IL-6 immunoturbidimetry detection kit prepared based on recombinant monoclonal antibody, and preparation method thereof

A technology for detection kits and monoclonal antibodies, applied in the fields of genetic engineering and immunological determination and analysis, can solve the problems of high cost, poor detection specificity, difficulty in popularization and application of chemiluminescence detection and time-resolved fluorescence chromatography, and achieve improved Detection sensitivity, good specificity, and the effect of improving application value

Pending Publication Date: 2021-04-02
ANHUI DAQIAN BIO ENG LIMITED
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, enzyme-linked immunoassay is suitable for scientific research, but the operation is cumbersome, and it is difficult to popularize and apply clinically.
The cost of chemiluminescence detection and time-resolved fluorescence chromatography is high, resulting in improved detection
[0004] Most of the IL-6 immunoturbidimetric detection kits currently used in clinical practice use the method of latex microspheres to label monoclonal antibodies, and the use of polyclonal antibodies has the problem of poor detection specificity

Method used

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  • IL-6 immunoturbidimetry detection kit prepared based on recombinant monoclonal antibody, and preparation method thereof
  • IL-6 immunoturbidimetry detection kit prepared based on recombinant monoclonal antibody, and preparation method thereof
  • IL-6 immunoturbidimetry detection kit prepared based on recombinant monoclonal antibody, and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Preparation of anti-IL-6 latex microsphere mother solution:

[0044] 1. Cleaning: Take latex microspheres with a particle size of 300nm into a 50ml centrifuge tube, add 10 times the volume of 20mMMES activation buffer, shake and mix, and centrifuge at 20000r / min for 30min.

[0045] 2. Activation: Add the same volume of activation buffer, blow and mix the microspheres with a Tip suction head, transfer to a 15ml centrifuge tube, and ultrasonically mix. Ultrasonic parameters are: power 60W, work for 1s, rest for 1s, total time 2min, 3 cycles of ultrasound for backup.

[0046] 3. Combine: Weigh 1.5 times the required amount of EDC, put it into a clean centrifuge tube, and dissolve it in 1ml of pure water. Add EDC to the sonicated microsphere suspension as needed, and shake for 30s. Place in a 37°C water bath and shake for 25 minutes. After activation, transfer the liquid to a 50ml centrifuge tube and centrifuge at 20000r / min for 30min. Add the same volume of binding buffe...

Embodiment 2

[0051] Preparation of IL-6 Immunoturbidimetric Kit Calibrator:

[0052] The IL-6 calibrator contains:

[0053]

Embodiment 3

[0054] Embodiment 3: Preparation of IL-6 immunoturbidimetric assay kit:

[0055] The IL-6 detection kit includes reagent R1 and reagent R2:

[0056] (1) Reagent R1 contains:

[0057]

[0058] (2) Reagent R2 contains:

[0059]

[0060] (3) Assay method of the kit:

[0061] Analysis method: two-point endpoint method;

[0062] Reaction direction: ascending reaction;

[0063] Calibration method: Logit-Log (4P) or SPLINE processing;

[0064] Measurement wavelength: 600nm;

[0065] Measuring temperature: 37°C;

[0066] Sample: Reagent R1: Reagent R2 = 8:200:40 (μL)

[0067] Test steps: Take 8 μL of sample, add 240 μL of reagent R1, incubate at 37°C for 3-5 minutes, add 60 μL of reagent R2, read the absorbance value A1 after 1 minute, read the absorbance value A2 after 5 minutes, and calculate ΔA.

[0068] Calibration method: 6-point calibration, use Hitachi 7180 automatic biochemical analyzer (or other brand models) for detection, set the concentrations of the calibrat...

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Abstract

The invention discloses an IL-6 immunoturbidimetry detection kit prepared based on a recombinant monoclonal antibody. The IL-6 immunoturbidimetry detection kit comprises a reagent R1 and a reagent R2.Two prepared anti-IL-6 antibody latex microsphere mother solutions are used as one component of reagent R2 of the kit, PEG 6000 is added to the reagent R1 according to a certain proportion, the reagent R1 is composed of a buffer solution, a surfactant, a stabilizer and a preservative, and the reagent R2 is composed of a buffer solution, a surfactant, a stabilizer, a preservative and an antibody.The invention also discloses a preparation method of the IL-6 immunoturbidimetry detection kit prepared on the basis of the recombinant monoclonal antibody. Compared with a similar kit for determiningIL-6 by latex enhanced immunotransmission turbidimetry, the kit disclosed by the invention adopts a mode of combining two anti-human IL-6 monoclonal antibodies, can be used for respectively identifying two antigen epitopes, has higher sensitivity than a single monoclonal antibody kit, and has better specificity than a polyclonal antibody kit.

Description

technical field [0001] The invention relates to the technical field of genetic engineering and the field of immunological determination and analysis, in particular to an IL-6 immunoturbidimetric assay kit based on recombinant monoclonal antibody and its preparation method. Background technique [0002] Interleukin-6, referred to as Interleukin 6 (IL-6), is a lymphokine produced by activated T cells and fibroblasts. It can make B cell precursors become antibody-producing cells; cooperate with colony-stimulating factors to promote the growth and differentiation of primitive bone marrow-derived cells, and enhance the lytic function of natural killer cells. IL-6 can regulate the growth and differentiation of various cells, and has the ability to regulate immune response and reverse interleukin-6 in the acute phase. It is a pleiotropic cytokine that can regulate a variety of cellular functions, including cell proliferation, cell differentiation, immune defense mechanisms, and h...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/577G01N33/543
CPCG01N33/6869G01N33/577G01N33/54346G01N2333/5412
Inventor 芮双印符修乐
Owner ANHUI DAQIAN BIO ENG LIMITED
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