ltd.] can be used, or a desired allergen can be prepared. For example, the antigenic material as a raw material is directly or after pulverization, and in some cases, after degreasing with an organic solvent (for example, ether, acetone, hexane, etc.), a buffer solution [for example, phosphate buffer solution Allergens can be prepared by stirring and extracting in (PBS) etc. and diluting the centrifugal supernatant to an appropriate protein concentration.
[0014]
Test sample The test sample that can be analyzed by the analysis method of the present invention is not particularly limited as long as it is a sample that may contain IgE specifically reacting with the allergen. Examples thereof include biological fluids generally used for clinical diagnosis, such as serum, blood, or filter paper blood.
[0015]
Anti-IgE antibody and labeling substance As a labeling substance used for labeling the anti-IgE antibody, a known labeling substance, for example, an enzyme (for example, alkaline phosphatase, β-galactosidase, peroxidase, or luciferase), A fluorescent substance (for example, europium), a light-emitting substance (for example, an acridinium derivative or adamantane), a radioactive substance (for example, ¹²⁵ I), and the like can be given. Further, according to the labeling substance, for example, a substrate solution or a luminescence inducing substance can be appropriately selected. As the anti-IgE antibody, either a polyclonal antibody or a monoclonal antibody can be used as long as it can specifically react with IgE to be analyzed, or a fragment thereof, for example, F (ab ′) ₂ , Fab, Fab ′, or Fv can also be used.
[0016]
Operation procedure : (1) Allergen binding step, (2) Test sample contact step, (3) Isolation step, (4) Labeled secondary antibody contact step, and (5) Analysis step The analysis method of the present invention will be described by taking an example of sequential implementation as an example.
In the allergen binding step in the analysis method of the present invention, different types of allergens are bound to a plurality of types of colored beads having different color tones.
In the subsequent test sample contact step in the analysis method of the present invention, the test sample that may contain allergen-binding colored beads obtained in the allergen binding step and IgE that specifically reacts with each of the allergens. Are brought into contact with each other. When IgE that specifically reacts with a specific allergen bound to an allergen-bound colored bead is present in a test sample, the allergen and the allergen-specific IgE are immunized on the colored bead. Each complex is formed.
[0017]
Subsequently, in the isolation step in the analysis method of the present invention, the allergen-bound colored beads are isolated for each color tone. The method for isolating allergen-bound colored beads for each color tone is not particularly limited, but can be isolated by visual observation using an isolation means (for example, tweezers or aspirator).
After the isolation of each allergen-bound colored bead, it is preferable to carry out a washing operation for each allergen-bound colored bead, if desired, before moving to the next labeled secondary antibody contact step. By this washing operation, unreacted substances remaining non-specifically on the allergen-bound colored beads can be removed.
[0018]
In the labeled secondary antibody contact step in the analysis method of the present invention, each allergen-binding colored bead isolated in the above step is brought into contact with an anti-IgE antibody labeled with a labeling substance.
Before moving to the next analysis step, it is preferable to carry out a washing operation for each allergen-bound colored bead, if desired. By this washing operation, unreacted substances remaining non-specifically on the allergen-bound colored beads can be removed.
[0019]
In the subsequent analysis step of the analysis method of the present invention, it is derived from a labeled substance contained in each immune complex consisting of each allergen-binding colored bead, IgE specifically reacting with the allergen, and a labeled anti-IgE antibody. Each signal or each signal derived from a labeling substance contained in an unreacted labeled anti-IgE antibody not contained in each immune complex is analyzed. The method for analyzing each signal derived from the labeling substance can be appropriately selected according to the type of the labeling substance used.
For example, among multiple types of allergens, a signal derived from a labeling substance contained in an immune complex formed on a first allergen-binding colored bead to which the first allergen is bound, or included in the immune complex By analyzing the signal derived from the labeling substance contained in the unreacted labeled anti-IgE antibody, the amount of IgE specifically reacting with the first allergen, that is, the first allergen-specific IgE is determined. be able to. Similarly, each signal derived from a labeled substance contained in each immune complex formed on each allergen-binding colored bead, or included in an unreacted labeled anti-IgE antibody not contained in each immune complex. By analyzing each signal derived from the labeling substance, the amount of each IgE specifically reacting with each allergen can be determined.
