Serological test for detecting anti-HPV antibody and test kit

Abstract

The present disclosure relates to serological assays for detecting anti-HPV antibodies. The serological test uses an antigen having a linear epitope that is reactive and that enables selective specific binding to multiple or many antibodies, each of which is effective against a different type of HPV. The disclosure also relates to a test kit (1; 1.1; 1.1; 1.2; 1.3, 2, 3; 1.4) of the method.

Description

Technical Field

[0001] The present disclosure relates to serological tests for detecting anti-HPV antibodies and test kits for such serological tests. In particular, the present disclosure includes methods for qualitatively and / or quantitatively determining anti-HPV antibodies contained in body fluids.

[0002] Serological tests use in vitro antigen-antibody reactions. These reactions can be used, for example, to identify autoantibodies in blood samples that are specific for certain antigens and affect pathogens such as viruses and bacteria. These antibodies are specifically produced by the human immune system as an immune response against these antigens to protect the body.

[0003] Serological tests are particularly used to detect antibodies against the human papillomavirus (abbreviated as HPV). For example, such a test can be learned from the publication WO 2012 / 097788 A1 and is used as the basis and description of the treatment control method in the publication WO 2018 / 108957A1. The publications are hereby incorporated by reference to supplement the present disclosure, and it should be noted that these publications may give different meanings to the same terms as those herein. The known test is capable of identifying antibodies that are particularly effective against HPV type 16.

[0004] Type 16 is one of the HPV types known as high-risk HPV. It is now known that high-risk types play an important role in the development of precancerous lesions (dysplasia) and resulting cancers, especially in the cervix. It is also now known that only 25% of HPV-induced precancerous lesions and cancers are associated with HPV type 16. 75% of HPV-induced precancerous lesions and cancers are caused by other high-risk types. So far, this important proportion has been ignored in the tests.

[0005] Therefore, there may be a need to use serological tests that also detect antibodies effective against HPV types other than HPV type 16. This is based on the expectation that by detecting as many HPV types as possible, a further improvement in the prediction of HPV-induced precancerous lesions and cancers in serological tests will be obtained.

[0006] Concept

[0007] A fundamental improvement in detection has been achieved by a serological method for detecting anti-HPV antibodies, which uses an antigen having linear epitopes, particularly having at least one linear epitope or only one linear epitope, wherein the linear epitope preferably has reactivity and enables multiple or many antibodies (each of the multiple or many antibodies being effective against different types of HPV) to selectively bind specifically thereto or be capable of selectively binding specifically thereto.

[0008] The term "serological method" as used in the present disclosure should be understood to mean or include a serological test or a serological test method or assay or immunoassay. The method is preferably an in vitro method, that is, a method carried out in vitro.

[0009] The term "in vitro" as used in the present disclosure should be particularly understood to mean that the method is carried out outside a living organism, that is, not on a human being. To carry out the method in vitro, at least one sample, such as a body fluid, is collected from the relevant person. For example, one drop (about 25 μL) is sufficient to carry out the method.

[0010] The term "antigen" as used in the present disclosure should be understood to mean any type of protein that triggers an immune response in the immune system of a person in contact with it. The term "antigen" as used in the present disclosure also includes virus-like particles (VLPs).

[0011] The term "virus-like particle" or "VLP" as used in the present disclosure should be understood to mean a particle composed of or including a viral capsid but not containing any nucleic acid. Thus, VLPs are able to present the above-mentioned linear epitopes and are not able to self-replicate.

[0012] The term "specifically binds" as used in the present disclosure should be particularly understood to mean that an antibody binds only to a specific binding site of an antigen. Thus, an antibody binds to an antigen only via such a binding site. Thus, with respect to a plurality or many antibodies that can bind to a linear epitope of an antigen, it should be understood that the antibodies considered herein are antibodies that bind to or can specifically bind to the linear epitope.

[0013] By using a specific linear epitope, a single serological test can be used to determine the presence of precancerous lesions and cancers caused by an entire group of HPV types. This is based on the unexpectedly discovered common feature of these HPV types, namely that they have the same amino acid sequence. The improved method makes use of this common feature, that is, the specific linear epitope of the antigen has the said amino acid sequence.

[0014] However, the linear epitope also offers other advantages. The antigen does not need to be stored in a physiologically active fluid to maintain the reactivity of the linear epitope. The antigen can also be in a dry or solid form. Storage of the antigen also does not require any refrigeration. The antigen can be stored, for example, at room temperature without any loss of potency or at least without any relevant loss of potency. These are the main differences from known tests using antigens with conformational epitopes. To maintain the reactivity of a conformational epitope, the antigen must be kept cold in a physiologically active fluid therein.

[0015] The antigen used in the improved method can be polymeric, that is, exist in a polymeric form. The term "polymeric" as used in the present disclosure should be understood to mean a molecule or molecular complex in which the above-mentioned linear epitope occurs at least twice.

[0016] Alternatively, the antigen can be monomeric, i.e., exist in monomeric form. The term "monomer" as used in the present disclosure should be understood to mean a molecule in which the above linear epitope occurs only once.

[0017] In particular, it is provided to use a labeled antibody that specifically binds to the linear epitope of the antigen or that can specifically bind to the linear epitope of the antigen. Thus, the labeled antibody is an antibody against an HPV type having an amino acid sequence presented or represented by the linear epitope of the antigen. These antibodies are preferably labeled using methods known per se, for example.

[0018] In particular, preferably by labeling, the labeled antibody causes, in the presence of reactants, for example, a measurable and / or user-perceivable reaction, or in the case of spatial accumulation, especially in the case of spatial accumulation of the bound labeled antibody, for example, a measurable and / or user-perceivable change. Thus, the labeled antibody is capable of identifying anti-HPV antibodies contained in body fluids. The reaction or change can be measured optically.

[0019] According to a first embodiment, the improved method can be a method for qualitatively and / or quantitatively determining anti-HPV antibodies contained in body fluids and particularly includes the following steps or consists of the following steps: providing a carrier medium, such as a test strip, on which a predetermined amount of the labeled antibody is presented in a mobile manner, and a predetermined amount of the antigen is present at a predetermined position, especially presented in an immobile manner, for example, fixed; bringing a preferably predetermined volume of body fluid into contact with the labeled antibody (i.e., the labeled antibody located on the carrier medium); and qualitatively and / or quantitatively identifying the anti-HPV antibodies by analyzing a change in a region of the carrier medium, for example, a preferably predetermined position.

[0020] In particular, in this case, the labeled antibody causes, for example, in the case of binding, i.e., in the case of binding to the linear epitope of the antigen, if there is spatial accumulation, i.e., spatial accumulation of the labeled antibody, they will cause a measurable and / or user-perceivable change. For example, the labeled antibody is labeled with gold particles and / or silver particles.

[0021] The step of bringing the body fluid into contact with the labeled antibody particularly includes bringing the body fluid into direct contact with the labeled antibody. This also particularly includes bringing the body fluid into indirect contact with the labeled antibody. This can be done via the carrier medium. For example, the body fluid is applied to a position on the carrier medium remote from the labeled antibody. For example, in this case, as the body fluid flows on the carrier medium towards the labeled antibody, the body fluid comes into contact with the labeled antibody. Additionally or alternatively, capillary action can be used. For example, the carrier medium comprises or consists of a material designed to produce a capillary effect on fluids.

[0022] This embodiment is based on a competition method known per se, in which any anti-HPV antibodies contained in the body fluid compete with the labeled antibody for binding to the linear epitope of the antigen. The improved method according to this embodiment can be carried out in only a few steps. The body fluid can be immediately applied to the carrier medium. For the body fluid, no prior steps are required. The carrier medium used, on which the labeled antibody is presented in a mobile manner and the antigen is presented in an immobile manner, enables a rapid test to be provided, in which a change that can be rapidly measured and / or perceived by the user occurs in the reaction zone.

[0023] According to a second alternative embodiment, the improved method can be a method for qualitatively and / or quantitatively determining anti-HPV antibodies contained in a body fluid, and particularly includes the following steps or consists of the following steps: providing a predetermined amount of antigen; providing a carrier medium, such as a test strip, on which a predetermined amount of labeled antibody is presented in a mobile manner and a predetermined amount of capture molecule is present at a predetermined position, particularly presented in an immobile manner, such as fixed; bringing a preferably predetermined volume of body fluid into contact with the provided antigen; bringing the body fluid and the provided antigen into contact with the labeled antibody (i.e., the labeled antibody located on the carrier medium); and qualitatively and / or quantitatively identifying the anti-HPV antibodies by analyzing a change in a region of the carrier medium, such as a preferably predetermined position.

