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CN-122017230-A - Immunodiagnosis reagent promoter and preparation method thereof

CN122017230ACN 122017230 ACN122017230 ACN 122017230ACN-122017230-A

Abstract

The invention provides an immunodiagnosis reagent accelerator and a preparation method thereof. Comprises an enhancer, a pH regulator, a protective protein, a surfactant, inorganic salts and a bacteriostatic agent. The immunodiagnosis reagent accelerator is a matched enhancement reagent of a common immunodiagnosis reagent, can effectively increase the binding efficiency of antigens and antibodies in the immunodiagnosis reagent, and increases detection signals through the improvement of the binding efficiency, thereby realizing the improvement of detection sensitivity and the reduction of detection time and improving the detection efficiency.

Inventors

  • YAN SHI
  • MENG YAN

Assignees

  • 北京昂领科技有限公司

Dates

Publication Date
20260512
Application Date
20260213

Claims (10)

  1. 1. An immunodiagnosis promoter is characterized by comprising an enhancer, a pH regulator, a protective protein, a surfactant, inorganic salts and a bacteriostatic agent.
  2. 2. An immunodiagnostic accelerator according to claim 1 wherein the enhancer is a high molecular polymer or copolymer of 1-20 carbon chain length, such as propanol, butanol or hexanol, a polyol of 1-2 carbon chain length, such as propylene glycol, hexylene glycol or tetraethylene glycol, polyethylene glycol (PEG, PEO), polypropylene glycol (PPG, PPO), polytetrahydrofuran (PTHF), polyvinylpyrrolidone (PVP), polyvinyl alcohol (PVA), polyacrylic acid (PAA), polymethacrylic acid (PMA) or Linear Polyacrylamide (LPAM), or a mixture of one or more of monosaccharides or polysaccharides such as glucose, fructose, sucrose, dextran or glycogen.
  3. 3. An immunodiagnostic accelerator according to claim 1 wherein the pH modifier is one or more of disodium hydrogen phosphate, sodium dihydrogen phosphate, sodium acetate, sodium citrate, 4-hydroxyethylpiperazine ethanesulfonic acid (HEPES), 2-morpholinoethanesulfonic acid (MES), 3-morpholinopropanesulfonic acid (MPOS), 2- [ [ Tris (hydroxymethyl) methyl ] amino ] ethanesulfonic acid (TES), piperazine-N, N' -bis (2-ethanesulfonic acid) (PIPES), tris (hydroxymethyl) aminomethane (Tris), tris (hydroxymethyl) aminomethane hydrochloride (Tris-HCl), tris (hydroxymethyl) aminopropanesulfonic acid (TAPS), and the like.
  4. 4. An immunodiagnostic accelerator according to claim 1, characterized in that the protective protein is a purified protein, polypeptide, oligopeptide, preferably human serum protein (HSA), bovine Serum Albumin (BSA), ovalbumin (OVA), gelatin, casein and derivatives thereof, oligopeptides, polypeptide, fish serum protein or a mixture of one or more protective proteins, preferably HSA, BSA, OVA, casein and derivatives thereof, fish serum protein, even more preferably HSA, BSA, fish serum protein.
  5. 5. An immunodiagnostic accelerator according to claim 1 wherein the surfactant is one or more of a nonionic surfactant, an anionic surfactant, a cationic surfactant, an amphoteric surfactant, wherein the nonionic surfactant comprises polysorbate, alkylphenol ethoxylates such as tween 20, tween 40, tween 60, tween 80, triton, preferably tween 20 and triton, wherein the anionic surfactant comprises a sulfonate surfactant, preferably Sodium Dodecyl Sulfonate (SDS), wherein the cationic surfactant comprises a quaternary ammonium salt surfactant comprising dodecyltrimethylammonium bromide (DTAB), tetradecyltrimethylammonium bromide, hexadecyltrimethylammonium bromide (CTAB), and the amphoteric surfactant comprises Tetronic 1307 (S9), even more preferably tween 20, tween 40, S9.