[0020]
When the order of the labeled secondary antibody contact step and the isolation step is reversed and the allergen binding step, the test sample contact step, the labeled secondary antibody contact step, the isolation step, and the analysis step are performed in this order. In the analysis step, each signal derived from a labeling substance contained in each immune complex consisting of each allergen-binding colored bead, IgE specifically reacting with the allergen, and a labeled anti-IgE antibody is analyzed. Thus, the allergen-specific IgE amount can be determined.
[0021]
The analysis method of the present invention includes (1) an allergen binding step, (2) a test sample contact step, (3) an isolation step, (4) a labeled secondary antibody contact step, and (5) an analysis step. Although not particularly limited, it can be carried out more specifically, for example, according to the following procedure.
[0022]
(Preparation of allergen-binding colored beads)
First, multiple types (for example, 7 types) of different allergens are respectively bound to multiple types (for example, 7 types) of beads having different colors.
(First reaction)
An appropriate amount (for example, 200 μL) of a sample diluent (also an extract) is dispensed into a sample cup in which an appropriate number (for example, one) of each of the obtained multiple types of allergen-binding colored beads is contained. (Eg 10 μL of serum or 1 punch of dry filter paper blood) is added and incubated at room temperature for several hours to overnight with agitation.
(Bead transfer)
Using an isolation means (eg tweezers etc.), allergen-bound colored beads are dispensed from the sample cup to a microtiter plate (eg 96-well microtiter plate) at 1 bead / well.
(First wash)
Remove unreacted material in each well. For example, it is washed with purified water or PBS (for example, 5 times with 300 μL) by an automatic washer.
(Addition of labeled antibody)
An appropriate amount (for example, 100 μL / well) of an anti-IgE antibody (for example, alkaline phosphatase-labeled anti-human IgE antibody) bound with a labeling substance is added, and incubated at room temperature for a predetermined time (for example, 1 hour) with stirring.
(Second cleaning)
Wash again with purified water, PBS, or the like (for example, 5 times with 300 μL) using, for example, an automatic washer.
(Luminescent substrate solution added)
If desired, a signal inducer is added. For example, when alkaline phosphatase is used as a labeling substance, for example, an adamantyl dioxetane derivative (AMPPD; for example, 100 μL / well) is added as a signal inducing substance, and the mixture is stirred at room temperature for a predetermined time (for example, 30 minutes). Incubate.
(Photometry)
The amount of luminescence of the beads in each well is measured with an automatic photometer (for example, Luminous CT-9000D; Diatron).
[0023]
The analysis kit of the present invention includes (1) a plurality of kinds of colored beads having different color tones and carrying different allergens on the surface thereof for each color tone, and (2) an anti-IgE antibody labeled with a labeling substance. Including.
The analysis kit of the present invention can be used, for example, in the analysis method of the present invention.
[0024]
【Example】
EXAMPLES Hereinafter, the present invention will be specifically described by way of examples, but these do not limit the scope of the present invention.
[Example 1]
In this example, serum was collected from 19 patients with allergic diseases, and the reactivity with respect to mushroom mites, egg whites, milk, wheat, rice, peanuts, and soybeans was measured according to the following procedure by the analysis method of the present invention. The amount of luminescence (cps) was observed. In addition, as a control, comparison was made with the measurement results (IU / mL) of AlaSTAT microplate (ALASTAT Microplate; Diagnostic Products Corporation).
[0025]
Seven kinds of polystyrene beads having a diameter of 3.2 mm [Immunobeads; A-26, A-26 GREEN, A-26 ORANGE, A-26 BLUE, A-26 PINK, A-26 RED, and A-26 YELLOW; Is added to each of the allergen solid phase solutions of mushroom leopard mite, egg white, milk, wheat, rice, peanut, and soy so as to be 20 / mL, stirred for 18 hours, and then washed with PBS. And dried at room temperature on moisture-absorbing paper.
