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A method and kit for detecting the blood drug concentration of PD-1 antibody

A PD-1, blood drug concentration technology, applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problem of high detection background

Active Publication Date: 2021-04-09
ACROBIOSYSTEMS INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Most of the preclinical and clinical pharmacokinetic studies of the traditional ELISA for the detection of PD-1 antibodies are indirect methods. As we all know, the indirect method has certain disadvantages, that is, the detection background is high, and the serum needs to be diluted 500-1000 times. to solve this problem

Method used

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  • A method and kit for detecting the blood drug concentration of PD-1 antibody
  • A method and kit for detecting the blood drug concentration of PD-1 antibody
  • A method and kit for detecting the blood drug concentration of PD-1 antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] 1. Materials

[0042] 1.1 Drugs and reagents: PD-1, biotinylated anti-PD-1 (biotinylated anti-PD-1 antibody); standard: PD-1 monoclonal antibody, 200mg; coating solution: 1.6g Na 2 CO 3 , 2.9 g NaHCO 3 , 0.5g NaN 3 , add double distilled water to 1L, adjust the pH value to 9.6, filter, subpackage, store at 4°C; PBS: 0.2722g KH2PO4, 3.58g Na2HPO4.12H2O, 8.0063g NaCl, 0.2066g ​​KCl, add double distilled water to 1L, Adjust the pH value to 7.4, filter, aliquot, and store at 4°C; washing solution (PBST): add 0.05% Tween 20 to PBS. Shake well before use; blocking solution: PBST solution containing 2% BSA; sample diluent: PBST solution containing 0.5% BSA; antibody diluent: PBST solution containing 0.5% BSA; antibody: Streptavidin Protein, HRPconjugate (21126) form Thermo, using a ratio of 1:10000; blank serum: purchased from Sunran (Shanghai) Biotechnology Co., Ltd. Wherein, the biotinylated anti-PD-1 is prepared by the following method: the antibody concentration is co...

Embodiment 2

[0095] 1. Establishment of quality inspection standards for raw liquids of each component

[0096] According to the 1.4 ELISA operation steps, the quality inspection of each component is carried out separately. The quality inspection standard of each component is that the experimental data deviation (RSD) of different days is less than 20%, and the stock solution that passes the quality inspection can be used as a component in the kit. The experimental results showed that the deviation of IC50 values ​​in different days was not more than 20% (Table 10).

[0097] Table 10 Deviation of stock solution components on different days

[0098]

[0099] 2. Kit components are freeze-dried

[0100] The freeze-drying process is as follows: the pre-freezing temperature is -60°C for 6 hours, the temperature of the cold trap is -70°C, the vacuum degree is guaranteed to be below 10Pa, the drying time is 30 hours, and the shape of the freeze-dried product is plump.

[0101] 3. Kit quality...

Embodiment 3

[0122] The method in this example is a general method, which can detect various PD-1 antibodies (provided by different manufacturers) under research and on the market. The specific experiment is as follows, according to the 1.4ELISA operation steps, various antibody verifications are carried out, and the results show ( figure 2 ), different antibody drugs may use this method to determine the drug content in serum.

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Abstract

The invention provides a method and a kit for detecting the blood drug concentration of PD-1 antibody. The method comprises using a biotin-labeled PD-1 neutralizing antibody to compete with the PD-1 antibody in the test sample, and using a competition ELISA method to detect the concentration of the PD-1 antibody. Among them, the biotin-labeled PD-1 neutralizing antibody competes with the PD-1 antibody in the serum to bind to the antigen immobilized on the microtiter plate, and horseradish peroxidase-labeled streptavidin is used as the detection antibody to detect biotin Labeled PD-1 neutralizing antibody to quantify PD-1 antibody in serum. The method of the invention has the advantages of good sensitivity, precision, repeated freeze-thaw stability, storage stability at room temperature, specificity and the like, and is beneficial to forming a commercial kit. The kit has the advantages of low background, good precision, batch-to-batch consistency, and stability.

Description

technical field [0001] The present invention relates to a method and a kit for detecting the blood drug concentration of PD-1 antibody, in particular to an enzyme-linked immunosorbent assay (ELISA) for measuring the concentration of PD-1 antibody in serum and the reagents used box. Background technique [0002] In recent years, monoclonal antibody drugs designed based on immune checkpoint proteins such as programmed apoptosis protein-1 (PD-1) and its ligands (PD-L1 / L2) have been widely used as an important immune regulation strategy for tumor suppression. Currently, FDA-approved monoclonal antibody drugs against PD-1 targets include Merck Keytruda (pembrolizumab) and Bristol-Myers Squibb Opdivo (nivolumab), but they have not yet been marketed in my country. In addition, anti-PD-1-related monoclonal antibody drugs are also being applied by multiple domestic pharmaceutical companies, but the current clinical research of PD-1 monoclonal antibody drugs is initiated by multiple ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/558
CPCG01N33/558
Inventor 陈宜顶葛平菊平志光
Owner ACROBIOSYSTEMS INC