[0024] Bringing the body fluid and the antigen into contact with the labeled antibody particularly includes bringing the body fluid and the antigen into direct contact with the labeled antibody. This also particularly includes bringing the body fluid and the antigen into indirect contact with the labeled antibody. This can be done via the carrier medium. For example, the body fluid and the antigen are applied to a position on the carrier medium remote from the labeled antibody. For example, in this case, as the body fluid and the antigen flow on the carrier medium towards the labeled antibody, the body fluid and the antigen come into contact with the labeled antibody. Additionally or alternatively, the capillary effect can also be used. For example, the carrier medium includes or consists of the following: a material designed to produce a capillary effect on the fluid.

[0025] Bringing the body fluid into contact with the antigen and bringing the body fluid and the antigen into contact with the labeled antibody can be understood to mean two preferably separate steps or one step. Bringing the body fluid into contact with the antigen and bringing the body fluid and the antigen into contact with the labeled antibody can be understood, for example, to mean performing the steps at different times. Particularly, first the body fluid is brought into contact with the antigen, and then the body fluid and the antigen are brought into contact with the labeled antibody. Particularly, the body fluid should be incubated with the antigen for a predetermined period and / or a freely selected period, and then brought into contact with the labeled antibody. If the body fluid contains antibodies that can bind to the linear epitope of the antigen, the interaction between these antibodies and the antigen can occur before the body fluid and the antigen come into contact with the labeled antibody.

[0026] Contacting the body fluid with the antigen and contacting the body fluid and the antigen with the labeled antibody can also be understood to mean performing the steps simultaneously or performing combined steps. For example, in this case, the body fluid, the antigen, and the labeled antibody are contacted simultaneously.

[0027] In particular, in this embodiment, the labeled antibody causes, for example, in the case of binding, that is, in the case of binding to the linear epitope of the antigen, if there is spatial accumulation, that is, spatial accumulation of the labeled antibody, they will cause a measurable and / or user-perceivable change. For example, the labeled antibody is labeled with gold particles and / or silver particles.

[0028] For example, a predetermined amount of antigen is stored (in particular dissolved) in a preferably predetermined volume of buffer fluid, in particular buffer solution. In this case, a buffer fluid containing the antigen is provided and a predetermined volume of body fluid is contacted therewith. Then the combined volume or a predetermined volume thereof is contacted with the labeled antibody located on the carrier medium. In principle, the buffer fluid and the antigen, the body fluid, and the labeled antibody contained therein are also contacted simultaneously.

[0029] This embodiment is also based on a competition method known per se, in which any anti-HPV antibodies contained in the body fluid compete with the labeled antibody for binding to the linear epitope of the antigen. The improved method according to this embodiment can also be carried out in only a few steps. The carrier medium used, on which the labeled antibody is presented in a mobile manner and the capture molecule is presented in an immobile manner, also enables a rapid test to be provided, in which a change that can be rapidly measured and / or perceived by the user occurs in the reaction zone.

[0030] If the body fluid is contacted with the antigen in a previous step, any anti-HPV antibodies contained in the body fluid will be directly spatially close to each other, that is, before the composition is contacted with the labeled antibody. As a result, the interaction between the antibody and the antigen in the body fluid will occur in advance and thus before any interaction between the antigen and the labeled antibody. If the body fluid contains anti-HPV antibodies, this makes it easier for a measurable and / or user-perceivable change to occur rapidly.

[0031] In one configuration, the capture molecules are such that they can each specifically bind to the linear epitope of the antigen. As a result, those antigens bound to the capture molecules remain at the position of the capture molecules on the carrier medium. In this case, the antigen is polymeric or present in polymeric form.

[0032] In another configuration, capture molecules are such that they can each specifically bind to different binding sites of the antigen other than linear epitopes. In this case, the antigen is monomeric or present in monomeric form. The additional binding sites can in principle be epitopes. In this case, the capture molecules are such that they can each specifically bind to different epitopes of the antigen other than linear epitopes. The additional epitopes can be linear epitopes or conformational epitopes.

[0033] According to a third alternative embodiment, the improved method can be a method for the qualitative and / or quantitative determination of anti-HPV antibodies contained in a body fluid, and particularly comprises the following steps or consists of the following steps: providing a predetermined amount of antigen, preferably presented immovably on or at a carrier medium such as a microtiter plate, for example immobilized on or at the carrier medium; bringing a preferably predetermined volume of the body fluid into contact with the provided antigen; bringing a predetermined amount of labeled antibody into contact with the body fluid and the provided antigen; washing the carrier medium; applying a predetermined volume of a reactant to the carrier medium; and qualitatively and / or quantitatively identifying anti-HPV antibodies by preferably analyzing the reaction of the carrier medium, for example.

[0034] Bringing the body fluid into contact with the antigen and bringing the labeled antibody into contact with the body fluid and the antigen can be understood to mean two preferably separate steps or one step. Bringing the body fluid into contact with the antigen and bringing the labeled antibody into contact with the body fluid and the antigen can, for example, be understood to mean performing the steps at different times. In particular, first the body fluid is brought into contact with the antigen, and then the labeled antibody is brought into contact with the body fluid and the antigen. In particular, the body fluid and the antigen should be incubated for a predetermined period and / or a freely chosen period before bringing the labeled antibody into contact with the composition. If the body fluid contains antibodies that can bind to the linear epitopes of the antigen, then the interaction between these antibodies and the antigen can thus occur before adding the labeled antibody to the body fluid.

[0035] In principle, it can be stipulated that the body fluid is first brought into contact with the antigen, that is, for example, placed on the carrier medium, and then, for example, after a predetermined period and / or a freely chosen period, the carrier medium is washed. It can also be stipulated that the labeled antibody is added only afterwards.

[0036] The term "washing" as used in the present disclosure should be understood to particularly mean removing substances present on the carrier medium (other than antibodies that specifically bind to the linear epitopes of the antigen) from the carrier medium. This removes substances contained in the body fluid that may, for example, interfere in an undesirable manner with the interaction of the labeled antibody with the linear epitopes of the antigen.

[0037] In particular, in this embodiment, the labeled antibodies are such that, in the presence of a reactant, they cause a measurable and / or user-perceivable reaction. The reactant used in the improved method is preferably such that, due to it, the labeled antibodies cause a measurable and / or user-perceivable reaction. The reaction can be a color reaction, for example, if the labeled antibody is labeled with an enzyme and the reactant is a reactant of the enzyme.

[0038] This embodiment is also based on a competition method known per se, in which any anti-HPV antibodies contained in the body fluid compete with the labeled antibodies for binding to the linear epitope of the antigen. The advantage of this embodiment is that the incubation time and / or incubation temperature can be freely selected. Therefore, in this embodiment, the improved method can be implemented by optimizing the settings of the incubation time and / or incubation temperature, so that only a relatively small amount of antigen is required.

[0039] According to a fourth alternative embodiment, the improved method can be a method for qualitatively and / or quantitatively determining anti-HPV antibodies contained in a body fluid, and particularly includes the following steps or consists of the following steps: providing a predetermined amount of antigen, preferably on or at a carrier medium, particularly presented in an immobile manner, such as immobilized on or at the carrier medium; bringing a preferably predetermined volume of body fluid into contact with the antigen; washing the carrier medium; applying a predetermined amount of labeled antibody to the carrier medium; washing, particularly re-washing or washing the carrier medium again; applying a predetermined volume of reactant to the carrier medium; and qualitatively and / or quantitatively identifying the anti-HPV antibodies by preferably analyzing the reaction of the carrier medium, for example.

[0040] In this case, the labeled antibody is such that it can specifically bind to a human antibody that specifically binds or can specifically bind to the linear epitope of the antigen. In this case, the labeled antibody is such that it can specifically bind to a different binding site of the human antibody, which is a binding site other than the binding site that specifically binds or can specifically bind to the linear epitope of the antigen. Therefore, if such human antibodies are contained in the body fluid and if these human antibodies bind to the linear epitope of the antigen, binding to these bound human antibodies will occur after the application of the labeled antibody.