Examples of the allergen solid phase solution include allergen extract purchased from Glia, ie, Dermatophagoides pteronyssinus, egg white (Egg, Chiken, White), milk (Milk, Wow), wheat (Milk, Wow) Rice (Rice), peanut (Peanut), and soy (Soybean) were used.
[0026]
Each of the 7 kinds of allergen-binding colored beads obtained (total 7 beads) was placed in a reaction tube (Eppendorf test tube, Safe-Lock, 2.0 mL) in PBS [0.8% NaCl, 0.02%. KCl, 0.115% Na ₂ HPO ₄ , 0.02% KH ₂ PO ₄ (pH 7.5)] was added together with 190 μL, and 10 μL of each serum collected from 19 patients was added, followed by stirring at room temperature for 18 hours.
Using tweezers, each bead was distributed to a 96-well microtiter plate (Round bottom plate, # 3792; Coaster) at 1 bead / well, washed with an automatic washing machine (washing solution: PBS), phosphatases labeled anti-human IgE monoclonal antibody (Diagnostic Products Corporation) solution (concentration = 0.05μg / mL) 100μL / well aliquots and stirred for 1 hour at room temperature. After washing with an automatic washer, 100 μL / well of a luminescent substrate (PPD reagent; Diagnostic Products Corporation) was dispensed. After stirring at room temperature for 30 minutes, the amount of luminescence was measured with a measuring device (Luminous CT-9000D; Diatron).
[0027]
Moreover, specific IgE concentration measurement was implemented by the conventional method (AlaSTAT method) about 19 types of serum used for the analysis method of the said this invention.
FIG. 1 to FIG. 7 show the correlation between the results obtained by the analysis method of the present invention and the results obtained by the conventional method for the specific IgE amount for each allergen together with the regression line and the correlation coefficient (R). Show. 1 to 7, the horizontal axis (x-axis) is a specific IgE concentration (unit: IU / mL) obtained by the conventional method, and the vertical axis (y-axis) is obtained by the analysis method of the present invention. The amount of emitted light (unit: cps). The results are for the leopard mite, egg white, milk, wheat, rice, peanuts, and soy, respectively. For all allergens and total allergens, the results obtained by the analysis method of the present invention showed a good correlation with the specific IgE concentration measurement results by the conventional method (AlaSTAT method).
[0028]
[Example 2]
In this example, the specific IgE recovery rate in a dry filter paper blood sample was examined.
Blood was collected from 70 healthy subjects and 70 allergic patients, dropped 50 μL each on a filter paper for blood collection [filter paper disk type for blood collection; manufactured by Toyo Filter Paper Co., Ltd.], and left to dry at room temperature for 1 hour. A sample cut into a circular shape with a diameter of 7.5 mm from the center of the dried blood site was put in a tube together with 200 μL of PBS, and thereafter, in the same manner as described in Example 1, reaction with 7 kinds of allergen-binding colored beads, beads , Reaction with labeled anti-human IgE antibody, and measurement of luminescence.
As a control, serum collected and separated at the same time was measured in the same manner as in the method described in Example 1, and the amount of luminescence was compared.
[0029]
8 to 14 show the correlation between the results obtained from the serum samples and the results obtained from the dried filter paper blood samples, together with the regression line and the correlation coefficient (R), for each allergen-specific IgE amount. 8 to 14, the horizontal axis (x-axis) is the luminescence amount (unit: cps) obtained from the serum sample, and the ordinate (y-axis) is the luminescence amount (unit) obtained from the dried filter paper blood. : Cps). Moreover, FIGS. 8-14 is the result regarding a mushroom leopard mite, egg white, milk, wheat, rice, a peanut, and soybean, respectively.
For both allergens and total allergen, light emission quantity obtained from the dried blood spot samples showed good correlation with those obtained from the serum sample.
[0030]
【The invention's effect】
According to the analysis method of the present invention, a specific IgE concentration that cannot be sufficiently measured by a normal measurement method can be analyzed simultaneously for a plurality of causal allergens.