  6. 6. An immunodiagnostic accelerator according to claim 1 wherein the inorganic salt is selected from the group consisting of sodium chloride, potassium chloride, magnesium chloride, sodium sulfate, potassium sulfate, sodium citrate, sodium ascorbate, potassium phosphate, sodium phosphate, ammonium sulfate, ammonium chloride, sodium gluconate, and combinations thereof; And/or the bacteriostatic agent may be an inorganic or organic compound having bacteriostatic activity, for example, sodium azide, potassium sorbate, sodium phenylpropionate, isothiazolinone, methyl chloroisothiazolinone, methyl isothiazolinone, proclin-300, etc., and the preferred bacteriostatic agent is sodium azide, methyl chloroisothiazolinone, methyl isothiazolinone, proclin-300, and even more preferred is sodium azide, proclin-300; Preferably, the immunodiagnostic accelerator comprises any one of (1) PB, naCl, BSA, tween 20, butanol, sodium azide, (2) MES, naCl, BSA, tween 20, butanol, proclin 300, (3) MES, naCl, ammonium sulfate, BSA, S9, PEG10000, proclin 300, (4) PB, naCl, ammonium sulfate, BSA, tween 40, PEG20000, proclin 300, (5) Tris, naCl, ammonium sulfate, HSA, tween 40, PEG10000, sodium azide, more preferably, the immunodiagnostic accelerator comprises any one of (1) PB, naCl, BSA, tween 20, butanol, sodium azide, (2) MES, naCl, BSA, tween 20, butanol, proclin 300; Preferably, the content of the enhancer is 0.01-30%, preferably 5-15%, more preferably 10%, the content of the pH regulator is 1-1000mM, preferably 10-200mM, more preferably 50mM, the content of the protective protein is 0.01-20%, preferably 0.5-5%, more preferably 1%, the content of the surfactant is 0.01-10%, preferably 0.05-2%, more preferably 0.1%, the content of the inorganic salt is 50mM-1M, preferably 150mM, and the content of the antibacterial agent is 0.005-5%, preferably 0.05%; preferably, the immunodiagnosis promoter comprises 1-1000mM MES,50mM-1M NaCl, 0.01-20% BSA, 0.01-10% Tween 20, 0.01-30% butanol, and 0.005-5% Proclin with pH of 6.0.
  7. 7. A method of preparing the immunodiagnostic enhancer comprising the step of mixing an enhancer, a pH adjuster, a protective protein, a surfactant, an inorganic salt, and a bacteriostatic agent.
  8. 8. A method of improving the efficiency of an immunodiagnostic reaction comprising the step of adding the immunodiagnostic accelerator of any one of claims 1-6 to an immunodiagnostic reaction system, wherein the immunodiagnostic reaction is selected from the group consisting of an enzyme-linked immune reaction, a chemiluminescent reaction, and an immunochromatographic reaction; preferably, the addition ratio of the immunodiagnosis promoter in the immunodiagnosis reaction system is 10%; Preferably, in the enzyme-linked immunosorbent assay, the immunodiagnosis promoter is added into a 96-well plate, and then a sample, an enzyme conjugate and a substrate are added for incubation and color development; displacing the immunodiagnostic enhancer with a solvent for the magnetic bead component in a chemiluminescent reaction; in an immunochromatographic reaction, adding the immunodiagnosis promoter to a sample buffer to form a mixed diluent; Preferably, the detection signal intensity is at least doubled after the immunodiagnostic enhancer is applied compared to the control group without the enhancer.
  9. 9. Use of the immunodiagnostic accelerator of any one of claims 1-6 in the preparation of a reagent that increases the efficiency of an immunodiagnostic reaction, wherein the immunodiagnostic reaction is selected from the group consisting of an enzyme-linked immune reaction, a chemiluminescent reaction, and an immunochromatographic reaction; Preferably, the improvement in the efficiency of the immunodiagnostic reaction is manifested as an increase in antigen-antibody binding efficiency and/or an increase in detection signal intensity, including OD value, relative luminescence value, or fluorescence intensity.
  10. 10. An immunodiagnostic kit comprising the immunodiagnostic accelerator of any one of claims 1-6 and one or more of an enzyme-linked immunosorbent, chemiluminescent reagent, or immunochromatographic reagent; Preferably, the kit further comprises a sample diluent, a magnetic bead component, an enzyme conjugate, a substrate or a sample buffer, wherein the immunodiagnosis enhancer is used in combination with the sample diluent, the magnetic bead component or the sample buffer.