[0041] In particular, the labeled antibodies are such that, in the presence of a reactant, they cause a measurable and / or user-perceivable reaction. The reactant used in the improved method is preferably such that, due to it, the labeled antibodies cause a measurable and / or user-perceivable reaction. The reaction can be a color reaction, for example, if the labeled antibody is labeled with an enzyme and the reactant is a reactant of the enzyme.

[0042] Due to the labeled antibodies used here, the improvement method of this embodiment is based on the sandwich method known per se, in which the labeled antibodies do not bind directly and specifically to the antigen, but indirectly via human antibodies.

[0043] The improvement method of this embodiment enables the qualitative and / or quantitative determination of any anti-HPV antibodies contained in body fluids. Additionally, it is also possible to indicate which type of antibody is present here. For example, it can be determined whether the identified antibody is immunoglobulin A (IgA) or immunoglobulin D (IgD) or immunoglobulin E (IgE) or immunoglobulin G (IgG) or immunoglobulin M (IgM). For this purpose, the labeled antibody only needs to be based on an antibody that specifically binds to a human antibody presenting IgA or IgD or IgE or IgG or IgM. If the human antibody presents, for example, IgA and a measurable and / or user-perceivable reaction occurs, it means that the detected anti-HPV antibody is an antibody of the IgA type.

[0044] Important information can be obtained by identifying the type of antibody. For example, the identified IgA-type antibody indicates that the mucosa of the person from whom the body fluid was obtained is affected. For example, the identified IgM-type antibody indicates a new infection. For example, the identified IgD-type antibody indicates a gastrointestinal infection.

[0045] In the improved method, it is specifically stipulated that the predetermined amount of antigen is less than or corresponds to the predetermined amount of labeled antibody. It has been proven that through this ratio between the antigen and the labeled antibody, the above-mentioned changes or reactions caused by the labeled antibody can be achieved to the desired extent in order to enable the qualitative and / or quantitative identification of anti-HPV antibodies. This is especially the case if the improved method is based on one of the above embodiments with a competitive method. In principle, the predetermined amount of antigen can also be more than the predetermined amount of labeled antibody.

[0046] In the improved method, the predetermined amount of antigen and the predetermined amount of labeled antibody are particularly compared and measured with the body fluid volume, such that if the amount of antibodies in the body fluid that specifically bind to the linear epitope of the antigen is higher than a predetermined minimum amount, in the presence of reactants, the labeled antibody causes a measurable and / or user-perceivable reaction, or in the case of spatial accumulation, leads to a measurable and / or user-perceivable change. Thereby, the sensitivity of the method can be adjusted. To adjust the predetermined minimum amount, data from confirmed clinical diagnoses and / or from proven healthy individuals can be used.

[0047] In particular, the predetermined minimum amount is set to a value higher than the amount required by healthy individuals. Therefore, the improved method has no effect on healthy individuals. In particular, the predetermined minimum amount is set to a value higher than the amount required by healthy individuals and lower than the amount of HPV-infected individuals. Therefore, the improved method has no effect on HPV-infected individuals.

[0048] The term "HPV infection" as used in the present disclosure should be specifically understood to mean that the person concerned has or has had an HPV-related disease, and thus, the person's immune system is producing or has produced anti-HPV antibodies. In other words, "HPV infection" specifically means that the person concerned has antibodies caused by an HPV-related disease, in particular antibodies of the person's own body.

[0049] In the improved method, the linear epitope of the antigen specifically enables multiple or many antibodies that selectively bind thereto to be effective against different types of HPV, said different types being from a category having at least two types of 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, 73 and 82. These HPV types form a high-risk group that plays a key role in the development of precancerous lesions (dysplasia) and the resulting cancer. If two of these types of HPV are covered by the linear epitope, the linear epitope targets an amino acid sequence that is also present in two of these types of HPV. The more of these types of HPV that are covered by the common amino acid sequence and this amino acid sequence is represented by the linear epitope, the higher the sensitivity of the improved method in identifying precancerous lesions and cancer.

[0050] In the improved method, the linear epitope of the antigen specifically enables multiple or many antibodies that selectively bind thereto to be effective against the L1 protein of HPV. Alternatively, the linear epitope of the antigen can enable multiple or many antibodies that selectively bind thereto to be effective against the L2 protein of HPV. Alternatively, the linear epitope of the antigen can enable multiple or many antibodies that selectively bind thereto to be effective against the E6 protein of HPV. Alternatively, the linear epitope of the antigen can enable multiple or many antibodies that selectively bind thereto to be effective against the E7 protein of HPV. Alternatively, the linear epitope of the antigen can enable multiple or many antibodies that selectively bind thereto to be effective against different HPV proteins from the L1, L2, E6 and E7 groups, respectively.

[0051] The term "body fluid" as used in the present disclosure should be specifically understood to mean water present in various compartments of the human body, including substances dissolved therein. Body fluid can be blood and / or lymph and / or saliva and / or urine and / or sweat. For example, body fluid is whole blood or serum and / or plasma of whole blood. For example, body fluid is serum and / or plasma of a blood sample obtained by venipuncture. Body fluid can also be capillary blood or whole blood from a human fingertip. In particular, body fluid is a body fluid sample.

[0052] Improved identification can be achieved using a test kit for detecting anti-HPV antibodies, the test kit comprising an antigen having a linear epitope, in particular having at least one linear epitope or only one linear epitope, the linear epitope preferably being reactive and enabling a plurality or many antibodies to selectively bind thereto specifically, wherein each of the plurality or many antibodies is effective against a different type of HPV. In particular, the antigen is the antigen described above for the improved method. The test kit is capable of implementing the improved method. In this regard, the advantages of the improved method can be achieved by means of the test kit.

[0053] Furthermore, due to the specific antigen used, the improved test kit has the advantage that it does not require refrigeration for storage. The test kit can be stored, for example, at room temperature without any loss of potency or at least without any relevant loss of potency. The improved test kit also has the advantage that it does not require any physiologically active fluid for storing the antigen to maintain the reactivity of the antigen. Even if the antigens are present, for example, in solid form and / or in a dry state and / or in a non-physiologically active fluid, they still maintain their reactivity.

[0054] In particular, the improved test kit comprises labeled antibodies which enable them to bind to the linear epitopes of the antigen. In particular, the labeled antibodies cause, for example, a measurable and / or user-perceivable reaction in the presence of reactants, or in the case of spatial accumulation, in particular in the case of spatial accumulation of the bound labeled antibodies, for example, a measurable and / or user-perceivable change. This thus enables the identification of anti-HPV antibodies contained in body fluids. In particular, the labeled antibodies are the labeled antibodies described above with respect to the improved method.

[0055] In one embodiment, the improved test kit comprises a carrier medium on which a predetermined amount of labeled antibodies is presented in a mobile manner and a predetermined amount of antigen is present at a predetermined position, in particular presented in a non-mobile manner, for example fixed. This enables the implementation of the improved method according to the first embodiment described above.

[0056] In particular, the labeled antibodies are arranged at a certain distance from the antigen. In particular, the labeled antibodies are arranged in the region of the body fluid application position. In particular, the labeled antibodies cause, for example, a measurable and / or user-perceivable change in the case of binding, that is to say in the case of binding to the linear epitope of the antigen, if there is spatial accumulation, that is to say spatial accumulation of the labeled antibodies.

[0057] The improved test kit of this embodiment can be simplified to the carrier medium as the only component. The antigen has been stored on the carrier medium. This is possible because the antigen does not require any physiologically active fluid to maintain its reactivity. For example, the antigen exists in a dry form, such as in a dry form on the carrier medium. In principle, the labeled antibody can also exist in a dry form, such as in a dry form on the carrier medium.

[0058] In another embodiment, the improved test kit includes a predetermined amount of antigen, for example, in a container, such as dissolved in a buffer fluid, especially a buffer solution. The improved test kit further includes a carrier medium on which a predetermined amount of labeled antibody is presented in a mobile manner, and a predetermined amount of capture molecules are present at a predetermined position, especially presented in an immobile manner, such as fixed. This enables the implementation of the improved method according to the second embodiment described above.