[Brief description of the drawings]
FIG. 1 is a graph showing the correlation between the results obtained by the analysis method of the present invention and the results obtained by the conventional method for the leopard mite allergen.
FIG. 2 is a graph showing the correlation between the results obtained by the analysis method of the present invention and the results obtained by the conventional method for egg white allergens.
FIG. 3 is a graph showing the correlation between the results obtained by the analysis method of the present invention and the results obtained by the conventional method for milk allergens.
FIG. 4 is a graph showing the correlation between the results obtained by the analysis method of the present invention and the results obtained by the conventional method for wheat allergens.
FIG. 5 is a graph showing the correlation between the results obtained by the analysis method of the present invention and the results obtained by the conventional method for rice allergens.
FIG. 6 is a graph showing the correlation between the results obtained by the analysis method of the present invention and the results obtained by the conventional method for peanut allergen.
FIG. 7 is a graph showing the correlation between the results obtained by the analysis method of the present invention and the results obtained by the conventional method for soybean allergens.
FIG. 8 is a graph showing the correlation between the results obtained from a serum sample and the results obtained from a dry filter paper blood sample for the leopard mite allergen.
FIG. 9 is a graph showing the correlation between the results obtained from serum samples and the results obtained from dried filter paper blood samples for egg white allergens.
FIG. 10 is a graph showing the correlation between the results obtained from serum samples and the results obtained from dry filter paper blood samples for milk allergens.
FIG. 11 is a graph showing the correlation between results obtained from serum samples and results obtained from dry filter paper blood samples for wheat allergens.
FIG. 12 is a graph showing the correlation between the results obtained from serum samples and the results obtained from dry filter paper blood samples for rice allergens.
FIG. 13 is a graph showing the correlation between the results obtained from serum samples and the results obtained from dry filter paper blood samples for peanut allergens.
FIG. 14 is a graph showing the correlation between the results obtained from serum samples and the results obtained from dry filter paper blood samples for soybean allergens.

Claims (3)

(1) A step of binding different kinds of allergens for each color tone to a plurality of types of colored beads having a diameter of 2 to 5 mm having different color tones;
(2) contacting the allergen-binding colored beads obtained in the step (1) with a test sample that may contain each immunoglobulin E that specifically reacts with each of the allergens;
(3) a step of isolating the allergen-binding colored beads for each color tone;
(4) contacting each allergen-binding colored bead isolated in the step (3) with an anti-immunoglobulin E antibody labeled with a labeling substance; and (5) each allergen-binding colored bead, Each signal derived from a labeling substance contained in each immune complex consisting of immunoglobulin E specifically reacting with allergen and labeled anti-immunoglobulin E antibody, or unreacted not contained in each immune complex A method for analyzing allergen-specific immunoglobulin E, comprising the step of analyzing each signal derived from a labeling substance contained in the labeled anti-immunoglobulin E antibody. (1) A step of binding different kinds of allergens for each color tone to a plurality of types of colored beads having a diameter of 2 to 5 mm having different color tones;
(2) contacting the allergen-binding colored beads obtained in the step (1) with a test sample that may contain each immunoglobulin E that specifically reacts with each of the allergens;
(3) a step of bringing the mixture obtained in the step (2) into contact with the anti-immunoglobulin E antibody labeled with a labeling substance;
(4) a step of isolating the allergen-binding colored beads for each color tone; and (5) each allergen-binding colored bead isolated in the step (4) and immunoglobulin E specifically reacting with the allergen. And a method for analyzing allergen-specific immunoglobulin E, comprising a step of analyzing each signal derived from a labeled substance contained in each immune complex comprising a labeled anti-immunoglobulin E antibody. (1) a plurality of colored beads having a diameter of 2 to 5 mm having different color tones and carrying different allergens on the surface for each color tone;
(2) A kit for analysis of immunoglobulin E that specifically reacts only with any one of the aforementioned allergens, comprising an anti-immunoglobulin E antibody labeled with a labeling substance.

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