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Procalcitonin detection kit, and preparation method and application thereof

A detection kit and procalcitonin technology, which is applied in the field of immunoassay, can solve problems such as insufficient sensitivity, large operating errors, and too many markers, so as to improve detection sensitivity and detection stability, reduce non-specific binding, and store good stability effect

Pending Publication Date: 2021-06-18
SHANGHAI UPPER BIO TECH PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the manual operation, the operation error is large, the precision deviation is high, the sensitivity and linear range cannot be guaranteed at the same time, and the detection results are easily affected by the external environment.
[0015] At present, although there are kits for detecting procalcitonin on the market, many of them have shortcomings such as not wide linear range, insufficient sensitivity, and high cost.
For example, CN101029897A has too many markers, and the reaction process is more complicated; CN107367620A reagent reaction system is more complicated, and high-throughput testing is difficult to achieve

Method used

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  • Procalcitonin detection kit, and preparation method and application thereof
  • Procalcitonin detection kit, and preparation method and application thereof
  • Procalcitonin detection kit, and preparation method and application thereof

Examples

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preparation example Construction

[0065] The present invention also provides a method for preparing the procalcitonin PCT detection kit according to the , wherein the preparation method includes:

[0066] Coating step of magnetic microspheres: coating the first antibody strain on the magnetic microspheres;

[0067] Labeling step with a tracer marker: labeling the tracer marker on the second antibody strain;

[0068] Prepare calibrator steps.

[0069] As one of the embodiments of the present invention, the coupling time of the first antibody strain and the magnetic microspheres in the step of coating the magnetic beads is 1-1.5 hours.

[0070]As one of the embodiments of the present invention, 2-(N-morpholino)ethanesulfonic acid (MES) buffer is used in the step of coating the magnetic microspheres with the first antibody strain.

[0071] As one of the embodiments of the present invention, a blocking solution is also used in the step of coating the magnetic microspheres with the first antibody strain, and the ...

Embodiment 1

[0086] This embodiment provides a method for preparing a PCT detection kit and a method for detecting procalcitonin in human blood.

[0087] Preparation 1: The first antibody-coated magnetic microspheres

[0088] (1) Measure 10mg of magnetic microspheres (average particle size 1.5μm, purchased from Bangs Laboratories, solid content 2.54%), and suspend in 1mL of 0.1M MES buffer, magnetize for 5-10min, discard After the supernatant, repeat the above cleaning steps 3 to 5 times, add 1 mL of the above buffer (0.1M MES buffer), and vortex to mix.

[0089] (2) Add 200 μg of the first strain of PCT monoclonal antibody (the amino acid position of procalcitonin recognized by the first strain of antibody is 21-40, that is, the sequence is the sequence shown in SEQ ID NO: 4), so that the magnetic The mass ratio of microspheres to antibodies was 50:1, vortexed and incubated at 37°C for 1 hour.

[0090] (3) Add 10 μL of 10 mg / mL 1-(3-dimethylaminopropyl)-3-ethyldiimine hydrochloride (EDC...

Embodiment 2

[0118] Panel screening of antibodies used in PCT detection kits

[0119] There are four kinds of PCT monoclonal antibodies in the process of preparation 1 and preparation 2 in this example: body 1 recognizes amino acid positions 21-40 (the sequence is as shown in SEQ ID NO: 4), and antibody 2 recognizes amino acid positions as 60-69 (the sequence is the sequence shown in SEQ ID NO: 5), the amino acid position recognized by antibody 3 is 72-81 (the sequence is the sequence shown in SEQ ID NO: 6), and the amino acid position recognized by antibody 4 is 96 -105 (the sequence is the sequence shown in SEQ ID NO: 7), and the amino acid positions recognized by antibody 5 are 102-111 (the sequence is the sequence shown in SEQ ID NO: 8). Antibody 1 was coated on magnetic microspheres to form antibody 1-magnetic microsphere suspension, and antibodies 2, 3, 4, and 5 were respectively labeled on acridinium esters to obtain antibody 2-acridinium esters, antibody 3- Acridine esters, Antibo...

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Abstract

The invention relates to a procalcitonin detection kit, and a preparation method and application thereof. The kit provided by the invention comprises a first antibody coated on magnetic microspheres, a second antibody marked with a tracing marker, and a calibrator; the specific binding sites of the procalcitonin identified by the first antibody and the second antibody are different; and the calibrator comprises a procalcitonin antigen A and a diluent, the procalcitonin antigen A has a sequence as shown in SEQ ID NO: 1, and the diluent comprises a buffer solution and a protein stabilizer. The calibration product is high in storage stability, the kit is low in matrix effect, and a detection result can be rapidly obtained.

Description

technical field [0001] The invention belongs to the field of immunoassay, and in particular relates to a procalcitonin detection kit, a preparation method and an application. Background technique [0002] Procalcitonin (PCT) is the precursor of calcitonin (CT), a glycoprotein consisting of 116 amino acid residues and a relative molecular mass of 13,000; it has no hormone activity and is The product encoded by the Calci gene on chromosome 11. Since Assicot first reported that PCT can be used as an early marker of bacterial infection in 1993, it has been used as a new inflammatory indicator and widely used in the diagnosis and differential diagnosis of infectious diseases. It is currently recognized as the most sensitive and effective marker. Specific indicators for the diagnosis of sepsis. [0003] Under normal circumstances, after the human thyroid parafollicular cells (C cells) transcribe and generate CalcimRNA, it is translated into PCT precursor, which is glycosylated a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/74G01N33/543
CPCG01N33/74G01N33/54326G01N2333/585G01N2800/7095G01N2800/52
Inventor 张瑞石晓强邓怡徐建新李福刚
Owner SHANGHAI UPPER BIO TECH PHARMA
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