[0059] In particular, the labeled antibody is arranged at a certain distance from the capture molecule. In particular, the labeled antibody is arranged in the region of the body fluid application position. In particular, the labeled antibody causes, for example, in the case of binding, that is, in the case of binding to the linear epitope of the antigen, if there is spatial accumulation, that is, the spatial accumulation of the labeled antibody, they will cause a measurable and / or user-perceivable change.

[0060] If the antigen is a polymer, that is, exists as a polymer, the capture molecules can each specifically bind to the linear epitope of the antigen. In this case, the improved method can be implemented according to the first configuration of the second embodiment described above. For example, the capture molecule is based on the same antibody as the basis for forming the labeled antibody. This saves development costs because only a single antibody needs to be developed.

[0061] Alternatively, the antigen can be monomeric. This results in a cost advantage because the antigen used is relatively inexpensive as a monomer. If the antigen is monomeric, that is, exists as a monomer, the capture molecules can each specifically bind to different binding sites of the antigen other than the linear epitope. In this case, the improved method can be implemented according to the second configuration of the second embodiment described above. The additional binding site can be an epitope. In this case, the capture molecule is such that it can each specifically bind to a different epitope of the antigen other than the linear epitope. The additional epitope can be a linear epitope or a conformational epitope.

[0062] The above buffer fluid can be a salt-based solution. Alternatively, the buffer fluid can be TRIS-BSA (tris(hydroxymethyl)aminomethane and bovine serum albumin) or a histidine buffer.

[0063] The carrier medium of the above-described embodiments of the improved test kit may be a test strip. For example, the test strip comprises or consists of a material that exerts a capillary force on the fluid applied thereto. For example, the test strip comprises or consists of blotting paper material. The test strip may be located in a test cassette.

[0064] In yet another embodiment, the improved test kit comprises a predetermined amount of antigen, which is preferably presented on a carrier medium, in particular in an immobile manner, for example immobilized on the carrier medium. The improved test kit further comprises a predetermined amount of labeled antibody. In particular, in the presence of a reactant, the labeled antibody causes a measurable and / or user-perceivable reaction. Additionally, the improved test kit comprises a predetermined volume of reactant, due to which the labeled antibody causes a measurable and / or user-perceivable reaction.

[0065] This embodiment enables the implementation of the improved method according to the above-described third embodiment. This embodiment enables the detection limit for identifying any anti-HPV antibodies contained in a body fluid to be changed (e.g., reduced) by adjusting the incubation time and / or the incubation temperature.

[0066] In this case, the carrier medium may be a reagent container, such as a microtiter plate. The antigen may be present therein, for example, on the bottom and / or the walls. Preferably, the antigen is immobilized in the reagent container, in particular on the bottom and / or the walls. For example, the bottom and / or the walls of the reagent container are coated with the antigen.

[0067] In a modification of the above-described embodiment, the improved test kit now comprises a labeled antibody that specifically binds to human antibodies, rather than a labeled antibody that binds to a linear epitope of an antigen, and these human antibodies specifically bind or can specifically bind to the linear epitope of the antigen. The labeled antibodies used herein in particular cause a measurable and / or user-perceivable reaction in the presence of a reactant.

[0068] This modification enables the implementation of the improved method according to the above-described fourth embodiment. In this regard, this modification enables any anti-HPV antibodies contained in a body fluid to be qualitatively and / or quantitatively determined, and additionally, it is also possible to state which type of antibody is present herein.

[0069] The above-described labeled antibodies may be based on labeled monoclonal antibodies, such as mouse monoclonal antibodies. These monoclonal antibodies may be obtained from a hybridoma cell line. This ensures that the antibodies obtained therefrom have no or at least no relevant differences from each other. The term "hybridoma cell line" should be understood to particularly mean a cell line obtained by means of hybridoma technology.

[0070] If the labeled antibodies are intended to produce a measurable and / or user-perceivable change in the case of spatial accumulation, these antibodies are labeled, for example, with colloidal gold and / or colloidal silver.

[0071] If the labeled antibodies are intended to cause a measurable and / or user-perceivable reaction in the presence of a reactant, these antibodies are labeled with an enzyme such as horseradish peroxidase (HRP) or alkaline phosphatase (AP). The reactant used can be 3,3′,5,5′-tetramethylbenzidine (TMB) or Fast Red, and thus the reaction caused by it is a color change.

[0072] If the reaction or change caused is to be measured, the improved test kit can include a photometer or some other optical photographic device.

[0073] According to one aspect, there is provided the use of an antigen in an improved method and / or an improved test kit. The antigen includes a linear epitope, in particular at least one linear epitope or only one linear epitope, which enables a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies is effective against a different type of HPV.

[0074] According to a further aspect, there is provided an antibody or a labeled antibody for use in a diagnostic method, in particular for a method for qualitatively and / or quantitatively determining anti-HPV antibodies contained in a body fluid. The antibody or the labeled antibody enables it to specifically bind to the linear epitope of the antigen. The linear epitope of the antigen enables a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies is effective against a different type of HPV.

[0075] In particular, the labeled antibody causes a measurable and / or user-perceivable reaction in the presence of a reactant, or in the case of spatial accumulation, especially in the case of spatial accumulation of this type of bound labeled antibody, resulting in a measurable and / or user-perceivable change.

[0076] According to a further aspect, there is provided the use of a labeled antibody in at least one of the above embodiments of an improved method and / or an improved test kit. The labeled antibody enables it to specifically bind to the linear epitope of the antigen. The linear epitope of the antigen enables a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies is effective against a different type of HPV.

[0077] In particular, the labeled antibody causes a measurable and / or user-perceivable reaction in the presence of a reactant, or in the case of spatial accumulation, especially in the case of spatial accumulation of this type of bound labeled antibody, resulting in a measurable and / or user-perceivable change.

[0078] According to a further aspect, there is provided the use of an antibody as a capture molecule in at least one of the above-described embodiments of an improved method and / or an improved test kit. The antibody specifically binds to a linear epitope of an antigen, wherein the linear epitope enables a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies is effective against a different type of HPV.

[0079] According to a further aspect, there is provided the use of a molecule as a capture molecule in at least one of the above-described embodiments of an improved method and / or an improved test kit. The molecule specifically binds to a different binding site of the antigen other than the linear epitope, the linear epitope enabling a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies is effective against a different type of HPV. In particular, the molecule is an antibody that enables it to specifically bind to a different epitope of the antigen other than the linear epitope. BRIEF DESCRIPTION OF THE DRAWINGS

[0080] Several exemplary embodiments are described below with reference to the accompanying drawings to clarify further details and features, wherein

[0081] Figure 1 A first exemplary embodiment of a test kit for detecting anti-HPV antibodies is shown in schematic form,

[0082] Figure 2 A second exemplary embodiment of a test kit for detecting anti-HPV antibodies is shown in schematic form,

[0083] Figure 3 A third exemplary embodiment of a test kit for detecting anti-HPV antibodies is shown in schematic form,

[0084] Figure 4 A fourth exemplary embodiment of a test kit for detecting anti-HPV antibodies is shown in schematic form,

[0085] Figure 5 A fifth exemplary embodiment of a test kit for detecting anti-HPV antibodies is shown in schematic form.

[0086] EXEMPLARY EMBODIMENTS

[0087] Figure 1Shows a first exemplary embodiment of a test kit 1 for detecting anti-HPV antibodies. The exemplary test kit 1 includes a carrier medium 2 on which a predetermined amount of antigen 10 and a predetermined amount of labeled antibody 20 are present. Preferably, the predetermined amount of antigen 10 is arranged at a predetermined position 3 of the carrier medium 2. For example, the predetermined position 3 is configured as a test line. The antigen 10 is preferably immobilized, especially fixed at the predetermined position 3.

[0088] Preferably, the labeled antibody 20 is presented in a mobile manner, and the predetermined amount of labeled antibody 20 is located at a certain spatial distance from the predetermined position 3. Preferably, the labeled antibody 20 is arranged in the region of the application position 4. The application position 4 represents the position of the carrier medium 2 for receiving the fluid to be examined.

[0089] For example, the carrier medium 2 is a test strip. For example, the test strip is provided in a test cassette. For example, the test cassette has a test opening in the region of the predetermined position 3. For example, the test cassette has an application opening in the region of the application position 4 through which the fluid to be examined can be applied to the test strip.