Description

Immunodiagnosis reagent promoter and preparation method thereof Technical Field The invention relates to the technical field of immunodiagnosis, in particular to an immunodiagnosis reagent accelerator and a preparation method thereof. Background In the current medical in-vitro diagnosis, the immunodiagnosis technology is the technical field with the most abundant project, the most wide application and the largest standard, and specifically is a method for completing capturing, enriching and detecting based on the specific combination among antigen, hapten and antibody, and the main stream detection method based on the immunodiagnosis technology comprises enzyme-linked immunity, chemiluminescence, immunochromatography, turbidimetry, flow fluorescence and the like. Immunodiagnosis technology has long been developed towards improving sensitivity, specificity and detection efficiency, and the key is to improve the binding efficiency and rate of antigen and antibody. The antigen and the antibody are mutually combined by non-bond acting forces such as dispersion force, electrostatic force and hydrogen bond, wherein the dispersion force and the electrostatic force play a role of pulling the two together in the combining process, the dispersion force and the electrostatic force tend to pull the position from the proper antigen and antibody together in early combining stage, the distance between the two is shortened, the relative positions of the antigen and the antibody are aligned, and when the optimal position is reached, the generated hydrogen bond can promote the two to be firmly combined. However, in most cases, the antigen and antibody are proteins, the surfaces of which are provided with a plurality of hydrophilic polar groups, such as-COOH, -NH 2, -OH and the like, which can form hydrogen bond interactions with water molecules, a layer of "fixed water molecules" with a thickness of about 1-1.5: 1.5 nm is formed on the surfaces of the proteins to form hydration layers, and on the other hand, different ions in the buffer aqueous solution can be dispersed on the surfaces of the proteins under the action of electrostatic adsorption. The existence of the hydration layer separates the protein particles in a correlated way and generates electrostatic mutual exclusion acting force, so that the protein is prevented from directly generating coagulation due to collision, the interaction between the antigen and the antibody is also hindered, and the binding efficiency of the antigen and the antibody is reduced. Disclosure of Invention Aiming at the defects existing in the prior art, the invention relates to an accelerator for an immunodiagnosis reagent, in particular to an accelerator which is added into a commercial immunodiagnosis reagent to damage a hydration layer on the surface of protein, effectively enhance the binding efficiency of antigen and antibody in the detection process, improve the performance of the reagent and not damage the stability of the protein. The immunodiagnosis promoter can be used in the common immunodiagnosis fields of enzyme-linked immunity, chemiluminescence, immunochromatography, immune turbidimetry and the like. In one aspect of the invention, an immunodiagnostic enhancer is provided that includes an enhancer, a pH adjustor, a protective protein, a surfactant, an inorganic salt, and a bacteriostat. According to an embodiment of the present invention, the reinforcing agent may be a high molecular polymer or copolymer such as a monohydric alcohol having a carbon chain length of 1 to 20, e.g., propanol, butanol, hexanol, etc., a polyhydric alcohol having a carbon chain length of 1 to 2, e.g., propylene glycol, hexylene glycol, tetraethylene glycol, etc., polyethylene glycol (PEG, PEO) having a polymerization degree of 10 to 200000, polypropylene glycol (PPG, PPO), polytetrahydrofuran (PTHF), polyvinylpyrrolidone (PVP), polyvinyl alcohol (PVA), polyacrylic acid (PAA), polymethacrylic acid (PMA), linear Polyacrylamide (LPAM), etc., or a mixture of one or more of monosaccharides or polysaccharides such as glucose, fructose, sucrose, dextran, glycogen, etc. According to an embodiment of the present invention, the pH adjuster may be one or more of disodium hydrogen phosphate, sodium dihydrogen phosphate, sodium acetate, sodium citrate, 4-hydroxyethyl piperazine ethane sulfonic acid (HEPES), 2-morpholinoethanesulfonic acid (MES), 3-morpholinopropanesulfonic acid (MPOS), 2- [ [ Tris (hydroxymethyl) methyl ] amino ] ethane sulfonic acid (TES), piperazine-N, N' -bis (2-ethane sulfonic acid) (PIPES), tris (hydroxymethyl) aminomethane (Tris), tris (hydroxymethyl) aminomethane hydrochloride (Tris-HCl), tris (hydroxymethyl) methylaminopropane sulfonic acid (TAPS), and the like. According to an embodiment of the present invention, the protecting protein may be purified protein, polypeptide, oligopeptide, protein source may be obtained by processes of purification of natural substances, chemical synthesi