CN-122017256-A - Kappa light chain detection kit and preparation method thereof

Abstract

The invention discloses a kappa light chain detection kit and a preparation method thereof, which relate to the technical field of clinical in-vitro detection reagents, wherein the kit is in the form of double reagents, the double reagents are a reagent R1 and a reagent R2 respectively, and the reagent R2 comprises a capture antibody and a detection antibody which can specifically bind different epitopes of the kappa light chain, and a sandwich structure of the capture antibody-antigen-detection antibody is formed so as to improve the hook effect in detection. The invention solves the problems that the existing kappa light chain detection reagent uses a single antibody, HOOK effect is easy to occur due to reduction of HOOK points, and a sandwich structure of the capture antibody, the antigen and the detection antibody is formed by two antibodies (capture antibody and detection antibody) aiming at different epitopes of the antigen, and the core is a specific binding mode, so that the recognition accuracy of the antigen can be improved, the non-specific binding is reduced, the detection sensitivity and the HOOK points are greatly improved, and the invention is favorable for further popularization and use in the market.

Inventors

• LIN YUAN
• LIU ZHEN
• LI XUERUI
• HE YUE

Assignees

• 重庆博士泰生物技术有限公司

Dates

Publication Date

20260512

Application Date

20260320

Claims (10)

1. 1. A kappa light chain detection kit, which is characterized in that the kit is in the form of double reagents, wherein the double reagents are a reagent R1 and a reagent R2 respectively, and the reagent R2 comprises a capture antibody and a detection antibody which can specifically bind different epitopes of a kappa light chain, and a sandwich structure of the capture antibody-antigen-detection antibody is formed so as to improve the hook effect in detection.
2. 2. The kappa light chain detection kit as recited in claim 1, wherein the capture antibody is a rabbit anti-human kappa light chain antibody and the detection antibody is a rabbit anti-human kappa free light chain antibody.
3. 3. The kappa light chain detection kit according to claim 1, wherein the concentration of the rabbit anti-human kappa light chain antibody in the reagent R2 is 0.3 to 0.9g/L and the concentration of the rabbit anti-human kappa free light chain antibody is 0.3 to 0.9g/L.
4. 4. The kappa light chain detection kit according to claim 1, wherein the reagent R2 further comprises sodium chloride at a concentration of 2 to 10g/L and sodium azide at a concentration of 0.05 to 0.2%.
5. 5. The kappa light chain detection kit according to claim 4, wherein the reagent R1 comprises 4-hydroxyethyl piperazine ethanesulfonic acid buffer at a concentration of 0.15mol/L, sodium chloride at a concentration of 4.8g/L, polyethylene glycol 20000 at a concentration of 30-50g/L, and Tween 80 at a concentration of 0.1%.
6. 6. A method for preparing the kit according to any one of claims 1 to 5, comprising the steps of preparing reagent R2: providing an antibody diluent; diluting the capture antibody and the detection antibody with the antibody diluent to obtain a solution A and a solution B respectively; The solution A and the solution B are mixed in proportion.
7. 7. The method of preparing a kit of claim 6, wherein the antibody diluent comprises 50mM phosphate buffer at pH 7.4 and 0.1% preservative.
8. 8. A method for detecting kappa light chain content in a sample, using a kit according to any one of claims 1 to 7, comprising the steps of: mixing a sample to be tested with the reagent R1 for pre-incubation; Adding the reagent R2 to start the reaction; the corresponding signal was measured and the concentration calculated.
9. 9. The method of claim 8, wherein the pre-incubation time is 5 minutes and the temperature is 37 ℃.
10. 10. The method of claim 8, wherein the sample is a blood sample or a urine sample.

Description

Kappa light chain detection kit and preparation method thereof Technical Field The invention relates to the technical field of clinical in-vitro detection reagents, in particular to a kappa light chain detection kit and a preparation method thereof. Background Immunoglobulin (Ig) molecular monomers are composed of 2 identical heavy chains (H chains) and 2 identical light chains (L chains) and are linked by disulfide bonds. Wherein the light chain that binds to the heavy chain is referred to as the binding light chain and the light chain that does not bind to the heavy chain is referred to as the free light chain. The sum of the bound light chain and the free light chain is the total light chain. Light chains can be classified into two types, kappa (kappa) and lambda (lambda), depending on their structure and antigenicity, and can be detected in serum and urine under normal conditions. Total light chains can be stably detected in normal human serum, and only a small amount of light chains exist in urine. When kidney injury or multiple myeloma diseases occur, abnormal secretion of plasma cells can cause abnormal content of immunoglobulin and light chain in serum, so that the content of the light chain in serum and urine is obviously different from that of normal people, and the kappa/lambda ratio can also be obviously changed. Currently, the market has more methods for measuring kappa light chains in blood and urine, such as an electroaggregation method, an immunoblotting method, an immunonephelometry method, a radioimmunoassay method, an enzyme-linked immunosorbent assay (ELISA), immunostationary electrophoresis, high performance liquid chromatography, a chemiluminescent immunoassay method and the like. Although ELISA method has the advantages of high sensitivity, high accuracy, rapid reaction, strong specificity and the like, special ELISA is needed, the reagent cost is high, and the immunoturbidimetry method has the advantages of simple operation, high automation degree, high detection speed, high sensitivity and precision, and is suitable for common automatic biochemical analyzers and common spectrophotometers, and the application is wide. Immunoturbidimetry is currently the most common method of total light chain detection in clinic. The anti-kappa light chain antibody is used to react with the antigen in the sample to form an antigen-antibody complex, resulting in increased turbidity of the solution. The light chain content can be calculated by detecting turbidity (change in scattered light intensity or transmitted light intensity). The method needs to add antigen with proper proportion into quantitative antibody solution to make the reaction proceed rapidly, and then form Immune Complex (IC) with the largest quantity, so that the optical signal obtained by detection reaches the strongest, and the result is ensured to be accurate. However, in actual detection, the imbalance of the addition ratio of the antigen in the sample can lead to the failure of the expected detection result. Especially when the antigen is excessive, the immune complex formed by combining the antigen and the antibody is reduced, the deviation of the detection result of the specific protein is directly caused, namely the HOOK effect (HOOK effect) occurs, and the false positive result and the false negative result of the strong positive specimen are easily misdetected, so that the real pathological state is directly covered. Total light chain (Total LIGHT CHAINS, TLC) is the sum of the bound and free light chains, the detection site of the free light chain is the hidden site inside the light chain, while the traditional kappa light chain reagent detection site is the exposed site outside the light chain binding to the heavy chain. Currently available immunoturbidimetry for kappa light chain detection assays use a single antibody, and when the antigen is in excess, a large number of antigens occupy the antibody sites in monovalent binding form, resulting in a reduction of the complex, possibly leading to false positive or false negative events. The hybrid protein in the complex sample may be non-specifically bound to a single antibody, indirectly consuming the amount of antibody, reducing the concentration of antibody that actually can bind to the target antigen, resulting in a reduction in the HOOK spot. In view of this, the present application has been proposed. Disclosure of Invention The invention aims to solve the technical problem that the existing kappa light chain detection reagent mostly uses a single antibody, HOOK effect is easy to occur when HOOK point is reduced, and the invention aims to provide a kappa light chain detection kit and a preparation method thereof. The invention is realized by the following technical scheme: in a first aspect, the invention provides a kappa light chain detection kit in the form of a double reagent, the double reagents being reagent R1 and reagent R2 respectively, the reagent R2 comprising a