[0090] For example, the carrier medium 2 comprises or consists of the following: a material that exerts a capillary force on the fluid. The fluid applied to the region of the application position 4 on the carrier medium 2 is thus supported to flow towards the predetermined position 4. For example, the material is blotting paper or the like.

[0091] In the exemplary test kit 1, the antigen 10 has a linear epitope. The linear epitope is reactive and enables a plurality or many antibodies to selectively bind thereto specifically, each of the plurality or many antibodies being effective against a different type of HPV. The antigen 10 can exist as a monomer or a multimer. Preferably, the antigen 10 is presented in a dry form on the carrier medium 2 and / or the antigen 10 is presented in a dry form on the carrier medium 2.

[0092] According to the antigen 10, the labeled antibody 20 enables it to specifically bind to the linear epitope of the antigen 10. The labeled antibody 20 also has a label such that the spatial accumulation of the labeled antibody 20 results in a measurable and / or user-perceivable change. For example, the labeled antibody 20 is a monoclonal mouse antibody provided with such a label. For example, the labeled antibody is labeled with colloidal gold and / or colloidal silver.

[0093] Preferably, the predetermined amount of antigen 10 is less than the predetermined amount of labeled antibody 20. In principle, the predetermined amount of antigen 10 can also correspond to or be more than the predetermined amount of labeled antibody 20. Preferably, the predetermined amount of antigen 10 and the predetermined amount of labeled antibody 20 are also compared and measured with respect to the volume of the fluid sample to be examined such that if the amount of antibodies contained in the fluid sample and specifically binding to the linear epitope of antigen 10 is higher than a predetermined minimum amount, the labeled antibody 20 causes a measurable and / or user-perceivable change in the case of spatial accumulation.

[0094] The exemplary test kit 1 is capable of qualitatively and / or quantitatively determining anti-HPV antibodies contained in body fluids by serological methods. By using an antigen 10 having its specific linear epitope, a single serological test can be used to judge the presence of precancerous lesions and cancers caused by the entire group of HPV types.

[0095] For the serological test to be carried out, a body fluid sample of a human being must be provided. Preferably, the body fluid is blood, such as whole blood, or serum and / or plasma from blood. The blood can be obtained by venipuncture or can be whole blood from a fingertip. The body fluid is applied to the carrier medium 2 via the application location 4. There, the body fluid comes into contact with the labeled antibody 20 and, together with at least some of the labeled antibody 20, optionally by utilizing the capillary forces caused by the material of the carrier medium 2, flows via the carrier medium 2 to a predetermined location 3 and comes into contact with the antigen 10. Due to the contact, on the one hand, an interaction occurs between the antigen 10, the labeled antibody 20, and any antibodies contained in the body fluid, and on the other hand, the labeled antibody 20 and the antibodies contained in the body fluid compete with each other to bind to the linear epitope of the antigen 10.

[0096] The more the labeled antibody 20 binds to the antigen 10, the more pronounced the measurable and / or user-perceivable change on the carrier medium 2. This change is caused by the local accumulation of the labeled antibody 20, which results from their binding to the antigen 10, and thus the antigen 10 remains at the accumulation location. The accumulation location can be the region of the predetermined location 3. For example, the accumulation has a linear extension, and thus the change occurs along a line, as Figure 1 shown by way of example using the predetermined location 3 shown as a dashed line in

[0097] In contrast, the less the labeled antibody 20 binds to the antigen 10, the weaker the change on the carrier medium 2. This effect is used to illustrate any anti-HPV antibodies contained in the body fluid. This is because the weaker the change on the carrier medium 2 means that there are more antibodies in the body fluid that can bind to the linear epitope of the antigen 10, and these antibodies now bind to the antigen 10 instead of the labeled antibody 20.

[0098] Preferably, the exemplary test kit 1 is adjusted or calibrated by measuring the amount of antigen 10 presented on the carrier medium 2 and / or by measuring the amount of labeled antibody 20 presented on the carrier medium 2, such that if the amount of antibodies in the body fluid that specifically bind to the linear epitope of antigen 10 is below the above-mentioned minimum amount, the measurable and / or user-perceivable change has or exceeds a preferably predetermined maximum intensity. Thus, the exemplary test kit 1 enables a qualitative determination of the anti-HPV antibodies contained in the body fluid, that is, whether such antibodies are contained in the body fluid.

[0099] In view of the fact that as the amount of antibodies in the body fluid that specifically bind to a specific epitope of antigen 10 increases, the measurable and / or user-perceivable change weakens compared to the maximum intensity, the exemplary test kit 1 is also capable of quantitatively determining the anti-HPV antibodies contained in the body fluid. In this case, the degree of weakening of the change compared to the maximum intensity can be used as a measure of the amount of anti-HPV antibodies contained in the body fluid.

[0100] On the one hand, the change that occurs, especially the intensity of its expression, can be perceived by the user, and then, based on the user's perception, a qualitative and / or quantitative identification of the anti-HPV antibodies is made. Additionally or alternatively, a photometer or other device for detecting the severity of the change can be used to qualitatively and / or quantitatively identify the anti-HPV antibodies.

[0101] Figure 2 A second exemplary embodiment of a test kit 1.1 for detecting anti-HPV antibodies is shown. The components or functional parts of the exemplary test kit 1.1 that are the same or functionally equivalent to those of Figure 1 the exemplary test kit 1 are provided with the same reference numerals; in this regard, reference is made to the description of Figure 1 the exemplary test kit 1.

[0102] In this exemplary test kit 1.1, a carrier medium 2' is provided, for example, in the manner of the carrier medium 2 of Figure 1 the exemplary test kit 1, and the labeled antibody 20 is also presented in a mobile manner in this carrier medium 2', for example, in the region of the application position 4.

[0103] Different from Figure 1 the exemplary test kit 1, in Figure 2 the exemplary test kit 1.1, a predetermined amount of capture molecules 30 is provided on the carrier medium 2'. Preferably, a predetermined number of capture molecules 30 are arranged at a predetermined position 3 of the carrier medium 2'. The capture molecules 30 are preferably immobilized, especially fixed at the predetermined position 3. The capture molecules 30 are preferably such that they can each specifically bind to the linear epitope of antigen 10'. For example, the capture molecules 30 are based on the same antibody that forms the basis of the labeled antibody 20.

[0104] Unlike Figure 1 exemplary test kit 1, in exemplary test kit 1.1 Figure 2 a predetermined amount of antigen 10’ is provided, which is present separately from the carrier medium 2’. Similar to antigen 10 of exemplary test kit 1 Figure 1 the antigen 10’ has the linear epitope that enables a plurality or many antibodies to selectively bind thereto specifically, each of the plurality or many antibodies being effective against a different type of HPV. Figure 2 Exemplary test kit 1.1 is based on the fact that antigen 10’ exists in the form of a multimer. Therefore, the linear epitope appears at least twice in the corresponding antigen 10’.

[0105] For example, antigen 10’ is dissolved in a preferably predetermined volume of buffer fluid 40. The buffer fluid 40 is used to keep antigen 10’ in solution and can utilize any capillary properties of the carrier medium 2’. The buffer fluid 40 does not need to have any physiological effect on antigen 10’ because the reactivity of the linear epitope of antigen 10’ can be retained even in a non-physiological buffer fluid. For example, the buffer fluid is a solution based on saline, tris (tris(hydroxymethyl)aminomethane) or histidine. For example, the buffer fluid 40 and antigen 10’ are accommodated in a preferably separate container 5. For example, the container 5 has an opening for filling and / or emptying.

[0106] Exemplary test kit 1.1 can be used as follows: A body fluid (especially a predetermined volume of body fluid) is brought into contact with antigen 10’ (especially the buffer fluid 40 in which antigen 10’ is dissolved). For example, for this purpose, the body fluid is introduced into the container 5. For example, the body fluid is the body fluid described above for Figure 1 exemplary test kit 1.

[0107] Then, preferably after the body fluid and antigen 10’ have interacted for a predetermined period of time, the mixture of the body fluid and antigen 10’, especially the mixture of the body fluid, buffer solution 40 and antigen 10’, or at least a portion of the volume of the mixture, is applied via the application location 4 to the carrier medium 2’. There, the mixture (especially the body fluid) comes into contact with the labeled antibody 20 and, together with at least some of the labeled antibody 20, optionally by utilizing the capillary force caused by the material of the carrier medium 2, flows through the carrier medium 2 to the predetermined location 3 and comes into contact with the capture molecule 30. Due to the contact, the capture body 30 binds to the linear epitopes of antigen 10’ that are still free at that time, especially those not yet occupied by one of the labeled antibody 20 or the antibodies contained in the body fluid.

[0108] In principle, the antigen 10' (especially the buffer fluid in which the antigen 10' is dissolved) and the body fluid separated therefrom can be applied to the carrier medium 2', that is to say, for example, in contact with the labeled antibody 20 at the same time. However, if the antigen 10' first contacts the body fluid and then contacts the labeled antibody, the interaction between the antigen 10' and any antibody contained in the body fluid and the binding to the linear epitope of the antigen 10' are advanced in time. Therefore, these interactions occur before any interaction between the antigen 10' and the labeled antibody 20.

[0109] Preferably, the exemplary test kit 1.1 is adjusted or calibrated by measuring the amount of the labeled antibody 20 presented on the carrier medium 2' and / or measuring the amount of the capture molecule 30 presented on the carrier medium 2', such that in the absence of the participation of the body fluid but in the presence of a fluid (such as a buffer fluid), the antigen 10' contacts the labeled antibody 20, and after the mixture flows to the capture body 30 due to the contact, on the one hand, the labeled antibody 20 specifically binds to the antigen 10', and on the other hand, there is still enough of this antigen other than these bound antigens that still has at least one free linear epitope such that the capture molecule specifically binds thereto. Only by binding to the capture molecule 30 will the labeled antibody 20 attach to the carrier medium 2', and local accumulation of the labeled antibody 20 will occur, resulting in a measurable and / or user-perceivable change on the carrier medium 2'.

[0110] If the antigen 10' is now brought into contact with the body fluid having the labeled antibody 20, then at least one linear epitope of at least some of the antigen 10' is occupied by any antibody contained in the body fluid and capable of binding to the linear epitope of the antigen 10', so that a smaller number of antigen 10' binds to the labeled antibody 20 and the capture molecule 30 at the same time, and thus the measurable and / or user-perceivable change is weaker.

[0111] Preferably, the exemplary test kit 1.1 is adjusted or measured by measuring the amount of the antigen 10' and comparing it with the amount of the labeled antibody 20 and / or the amount of the capture molecule, such that if the amount of the antibody specifically binding to the linear epitope of the antigen 10' contained in the body fluid is lower than the above minimum amount, the measurable and / or user-perceivable change has or exceeds the preferably predetermined maximum intensity. Therefore, the exemplary test kit 1.1 can qualitatively and / or quantitatively determine the anti-HPV antibodies contained in the body fluid in the same manner as Figure 1 the exemplary test kit 1. In this regard, regarding the analysis of the change caused by the accumulation of the labeled antibody, reference is made to Figure 1 the description of the exemplary test kit 1 in

[0112] Figure 3Shows a third exemplary embodiment of a test kit 1.2 for detecting anti-HPV antibodies. As compared with Figure 1 the exemplary test kit 1 or Figure 2 the components or functional parts of the test kit 1.1, the components or functional parts of the exemplary test kit 1.2 that are identical or functionally equivalent are provided with the same reference numerals; in this regard, reference is made to the description of Figure 1 the exemplary test kit 1 or Figure 2 the exemplary test kit 1.1.

[0113] The exemplary test kit 1.2 differs from Figure 2 the exemplary test kit 1.1 in that a predetermined amount of antigen 10” is provided, which exists in monomeric form and, in this regard, each antigen has only one of said linear epitopes. Instead of Figure 2 the antigen 10’ of the exemplary test kit 1.1, the antigen 10” may be dissolved in the buffer fluid 40 and / or accommodated in the container 5.

[0114] The exemplary test kit 1.2 further differs from Figure 2 the exemplary test kit 1.1 in that a carrier medium 2” is used, on which a predetermined amount of capture molecules 50 formed as antibodies are presented such that they can each specifically bind to different epitopes of the antigen 10” other than the said linear epitope. Preferably, the capture molecules 50 are arranged in the region of the predetermined position 3 of the carrier medium 2”’. Preferably, the capture molecules 50 are arranged at the predetermined position 3 of the carrier medium 2”’. The capture molecules 50 are preferably immobilized, in particular immobilized at the predetermined position 3.

[0115] The exemplary test kit 1.2 can be used to perform serological tests in the same manner as Figure 2 the exemplary test kit 1.1. For example, first, a preferably predetermined volume of body fluid is brought into contact with the antigen 10”, in particular the antigen 10” dissolved in the buffer solution 40, and incubated in particular. Then, the body fluid and the antigen 10”, in particular the body fluid, the antigen 10” and the buffer fluid 40, are brought into contact with the labeled antibody 20 located on the carrier medium 2”, and then the mixture flows on the carrier medium 2” to the predetermined position 3 and comes into contact with the capture molecules 50.

[0116] Those antigens 10 that form a binding complex with one of the labeled antibodies 20 will be retained by at least one capture molecule 50, provided that the epitope required for binding on the antigen of the binding complex remains unoccupied and at least one capture molecule binds specifically thereto. Subsequently, the measurable and / or user-perceivable changes in the carrier medium 2” caused by the accumulation of the retained labeled antibody can be analyzed according to the procedures described above for the exemplary test kit 1 or the exemplary test kit 1.1.

[0117] Figure 4 A fourth exemplary embodiment of a test kit 1.3 for detecting anti-HPV antibodies is shown. The exemplary test kit 1.3 includes a carrier medium 2”’, on which a predetermined amount of the above-mentioned antigen 10 is present. Preferably, the carrier medium 2”’ is a reagent container. For example, the reagent container is a component of a microtiter plate. Preferably, the antigen 10 is immobilized on the bottom and / or walls of the reagent container. For example, the bottom and / or walls of the reagent container 10 are coated with the antigen 10.

[0118] In addition, the exemplary test kit 1.3 includes a predetermined amount of labeled antibody 60 and a predetermined volume of reactant 70. Preferably, the labeled antibody 60 is separate from the carrier medium 2”’. Preferably, the reactant 70 is separate from the carrier medium 2”’ and the labeled antibody 60.

[0119] The labeled antibody 60 is based on the antibodies that are already the basis of the labeled antibodies 20 of the exemplary test kits 1, 1.1, and 1.2, and thus enables them to specifically bind to the linear epitopes of the antigen 10. Regarding the label, the labeled antibody 60 causes a measurable and / or user-perceivable reaction in the presence of the reactant. The provided reactant 70 causes this reaction to occur. For example, the labeled antibody 60 is labeled with an enzyme such as horseradish peroxidase (HRP). For example, the reactant is tetramethylbenzidine (TMB).

[0120] A serological test can be performed using the exemplary test kit 1.3 as follows: bringing a preferably predetermined volume of body fluid into contact with the antigen 10 of the carrier medium 2”’, in particular by incubation. For example, for this purpose, the body fluid is poured into the reagent container. The body fluid can be the above-mentioned body fluid.

[0121] Then the carrier medium 2”’ is washed. The antibodies contained in the body fluid and binding to the linear epitopes of the antigen 10 are retained on the carrier medium 2”’, in particular in the reagent container, provided that specific binding to the antigen 10 has occurred and these antibodies are thus retained.

[0122] After washing, the labeled antibody 70 is applied to the carrier medium 2”’, for example filled into a reagent container, in particular for incubation. More or less interaction occurs between the antigen 10 and the labeled antibody 70 depending on how much of the antigen 10 still allows specific binding, since the linear epitope or at least one of the linear epitopes has not been occupied by antibodies contained in the body fluid.

[0123] Then the carrier medium 2”’ is washed again. Those labeled antibodies 70 that are able to specifically bind to the antigen 10 remain on the carrier medium 2”’, in particular in the reagent container. Finally, the reactant 70 is applied to the carrier medium 2”’, in particular filled into the reagent container. Due to the reactant 70, those labeled antibodies 60 that remain on the carrier medium 2”’, in particular in the reagent container, will now show a measurable and / or user-perceivable reaction, such as a color reaction.

[0124] The more labeled antibodies 60 that are able to bind to the antigen 10, the stronger the measurable and / or user-perceivable reaction. In contrast, the fewer labeled antibodies 60 that bind to the antigen 10, the weaker the reaction. This effect is used to illustrate any anti-HPV antibodies contained in the body fluid. This is because the weaker the reaction means the more antibodies are contained in the body fluid that can bind to the linear epitope of the antigen 10, and these antibodies bind to the antigen 10 rather than the labeled antibody.

[0125] For the subsequent analysis of this reaction for the qualitative and / or quantitative determination of any anti-HPV antibodies contained in the body fluid, in principle, it can be carried out in the same way as the serological test based on the measurable and / or user-perceivable changes caused by the accumulation of the labeled antibody 20 described above.

[0126] Figure 5 Shows a fifth exemplary embodiment of the test kit 1.4 for detecting anti-HPV antibodies. The exemplary test kit 1.4 differs from Figure 4 the exemplary test kit 1.3 in that a pre-quantified labeled antibody 80 is provided, which enables it to specifically bind to human antibodies. For example, the labeled antibody 80 does not have the ability to specifically bind to the linear epitope of the antigen 10. The labeled antibody 80 can be based on monoclonal mouse antibodies. The labeled antibody 80 can be labeled in the same way as the labeled antibody 70 of the exemplary test kit 1.3 so that in the presence of the reactant, they will cause a measurable and / or user-perceivable reaction.

[0127] The exemplary test kit 1.4 can be used in the same way as Figure 4The exemplary test kit 1.3 performs serological tests in the same manner; in this regard, reference is made to the above description. Due to the labeled antibody 80, the exemplary test kit 1.4 is based on a sandwich method. This means that the labeled antibody 80 specifically binds to the antibodies contained in the body fluid, which themselves specifically bind to the linear epitopes of the antigen 10 and thus remain on the carrier medium 2”’, especially in the reagent container.

[0128] The more labeled antibodies 80 that bind to those antibodies derived from the body fluid, the stronger the measurable and / or user-perceivable reaction. Therefore, this means that the stronger the reaction, the more antibodies contained in the body fluid that specifically bind to the linear epitopes of the antigen. This effect is used here to illustrate any anti-HPV antibodies contained in the body fluid.

[0129] For the analysis of this reaction for the qualitative and / or quantitative determination of any anti-HPV antibodies contained in the body fluid, in principle, it can be carried out in the same manner as the above-described serological test based on the measurable and / or user-perceivable changes caused by the accumulation of the labeled antibody 20.

[0130] Inspection result

[0131] The performance of the above serological test is described below using the example of the examination of the body fluids of 138 women diagnosed with precancerous lesions or cancers caused by HPV. Whole blood samples are used as the body fluid. The serological test is carried out using a test kit corresponding to the exemplary test kit 1.1 in Figure 2 Among the antigens used, the linear epitopes enable various or many anti-HPV antibodies that can selectively bind to them to be effective against different HPVs among types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, 73, and 82. Thus, the entire high-risk group is covered.

[0132] The results of the serological test are summarized in Table 1. In the column titled "Diagnosis", cervical intraepithelial neoplasia ("CIN") and cervical cancer ("CxCa") are distinguished. Cervical intraepithelial neoplasia is divided into three severities. Severity 1 ("CIN 1") indicates only slight changes in the affected somatic cells, severity 2 ("CIN 2") indicates moderate changes in the affected somatic cells, and severity 3 ("CIN 3") indicates severe changes in the affected somatic cells.

[0133] The "Number" column indicates the number of women diagnosed with cervical intraepithelial neoplasia of grade 1 severity, cervical intraepithelial neoplasia of grade 2 severity, cervical intraepithelial neoplasia of grade 3 severity, or cervical cancer.

[0134] Table 1

[0135] Diagnosis Number Negative Positive Sensitivity CIN 1 21 0 21 100％ CIN 2 18 0 18 100％ CIN 3 82 11 71 86.6％ CxCa 17 3 14 82.4％ Total 138 14 124 89.9％

[0136] During the serological test, a qualitative determination was performed. The threshold for a positive result was set at 750 ng / ml of anti-HPV antibody in the body fluid, and the test kit used was adjusted to this value. Serological test results above 750 ng / ml were considered positive, and results below or equal to 750 ng / ml were considered negative.

[0137] The number of test results classified as positive is shown in the "Positive" column, and the number of test results classified as negative is shown in the "Negative" column. The detection rate is given in the "Sensitivity" column. As can be seen from Table 1, 100% of the women diagnosed with "CIN 1" were detected. 100% of the women diagnosed with "CIN 2" were also identified. A slightly lower detection rate of 86.6% was observed in women diagnosed with "CIN 3". For women diagnosed with cervical cancer, a detection rate of 82.4% could still be achieved. The overall detection rate was 89.9%.

[0138] List of reference numerals

[0139] 1 Test kit

[0140] 1.1 Test kit

[0141] 1.2 Test kit

[0142] 1.3 Test kit

[0143] 1.4 Test kit

[0144] 2 Carrier medium

[0145] 2’ Carrier medium

[0146] 2” Carrier medium

[0147] 2”’ Carrier medium

[0148] 3 Predetermined position

[0149] 4 Application position

[0150] 5 Container

[0151] 10 Antigen

[0152] 10’ Antigen, polymer

[0153] 10” Antigen, monomer

[0154] 20 Labeled antibody

[0155] 30 Capture molecule

[0156] 40 Buffer fluid

[0157] 50 Capture molecule

[0158] 60 Labeled antibody

[0159] 70 Reactant

[0160] 80 Labeled antibody.

Claims

1. A serological method for detecting anti-HPV antibodies, wherein, an antigen having a linear epitope is used, the linear epitope being reactive and enabling multiple or many antibodies to selectively bind specifically thereto, each of the multiple or many antibodies being effective against a different type of HPV.

2. The serological method according to claim 1, wherein, a labeled antibody is used, the labeled antibody enabling it to specifically bind to the linear epitope of the antigen and causing a measurable and / or user-perceivable reaction in the presence of reactants, or a measurable and / or user-perceivable change in the case of spatial accumulation.

3. A serological method for qualitatively and / or quantitatively determining anti-HPV antibodies contained in a body fluid, comprising the following steps or consisting of the following steps : i) Providing a carrier medium (2) on which a predetermined amount of a labeled antibody (20) is presented in a mobile manner, and a predetermined amount of an antigen (10) is present at a predetermined position (3), wherein the antigen (10) has a linear epitope, the linear epitope being reactive and enabling multiple or many antibodies to selectively bind specifically thereto, each of the multiple or many antibodies being effective against a different type of HPV, and wherein the labeled antibody (20) enables it to specifically bind to the linear epitope of the antigen (10) and causes a measurable and / or user-perceivable change in the case of spatial accumulation; ii) Contacting a preferably predetermined volume of the body fluid with the labeled antibody (20); iii) Qualitatively and / or quantitatively identifying the anti-HPV antibodies by analyzing the change in the region of the predetermined position (3) in the carrier medium (2).

4. A serological method for qualitatively and / or quantitatively determining anti-HPV antibodies contained in a body fluid, comprising the following steps or consisting of the following steps : i) Providing a predetermined amount of an antigen (10'), wherein the antigen (10') is polymeric and has a linear epitope, the linear epitope being reactive and enabling multiple or many antibodies to selectively bind specifically thereto, each of the multiple or many antibodies being effective against a different type of HPV; ii) Providing a carrier medium (2') on which a predetermined amount of a labeled antibody (20) is presented in a mobile manner, and a predetermined amount of a capture molecule (30) is present at a predetermined position (3), wherein the labeled antibody (20) and the capture molecule (30) enable them to each specifically bind to the linear epitope of the antigen (10'), and in the case of spatial accumulation, the labeled antibody (20) causes a measurable and / or user-perceivable change; iii) Contacting a preferably predetermined volume of the body fluid with the antigen (10'); iv) Contacting the body fluid and the antigen (10') with the labeled antibody (20); v) Qualitatively and / or quantitatively identifying the anti-HPV antibodies by analyzing the change in the region of the predetermined position (3) in the carrier medium (2').

5. A serological method for the qualitative and / or quantitative determination of anti-HPV antibodies contained in a body fluid, comprising the following steps or consisting of the following steps consisting of: i) providing a predetermined amount of antigen (10”), wherein the antigen (10”) is monomeric and has a linear epitope, the linear epitope being reactive and enabling a plurality or many antibodies to selectively bind thereto specifically, each of the plurality or many antibodies being effective against a different type of HPV; ii) providing a carrier medium (1”), on which a predetermined amount of labeled antibody (20) is presented in a mobile manner, and a predetermined amount of complementary capture molecule (50) is present at a predetermined position (3), wherein the labeled antibody (20) enables it to specifically bind to the linear epitope of the antigen (10”), and in the case of spatial accumulation, the labeled antibody (20) causes a measurable and / or user-perceivable change, wherein the capture molecule (50) enables it to specifically bind to a different binding site of the antigen (10”) other than the linear epitope; iii) contacting a preferably predetermined volume of body fluid with the antigen (10”); iv) contacting the body fluid and the antigen (10”) with the labeled antibody (20); v) qualitatively and / or quantitatively identifying anti-HPV antibodies by analyzing the change in the region of the predetermined position (3) in the carrier medium (2”).

6. A serological method for the qualitative and / or quantitative determination of anti-HPV antibodies contained in a body fluid, comprising the following steps or consisting of the following steps consisting of: i) providing a predetermined amount of antigen (10) on a carrier medium (2”’), wherein the antigen (10) has a linear epitope, the linear epitope being reactive and enabling a plurality or many antibodies to selectively bind thereto specifically, each of the plurality or many antibodies being effective against a different type of HPV; ii) contacting a preferably predetermined volume of body fluid with the antigen (10); iii) contacting a predetermined amount of labeled antibody (60) with the body fluid and the antigen (10), wherein the labeled antibody (60) enables it to specifically bind to the linear epitope of the antigen (10), and causes a measurable and / or user-perceivable reaction in the presence of a reactant (70); iv) washing the carrier medium (2”’); v) applying a predetermined volume of reactant (70) to the carrier medium (2”’), wherein the reactant (70) causes, due to it, the labeled antibody (60) to cause a measurable and / or user-perceivable reaction; vi) qualitatively and / or quantitatively identifying anti-HPV antibodies by analyzing the reaction of the carrier medium (2”’).

7. A serological method for the qualitative and / or quantitative determination of anti-HPV antibodies contained in a body fluid, comprising the following steps or consisting of the following steps consisting of: i) providing a predetermined amount of antigen (10) on a carrier medium (2”’), Wherein, the antigen (10) has a linear epitope, the linear epitope has reactivity and enables a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies is effective against different types of HPV; ii) contacting a preferably predetermined volume of body fluid with the antigen (10); iii) washing the carrier medium (2”’); iv) applying a predetermined amount of labeled antibody (80) to the carrier medium (2”’), wherein the labeled antibody (80) enables it to specifically bind to a human antibody that can specifically bind to the linear epitope of the antigen (10) and causes a measurable and / or user-perceivable reaction in the presence of a reactant (70); v) washing the carrier medium (2”’); vi) applying a predetermined volume of reactant (70) to the carrier medium (2”’), wherein the reactant (70) enables the labeled antibody (80) to cause a measurable and / or user-perceivable reaction due to it; vii) qualitatively and / or quantitatively identifying anti-HPV antibodies by analyzing the reaction of the carrier medium (2”’).

8. The serological method according to one of claims 3 to 7, wherein, the predetermined amount of antigen is less than or corresponds to the predetermined amount of labeled antibody.

9. The serological method according to one of claims 3 to 8, wherein, the predetermined amount of antigen and the predetermined amount of labeled antibody are compared and measured with the volume of body fluid, such that if the amount of antibodies contained in the body fluid and specifically binding to the linear epitope of the antigen is higher than a predetermined minimum amount, then in the presence of a reactant, the labeled antibody causes a measurable and / or user-perceivable reaction, or in the case of spatial accumulation, causes a measurable and / or user-perceivable change.

10. The serological method according to one of the foregoing claims, wherein, the linear epitope of the antigen enables a plurality or many antibodies that can selectively bind thereto to be effective against different types of HPV respectively, and the different types are from a category including at least two types among types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, 73 and 82.

11. The serological method according to one of the foregoing claims, wherein, the linear epitope of the antigen enables a plurality or many antibodies that can selectively bind thereto to be effective against the L1 protein of HPV.

12. A test kit (1; 1.1; 1.2; 1.3; 1.4) for detecting anti-HPV antibodies, particularly for implementing the method according to one of the foregoing claims, comprising an antigen (10; 10’; 10”) having a linear epitope, the linear epitope having reactivity and enabling a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies being effective against different types of HPV.

13. The test kit according to claim 12, particularly for implementing the method according to one of claims 1 to 6, comprising a labeled antibody (20; 60) that is capable of specifically binding to the linear epitope of the antigen (10; 10'; 10") and, in the presence of a reactant (70), causes a measurable and / or user-perceivable reaction, or in the case of spatial accumulation, results in a measurable and / or user-perceivable change.

14. The test kit according to claim 13, particularly for implementing the method according to claim 3, comprising a carrier medium (2) on which a predetermined amount of the labeled antibody (20) is presented in a mobile manner and a predetermined amount of the antigen (10) is present at a predetermined position (3).

15. The test kit according to claim 13, particularly for implementing the method according to claim 4, comprising a predetermined amount of the antigen (10') and a carrier medium (2'), on which a predetermined amount of the labeled antibody (20) is presented in a mobile manner and a predetermined amount of the capture molecule (30) is present at a predetermined position (3). wherein, the antigen (10') is polymeric, and the capture molecule (30) is capable of specifically binding to the linear epitope of the antigen (10') respectively.

16. The test kit according to claim 13, particularly for implementing the method according to claim 5, comprising a predetermined amount of the antigen (10") and a carrier medium (2"), on which a predetermined amount of the labeled antibody (20) is presented in a mobile manner and a predetermined amount of the capture molecule (50) is present at a predetermined position (3). wherein, the antigen (10") is monomeric, and the capture molecule (50) is capable of specifically binding to different binding sites of the antigen (10") other than the linear epitope respectively.

17. The test kit according to claim 13, particularly for implementing the method according to claim 6, comprising a predetermined amount of the antigen (10), preferably on a carrier medium (2"'), a predetermined amount of the labeled antibody (60), and a predetermined volume of the reactant (70), which causes the labeled antibody (60) to cause a measurable and / or user-perceivable reaction due to it.

18. The test kit according to claim 12, particularly for implementing the method according to claim 7, comprising a predetermined amount of the antigen (10), preferably on a carrier medium (2"'), a predetermined amount of the labeled antibody (80) that is capable of specifically binding to a human antibody that can specifically bind to the linear epitope of the antigen (10) and causes a measurable and / or user-perceivable reaction in the presence of a reactant (70), and comprising a predetermined volume of the reactant (70), which causes the reaction to occur due to it.

19. Use of an antigen in the method according to one of claims 1 to 11 and / or in the test kit according to one of claims 12 to 18, wherein the antigen comprises a linear epitope that enables a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies is effective against a different type of HPV.

20. An antibody or a labeled antibody that enables it to specifically bind to the linear epitope of an antigen, wherein the linear epitope enables a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies is effective against a different type of HPV.

21. Use of a labeled antibody in the method according to one of claims 2 to 6 and / or in the test kit according to one of claims 12 to 17, wherein the labeled antibody specifically binds to the linear epitope of an antigen, wherein the linear epitope enables a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies is effective against a different type of HPV.

22. Use of an antibody as a capture molecule in the method according to claim 4 and / or in the test kit according to claim 15, wherein the antibody specifically binds to the linear epitope of an antigen, wherein the linear epitope enables a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies is effective against a different type of HPV.

23. Use of a molecule as a capture molecule in the method according to claim 5 and / or in the test kit according to claim 16, wherein the molecule specifically binds to a different binding site of the antigen other than the linear epitope, wherein the linear epitope enables a plurality or many antibodies to selectively bind thereto specifically, and each of the plurality or many antibodies is effective against a different type of HPV.

24. The use according to claim 23, wherein the molecule is an antibody that enables it to specifically bind to a different epitope of the antigen other than the linear epitope.

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